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Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_042

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_043

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_044

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_045

Age-dependent biochemical and contractile properties in atrium of transgenic mice overexpressing junctin. Junctin is a transmembrane protein of the cardiac junctional sarcoplasmic reticulum (SR) that binds to the ryanodine receptor, calsequestrin, and triadin 1. This quaternary protein complex is thought to facilitate SR Ca2+ release. To improve our understanding of the contribution of junctin to the regulation of SR function, we examined the age-dependent effects of junctin overexpression in the atrium of 3-, 6-, and 18-wk-old transgenic mice. The ratio of atrial weight and body weight was unchanged between junctin-overexpressing (JCN) and wild-type (WT) mice at all ages investigated (n=6-8). The protein expression of triadin 1 was decreased starting in 3-wk-old JCN atria (by 69%), whereas the expression of the ryanodine receptor was diminished in 6- (by 48%) and 18-wk-old (by 57%) JCN atria compared with age-matched WT atria. Force of contraction was decreased by 35% in 18-wk-old JCN compared with age-matched WT left atrial muscle strips, which was accompanied by a prolonged time of relaxation (48.1 +/- 0.9 vs. 44.2 +/- 0.8 ms, respectively, n=6-8, P <0.05). The spontaneous beating rate of isolated right atria was higher in 18-wk-old JCN mice compared with age-matched WT mice (389 +/- 10 vs. 357 +/- 6 beats/min, respectively, n=6-8, P <0.05). Heart rate was lower by 9% in telemetric ECG recordings in 18-wk-old JCN mice during stress tests. Three-week-old JCN atria exhibited a higher potentiation of force of contraction at rest pauses of 30 s (by 13%) and of 300 s (by 35%), suggesting increased SR Ca2+ content. This was consistent with the higher force of contraction in 3-wk-old JCN atria (by 29%) compared with age-matched WT atria (by 10%) under the administration of caffeine. We conclude that in 3-wk-old atria, junctin overexpression was associated with a reduced expression of triadin 1 resulting in a higher SR Ca2+ load without changes in contractility or heart rate. In 6-wk-old JCN atria, the compensatory downregulation of the ryanodine receptor may offset the effects of junctin overexpression. Finally, the progressive decrease in ryanodine receptor density may contribute to the decreased atrial contractility and lower heart rate during stress in 18-wk-old JCN mice.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_047

Age-dependent biochemical and contractile properties in atrium of transgenic mice overexpressing junctin. Junctin is a transmembrane protein of the cardiac junctional sarcoplasmic reticulum (SR) that binds to the ryanodine receptor, calsequestrin, and triadin 1. This quaternary protein complex is thought to facilitate SR Ca2+ release. To improve our understanding of the contribution of junctin to the regulation of SR function, we examined the age-dependent effects of junctin overexpression in the atrium of 3-, 6-, and 18-wk-old transgenic mice. The ratio of atrial weight and body weight was unchanged between junctin-overexpressing (JCN) and wild-type (WT) mice at all ages investigated (n=6-8). The protein expression of triadin 1 was decreased starting in 3-wk-old JCN atria (by 69%), whereas the expression of the ryanodine receptor was diminished in 6- (by 48%) and 18-wk-old (by 57%) JCN atria compared with age-matched WT atria. Force of contraction was decreased by 35% in 18-wk-old JCN compared with age-matched WT left atrial muscle strips, which was accompanied by a prolonged time of relaxation (48.1 +/- 0.9 vs. 44.2 +/- 0.8 ms, respectively, n=6-8, P <0.05). The spontaneous beating rate of isolated right atria was higher in 18-wk-old JCN mice compared with age-matched WT mice (389 +/- 10 vs. 357 +/- 6 beats/min, respectively, n=6-8, P <0.05). Heart rate was lower by 9% in telemetric ECG recordings in 18-wk-old JCN mice during stress tests. Three-week-old JCN atria exhibited a higher potentiation of force of contraction at rest pauses of 30 s (by 13%) and of 300 s (by 35%), suggesting increased SR Ca2+ content. This was consistent with the higher force of contraction in 3-wk-old JCN atria (by 29%) compared with age-matched WT atria (by 10%) under the administration of caffeine. We conclude that in 3-wk-old atria, junctin overexpression was associated with a reduced expression of triadin 1 resulting in a higher SR Ca2+ load without changes in contractility or heart rate. In 6-wk-old JCN atria, the compensatory downregulation of the ryanodine receptor may offset the effects of junctin overexpression. Finally, the progressive decrease in ryanodine receptor density may contribute to the decreased atrial contractility and lower heart rate during stress in 18-wk-old JCN mice.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_048

Age-dependent biochemical and contractile properties in atrium of transgenic mice overexpressing junctin. Junctin is a transmembrane protein of the cardiac junctional sarcoplasmic reticulum (SR) that binds to the ryanodine receptor, calsequestrin, and triadin 1. This quaternary protein complex is thought to facilitate SR Ca2+ release. To improve our understanding of the contribution of junctin to the regulation of SR function, we examined the age-dependent effects of junctin overexpression in the atrium of 3-, 6-, and 18-wk-old transgenic mice. The ratio of atrial weight and body weight was unchanged between junctin-overexpressing (JCN) and wild-type (WT) mice at all ages investigated (n=6-8). The protein expression of triadin 1 was decreased starting in 3-wk-old JCN atria (by 69%), whereas the expression of the ryanodine receptor was diminished in 6- (by 48%) and 18-wk-old (by 57%) JCN atria compared with age-matched WT atria. Force of contraction was decreased by 35% in 18-wk-old JCN compared with age-matched WT left atrial muscle strips, which was accompanied by a prolonged time of relaxation (48.1 +/- 0.9 vs. 44.2 +/- 0.8 ms, respectively, n=6-8, P <0.05). The spontaneous beating rate of isolated right atria was higher in 18-wk-old JCN mice compared with age-matched WT mice (389 +/- 10 vs. 357 +/- 6 beats/min, respectively, n=6-8, P <0.05). Heart rate was lower by 9% in telemetric ECG recordings in 18-wk-old JCN mice during stress tests. Three-week-old JCN atria exhibited a higher potentiation of force of contraction at rest pauses of 30 s (by 13%) and of 300 s (by 35%), suggesting increased SR Ca2+ content. This was consistent with the higher force of contraction in 3-wk-old JCN atria (by 29%) compared with age-matched WT atria (by 10%) under the administration of caffeine. We conclude that in 3-wk-old atria, junctin overexpression was associated with a reduced expression of triadin 1 resulting in a higher SR Ca2+ load without changes in contractility or heart rate. In 6-wk-old JCN atria, the compensatory downregulation of the ryanodine receptor may offset the effects of junctin overexpression. Finally, the progressive decrease in ryanodine receptor density may contribute to the decreased atrial contractility and lower heart rate during stress in 18-wk-old JCN mice.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_049

Age-dependent biochemical and contractile properties in atrium of transgenic mice overexpressing junctin. Junctin is a transmembrane protein of the cardiac junctional sarcoplasmic reticulum (SR) that binds to the ryanodine receptor, calsequestrin, and triadin 1. This quaternary protein complex is thought to facilitate SR Ca2+ release. To improve our understanding of the contribution of junctin to the regulation of SR function, we examined the age-dependent effects of junctin overexpression in the atrium of 3-, 6-, and 18-wk-old transgenic mice. The ratio of atrial weight and body weight was unchanged between junctin-overexpressing (JCN) and wild-type (WT) mice at all ages investigated (n=6-8). The protein expression of triadin 1 was decreased starting in 3-wk-old JCN atria (by 69%), whereas the expression of the ryanodine receptor was diminished in 6- (by 48%) and 18-wk-old (by 57%) JCN atria compared with age-matched WT atria. Force of contraction was decreased by 35% in 18-wk-old JCN compared with age-matched WT left atrial muscle strips, which was accompanied by a prolonged time of relaxation (48.1 +/- 0.9 vs. 44.2 +/- 0.8 ms, respectively, n=6-8, P <0.05). The spontaneous beating rate of isolated right atria was higher in 18-wk-old JCN mice compared with age-matched WT mice (389 +/- 10 vs. 357 +/- 6 beats/min, respectively, n=6-8, P <0.05). Heart rate was lower by 9% in telemetric ECG recordings in 18-wk-old JCN mice during stress tests. Three-week-old JCN atria exhibited a higher potentiation of force of contraction at rest pauses of 30 s (by 13%) and of 300 s (by 35%), suggesting increased SR Ca2+ content. This was consistent with the higher force of contraction in 3-wk-old JCN atria (by 29%) compared with age-matched WT atria (by 10%) under the administration of caffeine. We conclude that in 3-wk-old atria, junctin overexpression was associated with a reduced expression of triadin 1 resulting in a higher SR Ca2+ load without changes in contractility or heart rate. In 6-wk-old JCN atria, the compensatory downregulation of the ryanodine receptor may offset the effects of junctin overexpression. Finally, the progressive decrease in ryanodine receptor density may contribute to the decreased atrial contractility and lower heart rate during stress in 18-wk-old JCN mice.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_050

Negatively charged amino acids within the intraluminal loop of ryanodine receptor are involved in the interaction with triadin. In mammalian striated muscles, ryanodine receptor (RyR), triadin, junctin, and calsequestrin form a quaternary complex in the lumen of sarcoplasmic reticulum. Such intermolecular interactions contribute not only to the passive buffering of sarcoplasmic reticulum luminal Ca2+, but also to the active Ca2+ release process during excitation-contraction coupling. Here we tested the hypothesis that specific charged amino acids within the luminal portion of RyR mediate its direct interaction with triadin. Using in vitro binding assay and site-directed mutagenesis, we found that the second intraluminal loop of the skeletal muscle RyR1 (amino acids 4860-4917), but not the first intraluminal loop of RyR1 (amino acids 4581-4640) could bind triadin. Specifically, three negatively charged residues Asp4878, Asp4907, and Glu4908 appear to be critical for the association with triadin. Using deletional approaches, we showed that a KEKE motif of triadin (amino acids 200-232) is essential for the binding to RyR1. Because the second intraluminal loop of RyR has been previously shown to contain the ion-conducting pore as well as the selectivity filter of the Ca2+ release channel, and Asp4878, Asp4907, and Glu4908 residues are predicted to locate at the periphery of the pore assembly of the channel, our data suggest that a physical interaction between RyR1 and triadin could play an active role in the overall Ca2+ release process of excitation-contraction coupling in muscle cells.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_053

Negatively charged amino acids within the intraluminal loop of ryanodine receptor are involved in the interaction with triadin. In mammalian striated muscles, ryanodine receptor (RyR), triadin, junctin, and calsequestrin form a quaternary complex in the lumen of sarcoplasmic reticulum. Such intermolecular interactions contribute not only to the passive buffering of sarcoplasmic reticulum luminal Ca2+, but also to the active Ca2+ release process during excitation-contraction coupling. Here we tested the hypothesis that specific charged amino acids within the luminal portion of RyR mediate its direct interaction with triadin. Using in vitro binding assay and site-directed mutagenesis, we found that the second intraluminal loop of the skeletal muscle RyR1 (amino acids 4860-4917), but not the first intraluminal loop of RyR1 (amino acids 4581-4640) could bind triadin. Specifically, three negatively charged residues Asp4878, Asp4907, and Glu4908 appear to be critical for the association with triadin. Using deletional approaches, we showed that a KEKE motif of triadin (amino acids 200-232) is essential for the binding to RyR1. Because the second intraluminal loop of RyR has been previously shown to contain the ion-conducting pore as well as the selectivity filter of the Ca2+ release channel, and Asp4878, Asp4907, and Glu4908 residues are predicted to locate at the periphery of the pore assembly of the channel, our data suggest that a physical interaction between RyR1 and triadin could play an active role in the overall Ca2+ release process of excitation-contraction coupling in muscle cells.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_055

Negatively charged amino acids within the intraluminal loop of ryanodine receptor are involved in the interaction with triadin. In mammalian striated muscles, ryanodine receptor (RyR), triadin, junctin, and calsequestrin form a quaternary complex in the lumen of sarcoplasmic reticulum. Such intermolecular interactions contribute not only to the passive buffering of sarcoplasmic reticulum luminal Ca2+, but also to the active Ca2+ release process during excitation-contraction coupling. Here we tested the hypothesis that specific charged amino acids within the luminal portion of RyR mediate its direct interaction with triadin. Using in vitro binding assay and site-directed mutagenesis, we found that the second intraluminal loop of the skeletal muscle RyR1 (amino acids 4860-4917), but not the first intraluminal loop of RyR1 (amino acids 4581-4640) could bind triadin. Specifically, three negatively charged residues Asp4878, Asp4907, and Glu4908 appear to be critical for the association with triadin. Using deletional approaches, we showed that a KEKE motif of triadin (amino acids 200-232) is essential for the binding to RyR1. Because the second intraluminal loop of RyR has been previously shown to contain the ion-conducting pore as well as the selectivity filter of the Ca2+ release channel, and Asp4878, Asp4907, and Glu4908 residues are predicted to locate at the periphery of the pore assembly of the channel, our data suggest that a physical interaction between RyR1 and triadin could play an active role in the overall Ca2+ release process of excitation-contraction coupling in muscle cells.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_056

Negatively charged amino acids within the intraluminal loop of ryanodine receptor are involved in the interaction with triadin. In mammalian striated muscles, ryanodine receptor (RyR), triadin, junctin, and calsequestrin form a quaternary complex in the lumen of sarcoplasmic reticulum. Such intermolecular interactions contribute not only to the passive buffering of sarcoplasmic reticulum luminal Ca2+, but also to the active Ca2+ release process during excitation-contraction coupling. Here we tested the hypothesis that specific charged amino acids within the luminal portion of RyR mediate its direct interaction with triadin. Using in vitro binding assay and site-directed mutagenesis, we found that the second intraluminal loop of the skeletal muscle RyR1 (amino acids 4860-4917), but not the first intraluminal loop of RyR1 (amino acids 4581-4640) could bind triadin. Specifically, three negatively charged residues Asp4878, Asp4907, and Glu4908 appear to be critical for the association with triadin. Using deletional approaches, we showed that a KEKE motif of triadin (amino acids 200-232) is essential for the binding to RyR1. Because the second intraluminal loop of RyR has been previously shown to contain the ion-conducting pore as well as the selectivity filter of the Ca2+ release channel, and Asp4878, Asp4907, and Glu4908 residues are predicted to locate at the periphery of the pore assembly of the channel, our data suggest that a physical interaction between RyR1 and triadin could play an active role in the overall Ca2+ release process of excitation-contraction coupling in muscle cells.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_057

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_059

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_060

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_061

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_062

Cardiac hypertrophy and impaired relaxation in transgenic mice overexpressing triadin 1. Triadin 1 is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum (SR), which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), junctin, and calsequestrin. To better understand the role of triadin 1 in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of triadin 1 to mouse atrium and ventricle, employing the alpha-myosin heavy chain promoter to drive protein expression. The protein was overexpressed 5-fold in mouse ventricles, and overexpression was accompanied by cardiac hypertrophy. The levels of two other junctional SR proteins, the ryanodine receptor and junctin, were reduced by 55% and 73%, respectively, in association with triadin 1 overexpression, whereas the levels of calsequestrin, the Ca(2+)-binding protein of junctional SR, and of phospholamban and SERCA2a, Ca(2+)-handling proteins of the free SR, were unchanged. Cardiac myocytes from triadin 1-overexpressing mice exhibited depressed contractility; Ca(2+) transients decayed at a slower rate, and cell shortening and relengthening were diminished. The extent of depression of cell shortening of triadin 1-overexpressing cardiomyocytes was rate-dependent, being more depressed under low stimulation frequencies (0.5 Hz), but reaching comparable levels at higher frequencies of stimulation (5 Hz). Spontaneously beating, isolated work-performing heart preparations overexpressing triadin 1 also relaxed at a slower rate than control hearts, and failed to adapt to increased afterload appropriately. The fast time inactivation constant, tau(1), of the l-type Ca(2+) channel was prolonged in transgenic cardiomyocytes. Our results provide evidence for the coordinated regulation of junctional SR protein expression in heart independent of free SR protein expression, and furthermore suggest an important role for triadin 1 in regulating the contractile properties of the heart during excitation-contraction coupling.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_065

Cardiac hypertrophy and impaired relaxation in transgenic mice overexpressing triadin 1. Triadin 1 is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum (SR), which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), junctin, and calsequestrin. To better understand the role of triadin 1 in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of triadin 1 to mouse atrium and ventricle, employing the alpha-myosin heavy chain promoter to drive protein expression. The protein was overexpressed 5-fold in mouse ventricles, and overexpression was accompanied by cardiac hypertrophy. The levels of two other junctional SR proteins, the ryanodine receptor and junctin, were reduced by 55% and 73%, respectively, in association with triadin 1 overexpression, whereas the levels of calsequestrin, the Ca(2+)-binding protein of junctional SR, and of phospholamban and SERCA2a, Ca(2+)-handling proteins of the free SR, were unchanged. Cardiac myocytes from triadin 1-overexpressing mice exhibited depressed contractility; Ca(2+) transients decayed at a slower rate, and cell shortening and relengthening were diminished. The extent of depression of cell shortening of triadin 1-overexpressing cardiomyocytes was rate-dependent, being more depressed under low stimulation frequencies (0.5 Hz), but reaching comparable levels at higher frequencies of stimulation (5 Hz). Spontaneously beating, isolated work-performing heart preparations overexpressing triadin 1 also relaxed at a slower rate than control hearts, and failed to adapt to increased afterload appropriately. The fast time inactivation constant, tau(1), of the l-type Ca(2+) channel was prolonged in transgenic cardiomyocytes. Our results provide evidence for the coordinated regulation of junctional SR protein expression in heart independent of free SR protein expression, and furthermore suggest an important role for triadin 1 in regulating the contractile properties of the heart during excitation-contraction coupling.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_066

Cardiac hypertrophy and impaired relaxation in transgenic mice overexpressing triadin 1. Triadin 1 is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum (SR), which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), junctin, and calsequestrin. To better understand the role of triadin 1 in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of triadin 1 to mouse atrium and ventricle, employing the alpha-myosin heavy chain promoter to drive protein expression. The protein was overexpressed 5-fold in mouse ventricles, and overexpression was accompanied by cardiac hypertrophy. The levels of two other junctional SR proteins, the ryanodine receptor and junctin, were reduced by 55% and 73%, respectively, in association with triadin 1 overexpression, whereas the levels of calsequestrin, the Ca(2+)-binding protein of junctional SR, and of phospholamban and SERCA2a, Ca(2+)-handling proteins of the free SR, were unchanged. Cardiac myocytes from triadin 1-overexpressing mice exhibited depressed contractility; Ca(2+) transients decayed at a slower rate, and cell shortening and relengthening were diminished. The extent of depression of cell shortening of triadin 1-overexpressing cardiomyocytes was rate-dependent, being more depressed under low stimulation frequencies (0.5 Hz), but reaching comparable levels at higher frequencies of stimulation (5 Hz). Spontaneously beating, isolated work-performing heart preparations overexpressing triadin 1 also relaxed at a slower rate than control hearts, and failed to adapt to increased afterload appropriately. The fast time inactivation constant, tau(1), of the l-type Ca(2+) channel was prolonged in transgenic cardiomyocytes. Our results provide evidence for the coordinated regulation of junctional SR protein expression in heart independent of free SR protein expression, and furthermore suggest an important role for triadin 1 in regulating the contractile properties of the heart during excitation-contraction coupling.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_067

Cardiac hypertrophy and impaired relaxation in transgenic mice overexpressing triadin 1. Triadin 1 is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum (SR), which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), junctin, and calsequestrin. To better understand the role of triadin 1 in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of triadin 1 to mouse atrium and ventricle, employing the alpha-myosin heavy chain promoter to drive protein expression. The protein was overexpressed 5-fold in mouse ventricles, and overexpression was accompanied by cardiac hypertrophy. The levels of two other junctional SR proteins, the ryanodine receptor and junctin, were reduced by 55% and 73%, respectively, in association with triadin 1 overexpression, whereas the levels of calsequestrin, the Ca(2+)-binding protein of junctional SR, and of phospholamban and SERCA2a, Ca(2+)-handling proteins of the free SR, were unchanged. Cardiac myocytes from triadin 1-overexpressing mice exhibited depressed contractility; Ca(2+) transients decayed at a slower rate, and cell shortening and relengthening were diminished. The extent of depression of cell shortening of triadin 1-overexpressing cardiomyocytes was rate-dependent, being more depressed under low stimulation frequencies (0.5 Hz), but reaching comparable levels at higher frequencies of stimulation (5 Hz). Spontaneously beating, isolated work-performing heart preparations overexpressing triadin 1 also relaxed at a slower rate than control hearts, and failed to adapt to increased afterload appropriately. The fast time inactivation constant, tau(1), of the l-type Ca(2+) channel was prolonged in transgenic cardiomyocytes. Our results provide evidence for the coordinated regulation of junctional SR protein expression in heart independent of free SR protein expression, and furthermore suggest an important role for triadin 1 in regulating the contractile properties of the heart during excitation-contraction coupling.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_068

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_071

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_072

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_073

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_074

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_075

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_076

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_080

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_081

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_082

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_083

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_084

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_086

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_087

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_088

Complex formation between junctin, triadin, calsequestrin, and the ryanodine receptor. Proteins of the cardiac junctional sarcoplasmic reticulum membrane. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995) J. Biol. Chem. 270, 30787-30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into "KEKE" motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027-9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_089

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_091

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_092

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_093

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_094

Dual role of junctin in the regulation of ryanodine receptors and calcium release in cardiac ventricular myocytes. Junctin, a 26 kDa intra-sarcoplasmic reticulum (SR) protein, forms a quaternary complex with triadin, calsequestrin and the ryanodine receptor (RyR) at the junctional SR membrane. The physiological role for junctin in the luminal regulation of RyR Ca(2+) release remains unresolved, but it appears to be essential for proper cardiac function since ablation of junctin results in increased ventricular automaticity. Given that the junctin levels are severely reduced in human failing hearts, we performed an in-depth study of the mechanisms affecting intracellular Ca(2+) homeostasis in junctin-deficient cardiomyocytes. In concurrence with sparks, JCN-KO cardiomyocytes display increased Ca(2+) transient amplitude, resulting from increased SR [Ca(2+)] ([Ca(2+)](SR)). Junctin ablation appears to affect how RyRs 'sense' SR Ca(2+) load, resulting in decreased diastolic SR Ca(2+) leak despite an elevated [Ca(2+)](SR). Surprisingly, the β-adrenergic enhancement of [Ca(2+)](SR) reverses the decrease in RyR activity and leads to spontaneous Ca(2+) release, evidenced by the development of spontaneous aftercontractions. Single channel recordings of RyRs from WT and JCN-KO cardiac SR indicate that the absence of junctin produces a dual effect on the normally linear response of RyRs to luminal [Ca(2+)]: at low luminal [Ca(2+)] (<1 mmol l(-1)), junctin-devoid RyR channels are less responsive to luminal [Ca(2+)]; conversely, high luminal [Ca(2+)] turns them hypersensitive to this form of channel modulation. Thus, junctin produces complex effects on Ca(2+) sparks, transients, and leak, but the luminal [Ca(2+)]-dependent dual response of junctin-devoid RyRs demonstrates that junctin normally acts as an activator of RyR channels at low luminal [Ca(2+)], and as an inhibitor at high luminal [Ca(2+)]. Because the crossover occurs at a [Ca(2+)](SR) that is close to that present in resting cells, it is possible that the activator-inhibitor role of junctin may be exerted under periods of prevalent parasympathetic and sympathetic activity, respectively.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_096

Dual role of junctin in the regulation of ryanodine receptors and calcium release in cardiac ventricular myocytes. Junctin, a 26 kDa intra-sarcoplasmic reticulum (SR) protein, forms a quaternary complex with triadin, calsequestrin and the ryanodine receptor (RyR) at the junctional SR membrane. The physiological role for junctin in the luminal regulation of RyR Ca(2+) release remains unresolved, but it appears to be essential for proper cardiac function since ablation of junctin results in increased ventricular automaticity. Given that the junctin levels are severely reduced in human failing hearts, we performed an in-depth study of the mechanisms affecting intracellular Ca(2+) homeostasis in junctin-deficient cardiomyocytes. In concurrence with sparks, JCN-KO cardiomyocytes display increased Ca(2+) transient amplitude, resulting from increased SR [Ca(2+)] ([Ca(2+)](SR)). Junctin ablation appears to affect how RyRs 'sense' SR Ca(2+) load, resulting in decreased diastolic SR Ca(2+) leak despite an elevated [Ca(2+)](SR). Surprisingly, the β-adrenergic enhancement of [Ca(2+)](SR) reverses the decrease in RyR activity and leads to spontaneous Ca(2+) release, evidenced by the development of spontaneous aftercontractions. Single channel recordings of RyRs from WT and JCN-KO cardiac SR indicate that the absence of junctin produces a dual effect on the normally linear response of RyRs to luminal [Ca(2+)]: at low luminal [Ca(2+)] (<1 mmol l(-1)), junctin-devoid RyR channels are less responsive to luminal [Ca(2+)]; conversely, high luminal [Ca(2+)] turns them hypersensitive to this form of channel modulation. Thus, junctin produces complex effects on Ca(2+) sparks, transients, and leak, but the luminal [Ca(2+)]-dependent dual response of junctin-devoid RyRs demonstrates that junctin normally acts as an activator of RyR channels at low luminal [Ca(2+)], and as an inhibitor at high luminal [Ca(2+)]. Because the crossover occurs at a [Ca(2+)](SR) that is close to that present in resting cells, it is possible that the activator-inhibitor role of junctin may be exerted under periods of prevalent parasympathetic and sympathetic activity, respectively.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_098

Dual role of junctin in the regulation of ryanodine receptors and calcium release in cardiac ventricular myocytes. Junctin, a 26 kDa intra-sarcoplasmic reticulum (SR) protein, forms a quaternary complex with triadin, calsequestrin and the ryanodine receptor (RyR) at the junctional SR membrane. The physiological role for junctin in the luminal regulation of RyR Ca(2+) release remains unresolved, but it appears to be essential for proper cardiac function since ablation of junctin results in increased ventricular automaticity. Given that the junctin levels are severely reduced in human failing hearts, we performed an in-depth study of the mechanisms affecting intracellular Ca(2+) homeostasis in junctin-deficient cardiomyocytes. In concurrence with sparks, JCN-KO cardiomyocytes display increased Ca(2+) transient amplitude, resulting from increased SR [Ca(2+)] ([Ca(2+)](SR)). Junctin ablation appears to affect how RyRs 'sense' SR Ca(2+) load, resulting in decreased diastolic SR Ca(2+) leak despite an elevated [Ca(2+)](SR). Surprisingly, the β-adrenergic enhancement of [Ca(2+)](SR) reverses the decrease in RyR activity and leads to spontaneous Ca(2+) release, evidenced by the development of spontaneous aftercontractions. Single channel recordings of RyRs from WT and JCN-KO cardiac SR indicate that the absence of junctin produces a dual effect on the normally linear response of RyRs to luminal [Ca(2+)]: at low luminal [Ca(2+)] (<1 mmol l(-1)), junctin-devoid RyR channels are less responsive to luminal [Ca(2+)]; conversely, high luminal [Ca(2+)] turns them hypersensitive to this form of channel modulation. Thus, junctin produces complex effects on Ca(2+) sparks, transients, and leak, but the luminal [Ca(2+)]-dependent dual response of junctin-devoid RyRs demonstrates that junctin normally acts as an activator of RyR channels at low luminal [Ca(2+)], and as an inhibitor at high luminal [Ca(2+)]. Because the crossover occurs at a [Ca(2+)](SR) that is close to that present in resting cells, it is possible that the activator-inhibitor role of junctin may be exerted under periods of prevalent parasympathetic and sympathetic activity, respectively.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_099

Dual role of junctin in the regulation of ryanodine receptors and calcium release in cardiac ventricular myocytes. Junctin, a 26 kDa intra-sarcoplasmic reticulum (SR) protein, forms a quaternary complex with triadin, calsequestrin and the ryanodine receptor (RyR) at the junctional SR membrane. The physiological role for junctin in the luminal regulation of RyR Ca(2+) release remains unresolved, but it appears to be essential for proper cardiac function since ablation of junctin results in increased ventricular automaticity. Given that the junctin levels are severely reduced in human failing hearts, we performed an in-depth study of the mechanisms affecting intracellular Ca(2+) homeostasis in junctin-deficient cardiomyocytes. In concurrence with sparks, JCN-KO cardiomyocytes display increased Ca(2+) transient amplitude, resulting from increased SR [Ca(2+)] ([Ca(2+)](SR)). Junctin ablation appears to affect how RyRs 'sense' SR Ca(2+) load, resulting in decreased diastolic SR Ca(2+) leak despite an elevated [Ca(2+)](SR). Surprisingly, the β-adrenergic enhancement of [Ca(2+)](SR) reverses the decrease in RyR activity and leads to spontaneous Ca(2+) release, evidenced by the development of spontaneous aftercontractions. Single channel recordings of RyRs from WT and JCN-KO cardiac SR indicate that the absence of junctin produces a dual effect on the normally linear response of RyRs to luminal [Ca(2+)]: at low luminal [Ca(2+)] (<1 mmol l(-1)), junctin-devoid RyR channels are less responsive to luminal [Ca(2+)]; conversely, high luminal [Ca(2+)] turns them hypersensitive to this form of channel modulation. Thus, junctin produces complex effects on Ca(2+) sparks, transients, and leak, but the luminal [Ca(2+)]-dependent dual response of junctin-devoid RyRs demonstrates that junctin normally acts as an activator of RyR channels at low luminal [Ca(2+)], and as an inhibitor at high luminal [Ca(2+)]. Because the crossover occurs at a [Ca(2+)](SR) that is close to that present in resting cells, it is possible that the activator-inhibitor role of junctin may be exerted under periods of prevalent parasympathetic and sympathetic activity, respectively.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_100

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_103

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_104

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_105

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_106

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_109

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_110

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_111

Regulatory roles of junctin in sarcoplasmic reticulum calcium cycling and myocardial function. Junctin (JCN), a 26-kd sarcoplasmic reticulum (SR) transmembrane protein, forms a quaternary protein complex with the ryanodine receptor, calsequestrin, and triadin in the SR lumen of cardiac muscle. Within this complex, calsequestrin, triadin, and JCN appear to be critical for normal regulation of ryanodine receptor-mediated calcium (Ca) release. Junctin and triadin exhibit 60% to 70% amino acid homology in their transmembrane domains, including repeated KEKE motifs important for macromolecular protein-protein interactions within their SR luminal tails. Recent studies have uncovered functional roles of both JCN and triadin in the mouse heart, using transgenic overexpression strategies, which exhibit varying phenotypes including mild SR structural alterations, prolongation of Ca transient decay, impaired relaxation, and cardiac hypertrophy and/or heart failure. More specifically, both in vitro adenoviral gene transfer and in vivo gene-targeting techniques to manipulate JCN expression levels have shown that JCN is an essential factor in maintaining normal cardiac Ca handling and cardiac function. This article reviews the new findings on the regulatory roles of JCN in cardiac SR Ca cycling and contractility, with special emphasis on the effects of JCN ablation on delayed after depolarization-induced arrhythmias and premature mortality in mouse models.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_112

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_115

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_116

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_117

Regulation of ryanodine receptors by calsequestrin: effect of high luminal Ca2+ and phosphorylation. Calsequestrin, the major calcium sequestering protein in the sarcoplasmic reticulum of muscle, forms a quaternary complex with the ryanodine receptor calcium release channel and the intrinsic membrane proteins triadin and junctin. We have investigated the possibility that calsequestrin is a luminal calcium concentration sensor for the ryanodine receptor. We measured the luminal calcium concentration at which calsequestrin dissociates from the ryanodine receptor and the effect of calsequestrin on the response of the ryanodine receptor to changes in luminal calcium. We provide electrophysiological and biochemical evidence that: 1), luminal calcium concentration of >/=4 mM dissociates calsequestrin from junctional face membrane, whereas in the range of 1-3 mM calsequestrin remains attached; 2), the association with calsequestrin inhibits ryanodine receptor activity, but amplifies its response to changes in luminal calcium concentration; and 3), under physiological calcium conditions (1 mM), phosphorylation of calsequestrin does not alter its ability to inhibit native ryanodine receptor activity when the anchoring proteins triadin and junctin are present. These data suggest that the quaternary complex is intact in vivo, and provides further evidence that calsequestrin is involved in the sarcoplasmic reticulum calcium signaling pathway and has a role as a luminal calcium sensor for the ryanodine receptor.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_118

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_120

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_121

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_122

Impaired relaxation in transgenic mice overexpressing junctin. OBJECTIVE: Junctin is a major transmembrane protein in cardiac junctional sarcoplasmic reticulum, which forms a quaternary complex with the ryanodine receptor (Ca(2+) release channel), triadin, and calsequestrin. METHODS: To better understand the role of junctin in excitation-contraction coupling in the heart, we generated transgenic mice with targeted overexpression of junctin to mouse heart, using the alpha-MHC promoter to drive protein expression. RESULTS: The protein was overexpressed 10-fold in mouse ventricles and overexpression was accompanied by cardiac hypertrophy (19%). The levels of two other junctional SR-proteins, the ryanodine receptor and triadin, were reduced by 32% and 23%, respectively. However, [3H]ryanodine binding and the expression levels of calsequestrin, phospholamban and SERCA2a remained unchanged. Cardiomyocytes from junctin-overexpressing mice exhibited impaired relaxation: Ca(2+) transients decayed at a slower rate and cell relengthening was prolonged. Isolated electrically stimulated papillary muscles from junctin-overexpressing hearts exhibited prolonged mechanical relaxation, and echocardiographic parameters of relaxation were prolonged in the living transgenic mice. The amplitude of caffeine-induced Ca(2+) transients was lower in cardiomyocytes from junctin-overexpressing mice. The inactivation kinetics of L-type Ca(2+) channel were prolonged in junctin-overexpressing cardiomyocytes using Ca(2+) or Ba(2+) as charge carriers. CONCLUSION: Our data provide evidence that cardiac-specific overexpression of junctin is accompanied by impaired myocardial relaxation with prolonged Ca(2+) transient kinetics on the cardiomyocyte level.

Which proteins participate in the formation of the ryanodine receptor quaternary macromolecular complex?

54f9d3eedd3fc62544000004_123

Autism and esophageal achalasia in childhood: a possible correlation? Report on three cases. Chronic gastrointestinal symptoms are commonly reported in autistic patients. Dysphagia is often present, and it is generally related to behavioral eating disorders. The association between autism and esophageal achalasia has not been described in literature yet. We report our experience with three cases of autistic children we recently treated for esophageal achalasia. In the first case (a 14-year-old male), achalasia was diagnosed with barium swallow and esophageal manometry and was successfully treated with three pneumatic endoscopic dilatations (follow-up: 3 years). In the second case (a 12-year-old female), achalasia was diagnosed with barium swallow and esophageal manometry and was treated with Heller myotomy after two unsuccessful pneumatic endoscopic attempts (follow-up: 3 months). In the last case, a 15-year-old male underwent barium swallow and endoscopy that confirmed achalasia. He was treated with Heller myotomy, and he is asymptomatic at a 6-month follow-up. To our knowledge, this is the first report of a possible association between autism and esophageal achalasia. Because of the rarity of both diseases, their association in the same patient is unlikely to be casual even if speculation on their common etiology is impossible at present. This finding needs further confirmation, but it is sufficient, in our opinion, to indicate proper evaluation with barium swallow and/or manometry in any autistic children with eating difficulty.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_001

Autism and esophageal achalasia in childhood: a possible correlation? Report on three cases. Chronic gastrointestinal symptoms are commonly reported in autistic patients. Dysphagia is often present, and it is generally related to behavioral eating disorders. The association between autism and esophageal achalasia has not been described in literature yet. We report our experience with three cases of autistic children we recently treated for esophageal achalasia. In the first case (a 14-year-old male), achalasia was diagnosed with barium swallow and esophageal manometry and was successfully treated with three pneumatic endoscopic dilatations (follow-up: 3 years). In the second case (a 12-year-old female), achalasia was diagnosed with barium swallow and esophageal manometry and was treated with Heller myotomy after two unsuccessful pneumatic endoscopic attempts (follow-up: 3 months). In the last case, a 15-year-old male underwent barium swallow and endoscopy that confirmed achalasia. He was treated with Heller myotomy, and he is asymptomatic at a 6-month follow-up. To our knowledge, this is the first report of a possible association between autism and esophageal achalasia. Because of the rarity of both diseases, their association in the same patient is unlikely to be casual even if speculation on their common etiology is impossible at present. This finding needs further confirmation, but it is sufficient, in our opinion, to indicate proper evaluation with barium swallow and/or manometry in any autistic children with eating difficulty.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_002

The significance of ileo-colonic lymphoid nodular hyperplasia in children with autistic spectrum disorder. BACKGROUND: Intestinal mucosal pathology, characterized by ileo-colonic lymphoid nodular hyperplasia (LNH) and mild acute and chronic inflammation of the colorectum, small bowel and stomach, has been reported in children with autistic spectrum disorder (ASD). AIM: To assess ileo-colonic LNH in ASD and control children and to test the hypothesis that there is an association between ileo-colonic LNH and ASD in children. PATIENTS AND METHODS: One hundred and forty-eight consecutive children with ASD (median age 6 years; range 2-16; 127 male) with gastrointestinal symptoms were investigated by ileo-colonoscopy. Macroscopic and histological features were scored and compared with 30 developmentally normal (non-inflammatory bowel disease, non-coeliac disease) controls (median age 7 years; range 1-11; 25 male) showing mild non-specific colitis in 16 cases (13 male) and normal colonic histology in 14 cases (12 male). Seventy-four ASD children and 23 controls also underwent upper gastrointestinal endoscopy. The influence on ileal LNH of dietary restriction, age at colonoscopy, and co-existent LNH elsewhere in the intestine, was examined. RESULTS: The prevalence of LNH was significantly greater in ASD children compared with controls in the ileum (129/144 (90%) vs. 8/27 (30%), P < 0.0001) and colon (88/148 (59%) vs. 7/30 (23%), P = 0.0003), whether or not controls had co-existent colonic inflammation. The severity of ileal LNH was significantly greater in ASD children compared with controls, with moderate to severe ileal LNH present in 98 of 144 (68%) ASD children versus 4 of 27 (15%) controls (P < 0.0001). Severe ileal LNH was associated with co-existent colonic LNH in ASD children (P = 0.01). The presence and severity of ileal LNH was not influenced by either diet or age at colonoscopy (P = 0.2). Isolated ileal LNH without evidence of pathology elsewhere in the intestine was a rare event, occurring in less than 3% of children overall. On histopathological examination, hyperplastic lymphoid follicles are significantly more prevalent in the ileum of ASD children (84/138; 61%) compared with controls (2/23; 9%, P = 0.0001). CONCLUSION: Ileo-colonic LNH is a characteristic pathological finding in children with ASD and gastrointestinal symptoms, and is associated with mucosal inflammation. Differences in age at colonoscopy and diet do not account for these changes. The data support the hypothesis that LNH is a significant pathological finding in ASD children.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_003

Gastrointestinal abnormalities in children with autistic disorder. OBJECTIVES: Our aim was to evaluate the structure and function of the upper gastrointestinal tract in a group of patients with autism who had gastrointestinal symptoms. STUDY DESIGN: Thirty-six children (age: 5.7 +/- 2 years, mean +/- SD) with autistic disorder underwent upper gastrointestinal endoscopy with biopsies, intestinal and pancreatic enzyme analyses, and bacterial and fungal cultures. The most frequent gastrointestinal complaints were chronic diarrhea, gaseousness, and abdominal discomfort and distension. RESULTS: Histologic examination in these 36 children revealed grade I or II reflux esophagitis in 25 (69.4%), chronic gastritis in 15, and chronic duodenitis in 24. The number of Paneth's cells in the duodenal crypts was significantly elevated in autistic children compared with non-autistic control subjects. Low intestinal carbohydrate digestive enzyme activity was reported in 21 children (58.3%), although there was no abnormality found in pancreatic function. Seventy-five percent of the autistic children (27/36) had an increased pancreatico-biliary fluid output after intravenous secretin administration. Nineteen of the 21 patients with diarrhea had significantly higher fluid output than those without diarrhea. CONCLUSIONS: Unrecognized gastrointestinal disorders, especially reflux esophagitis and disaccharide malabsorption, may contribute to the behavioral problems of the non-verbal autistic patients. The observed increase in pancreatico-biliary secretion after secretin infusion suggests an upregulation of secretin receptors in the pancreas and liver. Further studies are required to determine the possible association between the brain and gastrointestinal dysfunctions in children with autistic disorder.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_004

Gastrointestinal abnormalities in children with autistic disorder. OBJECTIVES: Our aim was to evaluate the structure and function of the upper gastrointestinal tract in a group of patients with autism who had gastrointestinal symptoms. STUDY DESIGN: Thirty-six children (age: 5.7 +/- 2 years, mean +/- SD) with autistic disorder underwent upper gastrointestinal endoscopy with biopsies, intestinal and pancreatic enzyme analyses, and bacterial and fungal cultures. The most frequent gastrointestinal complaints were chronic diarrhea, gaseousness, and abdominal discomfort and distension. RESULTS: Histologic examination in these 36 children revealed grade I or II reflux esophagitis in 25 (69.4%), chronic gastritis in 15, and chronic duodenitis in 24. The number of Paneth's cells in the duodenal crypts was significantly elevated in autistic children compared with non-autistic control subjects. Low intestinal carbohydrate digestive enzyme activity was reported in 21 children (58.3%), although there was no abnormality found in pancreatic function. Seventy-five percent of the autistic children (27/36) had an increased pancreatico-biliary fluid output after intravenous secretin administration. Nineteen of the 21 patients with diarrhea had significantly higher fluid output than those without diarrhea. CONCLUSIONS: Unrecognized gastrointestinal disorders, especially reflux esophagitis and disaccharide malabsorption, may contribute to the behavioral problems of the non-verbal autistic patients. The observed increase in pancreatico-biliary secretion after secretin infusion suggests an upregulation of secretin receptors in the pancreas and liver. Further studies are required to determine the possible association between the brain and gastrointestinal dysfunctions in children with autistic disorder.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_005

Gastrointestinal abnormalities in children with autistic disorder. OBJECTIVES: Our aim was to evaluate the structure and function of the upper gastrointestinal tract in a group of patients with autism who had gastrointestinal symptoms. STUDY DESIGN: Thirty-six children (age: 5.7 +/- 2 years, mean +/- SD) with autistic disorder underwent upper gastrointestinal endoscopy with biopsies, intestinal and pancreatic enzyme analyses, and bacterial and fungal cultures. The most frequent gastrointestinal complaints were chronic diarrhea, gaseousness, and abdominal discomfort and distension. RESULTS: Histologic examination in these 36 children revealed grade I or II reflux esophagitis in 25 (69.4%), chronic gastritis in 15, and chronic duodenitis in 24. The number of Paneth's cells in the duodenal crypts was significantly elevated in autistic children compared with non-autistic control subjects. Low intestinal carbohydrate digestive enzyme activity was reported in 21 children (58.3%), although there was no abnormality found in pancreatic function. Seventy-five percent of the autistic children (27/36) had an increased pancreatico-biliary fluid output after intravenous secretin administration. Nineteen of the 21 patients with diarrhea had significantly higher fluid output than those without diarrhea. CONCLUSIONS: Unrecognized gastrointestinal disorders, especially reflux esophagitis and disaccharide malabsorption, may contribute to the behavioral problems of the non-verbal autistic patients. The observed increase in pancreatico-biliary secretion after secretin infusion suggests an upregulation of secretin receptors in the pancreas and liver. Further studies are required to determine the possible association between the brain and gastrointestinal dysfunctions in children with autistic disorder.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_006

Gastrointestinal abnormalities in children with autistic disorder. OBJECTIVES: Our aim was to evaluate the structure and function of the upper gastrointestinal tract in a group of patients with autism who had gastrointestinal symptoms. STUDY DESIGN: Thirty-six children (age: 5.7 +/- 2 years, mean +/- SD) with autistic disorder underwent upper gastrointestinal endoscopy with biopsies, intestinal and pancreatic enzyme analyses, and bacterial and fungal cultures. The most frequent gastrointestinal complaints were chronic diarrhea, gaseousness, and abdominal discomfort and distension. RESULTS: Histologic examination in these 36 children revealed grade I or II reflux esophagitis in 25 (69.4%), chronic gastritis in 15, and chronic duodenitis in 24. The number of Paneth's cells in the duodenal crypts was significantly elevated in autistic children compared with non-autistic control subjects. Low intestinal carbohydrate digestive enzyme activity was reported in 21 children (58.3%), although there was no abnormality found in pancreatic function. Seventy-five percent of the autistic children (27/36) had an increased pancreatico-biliary fluid output after intravenous secretin administration. Nineteen of the 21 patients with diarrhea had significantly higher fluid output than those without diarrhea. CONCLUSIONS: Unrecognized gastrointestinal disorders, especially reflux esophagitis and disaccharide malabsorption, may contribute to the behavioral problems of the non-verbal autistic patients. The observed increase in pancreatico-biliary secretion after secretin infusion suggests an upregulation of secretin receptors in the pancreas and liver. Further studies are required to determine the possible association between the brain and gastrointestinal dysfunctions in children with autistic disorder.

List the endoscopic diagnoses that have been reported in children with autism

515deafd298dcd4e51000025_007

Network analysis reveals patterns of antiepileptic drug use in children with medically intractable epilepsy. Network analysis is an emerging tool for the study of complex systems. Antiepileptic drug (AED) polytherapy in children with medically intractable epilepsy may be considered a complex system, given the heterogeneity of drug combinations that are frequently modified according to clinical indications. The current article presents a concise review of network theory and its application to the characterization of AED use in children with refractory epilepsy. Current and previous AEDs prescribed to 27 children with refractory, localization-related epilepsy were recorded, and network theory was applied to identify patterns of drug administration. Of the fifteen unique AEDs prescribed, levetiracetam possessed the highest betweenness centrality within the network. Furthermore, first generation AEDs were often discontinued, while lacosamide and topiramate were most likely to be initiated. We also identified three subnetworks of AEDs that were commonly coadministered. We conclude that network analysis is an effective method to characterize the complexity of AED administration patterns in children with epilepsy with many promising future applications.

What are the main indications of lacosamide?

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Therapeutic drug monitoring of antiepileptic drugs by use of saliva. Blood (serum/plasma) antiepileptic drug (AED) therapeutic drug monitoring (TDM) has proven to be an invaluable surrogate marker for individualizing and optimizing the drug management of patients with epilepsy. Since 1989, there has been an exponential increase in AEDs with 23 currently licensed for clinical use, and recently, there has been renewed and extensive interest in the use of saliva as an alternative matrix for AED TDM. The advantages of saliva include the fact that for many AEDs it reflects the free (pharmacologically active) concentration in serum; it is readily sampled, can be sampled repetitively, and sampling is noninvasive; does not require the expertise of a phlebotomist; and is preferred by many patients, particularly children and the elderly. For each AED, this review summarizes the key pharmacokinetic characteristics relevant to the practice of TDM, discusses the use of other biological matrices with particular emphasis on saliva and the evidence that saliva concentration reflects those in serum. Also discussed are the indications for salivary AED TDM, the key factors to consider when saliva sampling is to be undertaken, and finally, a practical protocol is described so as to enable AED TDM to be applied optimally and effectively in the clinical setting. Overall, there is compelling evidence that salivary TDM can be usefully applied so as to optimize the treatment of epilepsy with carbamazepine, clobazam, ethosuximide, gabapentin, lacosamide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, primidone, topiramate, and zonisamide. Salivary TDM of valproic acid is probably not helpful, whereas for clonazepam, eslicarbazepine acetate, felbamate, pregabalin, retigabine, rufinamide, stiripentol, tiagabine, and vigabatrin, the data are sparse or nonexistent.

What are the main indications of lacosamide?

52bf202003868f1b06000018_003

Evaluation of anticonvulsants for possible use in neuropathic pain. Neuropathic pain is a kind of pain related with functional abnormality of neurons. Despite large progress in pharmacotherapy, neuropathic pain is still considered an unmet need. Nowadays, there are few drugs registered for this condition, such as pregabalin, gabapentin, duloxetine, carbamazepine, and lidocaine. Among them, pregabalin, gabapentin and carbamazepine are well known antiepileptic drugs. Among the group of new antiepileptic drugs, which are addressed to 1% of human world population suffering from seizures, it turned out that 30% of the seizures resistant to pharmacotherapy has not enough market to justify the costs of drug development. Therefore, it is already a phenomenon that researchers turn their projects toward a larger market, related with possible similar mechanism. Anticonvulsant mechanism of action is in the first place among primary indications for drugs revealing potential analgesic activity. Therefore, many drug candidates for epilepsy, still in preclinical stage, are being evaluated for activity in neuropathic pain. This review is focusing on antiepileptic drugs, which are evaluated for their analgesic activity in major tests related with neuropathic pain. Relation between structure, mechanism of action and result in tests such as the Chung model (spinal nerve ligation SNL), the Bennett model (chronic constriction injury of sciatic nerve CCI) and other tests are considered. The first examples are carbamazepine, gabapentin, and lacosamide as drugs well established in epilepsy market as well as drug candidates such as valnoctamide, and other valproic acid derivatives, novel biphenyl pyrazole derivatives, etc. Moreover, clinical efficacy related with listed animal models has been discussed.

What are the main indications of lacosamide?

52bf202003868f1b06000018_004

Collapsin response mediator protein-2: an emerging pathologic feature and therapeutic target for neurodisease indications. Collapsin response mediator protein-2 (DPYSL2 or CRMP2) is a multifunctional adaptor protein within the central nervous system. In the developing brain or cell cultures, CRMP2 performs structural and regulatory functions related to cytoskeletal dynamics, vesicle trafficking and synaptic physiology whereas CRMP2 functions in adult brain are still being elucidated. CRMP2 has been associated with several neuropathologic or psychiatric conditions including Alzheimer's disease (AD) and schizophrenia, either at the level of genetic polymorphisms; protein expression; post-translational modifications; or protein/protein interactions. In AD, CRMP2 is phosphorylated by glycogen synthase kinase-3β (GSK3β) and cyclin dependent protein kinase-5 (CDK5), the same kinases that act on tau protein in generating neurofibrillary tangles (NFTs). Phosphorylated CRMP2 collects in NFTs in association with the synaptic structure-regulating SRA1/WAVE1 (specifically Rac1-associated protein-1/WASP family verprolin-homologous protein-1) complex. This phenomenon could plausibly contribute to deficits in neural and synaptic structure that have been well documented in AD. This review discusses the essential biology of CRMP2 in the context of nascent data implicating CRMP2 perturbations as either a correlate of, or plausible contributor to, diverse neuropathologies. A discussion is made of recent findings that the atypical antidepressant tianeptine increases CRMP2 expression, whereas other, neuroactive small molecules including the epilepsy drug lacosamide and the natural brain metabolite lanthionine ketimine appear to bind CRMP2 directly with concomitant affects on neural structure. These findings constitute proofs-of-concept that pharmacological manipulation of CRMP2 is possible and hence, may offer new opportunities for therapy development against certain neurological diseases.

What are the main indications of lacosamide?

52bf202003868f1b06000018_006

Lacosamide, a newer antiepileptic. Lacosamide (LCM) is a newer antiepileptic drug with a dual mode of action. It selectively enhances slow inactivation of voltage-gated sodium channels without affecting fast inactivation, and modulates collapsing response mediator protein 2 (CRMP-2). It has a high oral bioavailability of approximately 100%. It has shown potent and broad neuroprotective effects in vitro and in vivo animal models making it a potential candidate for long term treatment of epilepsy. In addition to this, it has demonstrated analgesic activity in various animal models. Apart from this, LCM has demonstrated potent effects in animal models for a variety of CNS disorders like schizophrenia and stress induced anxiety. Various safety pharmacology and toxicology studies have shown that LCM is well tolerated. Clinical trials have also suggested that LCM is a safe, effective, and well tolerated adjunctive treatment for reduction of seizure frequency in patients with highly refractory, partial seizures. Other potential indications of LCM are being investigated.

What are the main indications of lacosamide?

52bf202003868f1b06000018_007

Lacosamide, a newer antiepileptic. Lacosamide (LCM) is a newer antiepileptic drug with a dual mode of action. It selectively enhances slow inactivation of voltage-gated sodium channels without affecting fast inactivation, and modulates collapsing response mediator protein 2 (CRMP-2). It has a high oral bioavailability of approximately 100%. It has shown potent and broad neuroprotective effects in vitro and in vivo animal models making it a potential candidate for long term treatment of epilepsy. In addition to this, it has demonstrated analgesic activity in various animal models. Apart from this, LCM has demonstrated potent effects in animal models for a variety of CNS disorders like schizophrenia and stress induced anxiety. Various safety pharmacology and toxicology studies have shown that LCM is well tolerated. Clinical trials have also suggested that LCM is a safe, effective, and well tolerated adjunctive treatment for reduction of seizure frequency in patients with highly refractory, partial seizures. Other potential indications of LCM are being investigated.

What are the main indications of lacosamide?

52bf202003868f1b06000018_008

Lacosamide, a newer antiepileptic. Lacosamide (LCM) is a newer antiepileptic drug with a dual mode of action. It selectively enhances slow inactivation of voltage-gated sodium channels without affecting fast inactivation, and modulates collapsing response mediator protein 2 (CRMP-2). It has a high oral bioavailability of approximately 100%. It has shown potent and broad neuroprotective effects in vitro and in vivo animal models making it a potential candidate for long term treatment of epilepsy. In addition to this, it has demonstrated analgesic activity in various animal models. Apart from this, LCM has demonstrated potent effects in animal models for a variety of CNS disorders like schizophrenia and stress induced anxiety. Various safety pharmacology and toxicology studies have shown that LCM is well tolerated. Clinical trials have also suggested that LCM is a safe, effective, and well tolerated adjunctive treatment for reduction of seizure frequency in patients with highly refractory, partial seizures. Other potential indications of LCM are being investigated.

What are the main indications of lacosamide?

52bf202003868f1b06000018_009

Lacosamide: a review of preclinical properties. Lacosamide (LCM), (SPM 927, (R)-2-acetamido-N-benzyl-3-methoxypropionamide, previously referred to as harkoseride or ADD 234037) is a member of a series of functionalized amino acids that were specifically synthesized as anticonvulsive drug candidates. LCM has demonstrated antiepileptic effectiveness in different rodent seizure models and antinociceptive potential in experimental animal models that reflect distinct types and symptoms of neuropathic as well as chronic inflammatory pain. Recent results suggest that LCM has a dual mode of action underlying its anticonvulsant and analgesic activity. It was found that LCM selectively enhances slow inactivation of voltage-gated sodium channels without affecting fast inactivation. Furthermore, employing proteomic affinity-labeling techniques, collapsin-response mediator protein 2 (CRMP-2 alias DRP-2) was identified as a binding partner. Follow-up experiments confirmed a functional interaction of LCM with CRMP-2 in vitro. LCM did not inhibit or induce a wide variety of cytochrome P450 enzymes at therapeutic concentrations. In safety pharmacology and toxicology studies conducted in mice, rats, rabbits, and dogs, LCM was well tolerated. Either none or only minor side effects were observed in safety studies involving the central nervous, respiratory, gastrointestinal, and renal systems and there is no indication of abuse liability. Repeated dose toxicity studies demonstrated that after either intravenous or oral administration of LCM the adverse events were reversible and consisted mostly of exaggerated pharmacodynamic effects on the CNS. No genotoxic or carcinogenic effects were observed in vivo, and LCM showed a favorable profile in reproductive and developmental animal studies. Currently, LCM is in a late stage of clinical development as an adjunctive treatment for patients with uncontrolled partial-onset seizures, and it is being assessed as monotherapy in patients with painful diabetic neuropathy. Further trials to identify LCM's potential in pain and for other indications have been initiated.

What are the main indications of lacosamide?

52bf202003868f1b06000018_010

Lacosamide, a novel anti-convulsant drug, shows efficacy with a wide safety margin in rodent models for epilepsy. This paper comprises a series of experiments in rodent models of partial and generalized epilepsy which were designed to describe the anti-convulsant profile of the functionalized amino acid lacosamide. Lacosamide was effective against sound-induced seizures in the genetically susceptible Frings mouse, against maximal electroshock test (MES)-induced seizures in rats and mice, in the rat hippocampal kindling model of partial seizures, and in the 6Hz model of psychomotor seizures in mice. The activity in the MES test in both mice (4.5mg/kg i.p.) and rats (3.9 mg/kg p.o.) fell within the ranges previously reported for most clinically available anti-epileptic drugs. At both the median effective dose for MES protection, as well as the median toxic dose for rotorod impairment, lacosamide elevated the seizure threshold in the i.v. pentylenetetrazol seizure test, suggesting that it is unlikely to be pro-convulsant at high doses. Lacosamide was inactive against clonic seizures induced by subcutaneous administration of the chemoconvulsants pentylenetetrazol, bicuculline, and picrotoxin, but it did inhibit NMDA-induced seizures in mice and showed full efficacy in the homocysteine model of epilepsy. In summary, the overall anti-convulsant profile of lacosamide appeared to be unique, and the drug displayed a good margin of safety in those tests in which it was effective. These results suggest that lacosamide may have the potential to be clinically useful for at least the treatment of generalized tonic-clonic and partial-onset epilepsies, and support ongoing clinical trials in these indications.

What are the main indications of lacosamide?

52bf202003868f1b06000018_011

Hemolytic uremic syndrome. Hemolytic uremic syndrome (HUS) is a clinical syndrome characterized by the triad of thrombotic microangiopathy, thrombocytopenia, and acute kidney injury. Hemolytic uremic syndrome represents a heterogeneous group of disorders with variable etiologies that result in differences in presentation, management and outcome. In recent years, better understanding of the HUS, especially those due to genetic mutations in the alternative complement pathway have provided an update on the terminology, classification, and treatment of the disease. This review will provide the updated classification of the disease and the current diagnostic and therapeutic approaches on the complement-mediated HUS in addition to STEC-HUS which is the most common cause of the HUS in childhood.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_001

[Atypical HUS caused by complement-related abnormalities]. Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. The term aHUS was historically used to distinguish this disorder from Shiga-toxin producing Escherichia coli (STEC)-HUS. Many aHUS cases (approximately 70%) are reportedly caused by uncontrolled complement activation due to genetic mutations in the alternative pathway, including complement factor H (CFH), complement factor I (CFI), membrane cofactor protein (MCP), thrombomodulin (THBD), complement component C3 (C3), and complement factor B (CFB). Mutations in the coagulation pathway, such as diacylglycerol kinase ε (DGKE) and plasminogen, are also reported to be causes of aHUS. In this review, we have focused on aHUS due to complement dysfunction. aHUS is suspected based on plasma ADAMTS13 activity of 10% or more, and being negative for STEC-HUS, in addition to the aforementioned triad. Complement genetic studies provide a more specific diagnosis of aHUS. Plasma therapy is the first-line treatment for patients with aHUS and should be initiated as soon as the diagnosis is suspected. Recently, eculizumab, a humanized monoclonal antibody against C5, was shown to be an effective treatment for aHUS. Therefore, early diagnosis and identification of the underlying pathogenic mechanism is important for improving the outcome of aHUS.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_002

[Atypical HUS caused by complement-related abnormalities]. Atypical hemolytic uremic syndrome (aHUS) is a rare disease characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. The term aHUS was historically used to distinguish this disorder from Shiga-toxin producing Escherichia coli (STEC)-HUS. Many aHUS cases (approximately 70%) are reportedly caused by uncontrolled complement activation due to genetic mutations in the alternative pathway, including complement factor H (CFH), complement factor I (CFI), membrane cofactor protein (MCP), thrombomodulin (THBD), complement component C3 (C3), and complement factor B (CFB). Mutations in the coagulation pathway, such as diacylglycerol kinase ε (DGKE) and plasminogen, are also reported to be causes of aHUS. In this review, we have focused on aHUS due to complement dysfunction. aHUS is suspected based on plasma ADAMTS13 activity of 10% or more, and being negative for STEC-HUS, in addition to the aforementioned triad. Complement genetic studies provide a more specific diagnosis of aHUS. Plasma therapy is the first-line treatment for patients with aHUS and should be initiated as soon as the diagnosis is suspected. Recently, eculizumab, a humanized monoclonal antibody against C5, was shown to be an effective treatment for aHUS. Therefore, early diagnosis and identification of the underlying pathogenic mechanism is important for improving the outcome of aHUS.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_003

Refractory atypical hemolytic uremic syndrome with monoclonal gammopathy responsive to bortezomib-based therapy. Atypical hemolytic uremic syndrome (aHUS) is a relatively rare disorder described by the triad of hemolytic anemia, thrombocytopenia, and renal failure. Atypical HUS could be genetic, acquired, or idiopathic (without known genetic changes or environmental triggers). Monoclonal protein has uncommonly been reported as a cause of microangiopathic hemolytic anemia (MAHA). We report a 59-year-old white man who presented with acute kidney injury (AKI) with MAHA and was given a diagnosis of aHUS with monoclonal gammopathy. His kidney function and proteinuria worsened with persistent hemolysis despite eculizumab and later cyclophosphamide and prednisone treatment. He responded well to VRD (bortezomib, lenalidomide, and dexamethasone) regimen. Renal function, proteinuria, and hemolysis all improved, and he was been in remission for more than 15 months. To our knowledge, this is the first report of successful treatment with bortezomib-based regimen for a patient with aHUS and monoclonal protein refractory to eculizumab therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_004

Refractory atypical hemolytic uremic syndrome with monoclonal gammopathy responsive to bortezomib-based therapy. Atypical hemolytic uremic syndrome (aHUS) is a relatively rare disorder described by the triad of hemolytic anemia, thrombocytopenia, and renal failure. Atypical HUS could be genetic, acquired, or idiopathic (without known genetic changes or environmental triggers). Monoclonal protein has uncommonly been reported as a cause of microangiopathic hemolytic anemia (MAHA). We report a 59-year-old white man who presented with acute kidney injury (AKI) with MAHA and was given a diagnosis of aHUS with monoclonal gammopathy. His kidney function and proteinuria worsened with persistent hemolysis despite eculizumab and later cyclophosphamide and prednisone treatment. He responded well to VRD (bortezomib, lenalidomide, and dexamethasone) regimen. Renal function, proteinuria, and hemolysis all improved, and he was been in remission for more than 15 months. To our knowledge, this is the first report of successful treatment with bortezomib-based regimen for a patient with aHUS and monoclonal protein refractory to eculizumab therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_005

A mechanistic approach to the diagnosis and management of atypical hemolytic uremic syndrome. Until recently, atypical hemolytic uremic syndrome (aHUS), conventionally defined in the pediatric literature as a syndrome of the triad of renal failure, microangiopathic hemolytic anemia, and thrombocytopenia without a prodrome of hemorrhagic diarrhea, has received little attention in adult practice because the patients are commonly given the diagnosis of thrombotic thrombocytopenic purpura (TTP) or TTP/HUS and treated as TTP with plasma exchange, augmented in refractory cases with rituximab and sometimes even splenectomy. Molecular studies have shown that the regulation of the alternative complement pathway is defective in many patients with conventionally defined aHUS. With this new knowledge and the findings of ADAMTS13 autoinhibitors or mutations in TTP, it is time to redefine aHUS as a disorder with propensity to the development of thrombotic microangiopathy due to defective regulation of the alternative complement pathway and TTP as a disorder with propensity to arteriolar and capillary thrombosis due to ADAMTS13 deficiency. This new definition provides a clear distinction of aHUS from TTP, encompasses patients without all 3 components of the triad, and provides the rationale for management with anticomplement therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_006

A mechanistic approach to the diagnosis and management of atypical hemolytic uremic syndrome. Until recently, atypical hemolytic uremic syndrome (aHUS), conventionally defined in the pediatric literature as a syndrome of the triad of renal failure, microangiopathic hemolytic anemia, and thrombocytopenia without a prodrome of hemorrhagic diarrhea, has received little attention in adult practice because the patients are commonly given the diagnosis of thrombotic thrombocytopenic purpura (TTP) or TTP/HUS and treated as TTP with plasma exchange, augmented in refractory cases with rituximab and sometimes even splenectomy. Molecular studies have shown that the regulation of the alternative complement pathway is defective in many patients with conventionally defined aHUS. With this new knowledge and the findings of ADAMTS13 autoinhibitors or mutations in TTP, it is time to redefine aHUS as a disorder with propensity to the development of thrombotic microangiopathy due to defective regulation of the alternative complement pathway and TTP as a disorder with propensity to arteriolar and capillary thrombosis due to ADAMTS13 deficiency. This new definition provides a clear distinction of aHUS from TTP, encompasses patients without all 3 components of the triad, and provides the rationale for management with anticomplement therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_007

Diagnostic criteria for atypical hemolytic uremic syndrome proposed by the Joint Committee of the Japanese Society of Nephrology and the Japan Pediatric Society. Atypical hemolytic uremic syndrome (aHUS) is rare and comprises the triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute kidney injury. Recently, abnormalities in the mechanisms underlying complement regulation have been focused upon as causes of aHUS. The prognosis for patients who present with aHUS is very poor, with the first aHUS attack being associated with a mortality rate of approximately 25%, and with approximately 50% of cases resulting in end-stage renal disease requiring dialysis. If treatment is delayed, there is a high risk of this syndrome progressing to renal failure. Therefore, we have developed diagnostic criteria for aHUS to enable its early diagnosis and to facilitate the timely initiation of appropriate treatment. We hope these diagnostic criteria will be disseminated to as many clinicians as possible and that they will be used widely.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_008

Adult hemolytic uremic syndrome associated with Streptococcus pneumoniae. Hemolyitic uremic syndrome (HUS), characterized by triad of acute kidney injury, thrombocytopenia, and hemolytic anemia, has considerable morbidity and mortality and is known to be associated with diarrheal illness. It usually occurs after a diarrheal illness due to Shiga-toxin-producing Escherichia coli. Streptococcus pneumoniae is a rare but well recognized trigger for non-diarrhea associated HUS in children, but has not been reported in adults. We report a case of an adult presenting with pneumococcal pneumonia complicated by HUS and required renal replacement therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_009

Gemcitabine-induced hemolytic uremic syndrome in pancreatic cancer: a case report and review of the literature. Hemolytic uremic syndrome (HUS) is a rare thrombotic complication characterized by a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. HUS may be caused by several different conditions, including infection, malignancy, and chemotherapeutic agents, such as mitomycin, cisplatin, and most recently, gemcitabine. The outcome of gemcitabine-induced HUS is poor, and the disease has a high mortality rate. This study reports a case of gemcitabine-induced HUS in a patient with pancreatic cancer in Korea.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_010

Gemcitabine-induced hemolytic uremic syndrome in pancreatic cancer: a case report and review of the literature. Hemolytic uremic syndrome (HUS) is a rare thrombotic complication characterized by a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. HUS may be caused by several different conditions, including infection, malignancy, and chemotherapeutic agents, such as mitomycin, cisplatin, and most recently, gemcitabine. The outcome of gemcitabine-induced HUS is poor, and the disease has a high mortality rate. This study reports a case of gemcitabine-induced HUS in a patient with pancreatic cancer in Korea.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_011

A mechanistic approach to the diagnosis and management of atypical hemolytic uremic syndrome. Until recently, atypical hemolytic uremic syndrome (aHUS), conventionally defined in the pediatric literature as a syndrome of the triad of renal failure, microangiopathic hemolytic anemia, and thrombocytopenia without a prodrome of hemorrhagic diarrhea, has received little attention in adult practice because the patients are commonly given the diagnosis of thrombotic thrombocytopenic purpura (TTP) or TTP/HUS and treated as TTP with plasma exchange, augmented in refractory cases with rituximab and sometimes even splenectomy. Molecular studies have shown that the regulation of the alternative complement pathway is defective in many patients with conventionally defined aHUS. With this new knowledge and the findings of ADAMTS13 autoinhibitors or mutations in TTP, it is time to redefine aHUS as a disorder with propensity to the development of thrombotic microangiopathy due to defective regulation of the alternative complement pathway and TTP as a disorder with propensity to arteriolar and capillary thrombosis due to ADAMTS13 deficiency. This new definition provides a clear distinction of aHUS from TTP, encompasses patients without all 3 components of the triad, and provides the rationale for management with anticomplement therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_012

A mechanistic approach to the diagnosis and management of atypical hemolytic uremic syndrome. Until recently, atypical hemolytic uremic syndrome (aHUS), conventionally defined in the pediatric literature as a syndrome of the triad of renal failure, microangiopathic hemolytic anemia, and thrombocytopenia without a prodrome of hemorrhagic diarrhea, has received little attention in adult practice because the patients are commonly given the diagnosis of thrombotic thrombocytopenic purpura (TTP) or TTP/HUS and treated as TTP with plasma exchange, augmented in refractory cases with rituximab and sometimes even splenectomy. Molecular studies have shown that the regulation of the alternative complement pathway is defective in many patients with conventionally defined aHUS. With this new knowledge and the findings of ADAMTS13 autoinhibitors or mutations in TTP, it is time to redefine aHUS as a disorder with propensity to the development of thrombotic microangiopathy due to defective regulation of the alternative complement pathway and TTP as a disorder with propensity to arteriolar and capillary thrombosis due to ADAMTS13 deficiency. This new definition provides a clear distinction of aHUS from TTP, encompasses patients without all 3 components of the triad, and provides the rationale for management with anticomplement therapy.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_013

Atypical hemolytic uremic syndrome in the Tunisian population. BACKGROUND: Hemolytic uremic syndrome consists of a triad of acquired hemolytic anemia, thrombocytopenia and renal failure. AIM: Our objectives were to determine epidemiology, clinical and laboratory characteristics of patients with atypical hemolytic uremic syndrome (aHUS) to determine the relationship between the complement protein deficit and aHUS in the Tunisian population. METHODS: We studied retrospectively four cases of atypical HUS in adults admitted in the Nephrology Department of Fattouma Bourguiba Universitary Hospital in Monastir between 2000 and 2008. RESULTS: Three patients had renal failure that required dialysis. One of them received kidney transplantation with no further recurrence of aHUS. Three patients had normal C3, C4, CFH, and FB levels, and in all patients anti-FH autoantibodies were absent. The kidney biopsy of one patient showed in addition to lupus glomerulonephritis histological findings consistent with TMA. A decrease in C3, C4 and CFH levels in this patient was found both before and after the cure. CONCLUSION: Nephrologists should be aware of autoimmune conditions and genetic abnormalities of the complement regulatory genes as possible pathogenic mechanisms in atypical HUS patients.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_014

Atypical hemolytic uremic syndrome in the Tunisian population. BACKGROUND: Hemolytic uremic syndrome consists of a triad of acquired hemolytic anemia, thrombocytopenia and renal failure. AIM: Our objectives were to determine epidemiology, clinical and laboratory characteristics of patients with atypical hemolytic uremic syndrome (aHUS) to determine the relationship between the complement protein deficit and aHUS in the Tunisian population. METHODS: We studied retrospectively four cases of atypical HUS in adults admitted in the Nephrology Department of Fattouma Bourguiba Universitary Hospital in Monastir between 2000 and 2008. RESULTS: Three patients had renal failure that required dialysis. One of them received kidney transplantation with no further recurrence of aHUS. Three patients had normal C3, C4, CFH, and FB levels, and in all patients anti-FH autoantibodies were absent. The kidney biopsy of one patient showed in addition to lupus glomerulonephritis histological findings consistent with TMA. A decrease in C3, C4 and CFH levels in this patient was found both before and after the cure. CONCLUSION: Nephrologists should be aware of autoimmune conditions and genetic abnormalities of the complement regulatory genes as possible pathogenic mechanisms in atypical HUS patients.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_015

Atypical hemolytic uremic syndrome. Hemolytic uremic syndrome (HUS) is a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. The atypical form of HUS is a disease characterized by complement overactivation. Inherited defects in complement genes and acquired autoantibodies against complement regulatory proteins have been described. Incomplete penetrance of mutations in all predisposing genes is reported, suggesting that a precipitating event or trigger is required to unmask the complement regulatory deficiency. The underlying genetic defect predicts the prognosis both in native kidneys and after renal transplantation. The successful trials of the complement inhibitor eculizumab in the treatment of atypical HUS will revolutionize disease management.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_016

Atypical hemolytic uremic syndrome. Hemolytic uremic syndrome (HUS) is a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. The atypical form of HUS is a disease characterized by complement overactivation. Inherited defects in complement genes and acquired autoantibodies against complement regulatory proteins have been described. Incomplete penetrance of mutations in all predisposing genes is reported, suggesting that a precipitating event or trigger is required to unmask the complement regulatory deficiency. The underlying genetic defect predicts the prognosis both in native kidneys and after renal transplantation. The successful trials of the complement inhibitor eculizumab in the treatment of atypical HUS will revolutionize disease management.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_017

Thrombomodulin mutations in atypical hemolytic-uremic syndrome. BACKGROUND: The hemolytic-uremic syndrome consists of the triad of microangiopathic hemolytic anemia, thrombocytopenia, and renal failure. The common form of the syndrome is triggered by infection with Shiga toxin-producing bacteria and has a favorable outcome. The less common form of the syndrome, called atypical hemolytic-uremic syndrome, accounts for about 10% of cases, and patients with this form of the syndrome have a poor prognosis. Approximately half of the patients with atypical hemolytic-uremic syndrome have mutations in genes that regulate the complement system. Genetic factors in the remaining cases are unknown. We studied the role of thrombomodulin, an endothelial glycoprotein with anticoagulant, antiinflammatory, and cytoprotective properties, in atypical hemolytic-uremic syndrome. METHODS: We sequenced the entire thrombomodulin gene (THBD) in 152 patients with atypical hemolytic-uremic syndrome and in 380 controls. Using purified proteins and cell-expression systems, we investigated whether thrombomodulin regulates the complement system, and we characterized the mechanisms. We evaluated the effects of thrombomodulin missense mutations associated with atypical hemolytic-uremic syndrome on complement activation by expressing thrombomodulin variants in cultured cells. RESULTS: Of 152 patients with atypical hemolytic-uremic syndrome, 7 unrelated patients had six different heterozygous missense THBD mutations. In vitro, thrombomodulin binds to C3b and factor H (CFH) and negatively regulates complement by accelerating factor I-mediated inactivation of C3b in the presence of cofactors, CFH or C4b binding protein. By promoting activation of the plasma procarboxypeptidase B, thrombomodulin also accelerates the inactivation of anaphylatoxins C3a and C5a. Cultured cells expressing thrombomodulin variants associated with atypical hemolytic-uremic syndrome had diminished capacity to inactivate C3b and to activate procarboxypeptidase B and were thus less protected from activated complement. CONCLUSIONS: Mutations that impair the function of thrombomodulin occur in about 5% of patients with atypical hemolytic-uremic syndrome.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_018

Thrombomodulin mutations in atypical hemolytic-uremic syndrome. BACKGROUND: The hemolytic-uremic syndrome consists of the triad of microangiopathic hemolytic anemia, thrombocytopenia, and renal failure. The common form of the syndrome is triggered by infection with Shiga toxin-producing bacteria and has a favorable outcome. The less common form of the syndrome, called atypical hemolytic-uremic syndrome, accounts for about 10% of cases, and patients with this form of the syndrome have a poor prognosis. Approximately half of the patients with atypical hemolytic-uremic syndrome have mutations in genes that regulate the complement system. Genetic factors in the remaining cases are unknown. We studied the role of thrombomodulin, an endothelial glycoprotein with anticoagulant, antiinflammatory, and cytoprotective properties, in atypical hemolytic-uremic syndrome. METHODS: We sequenced the entire thrombomodulin gene (THBD) in 152 patients with atypical hemolytic-uremic syndrome and in 380 controls. Using purified proteins and cell-expression systems, we investigated whether thrombomodulin regulates the complement system, and we characterized the mechanisms. We evaluated the effects of thrombomodulin missense mutations associated with atypical hemolytic-uremic syndrome on complement activation by expressing thrombomodulin variants in cultured cells. RESULTS: Of 152 patients with atypical hemolytic-uremic syndrome, 7 unrelated patients had six different heterozygous missense THBD mutations. In vitro, thrombomodulin binds to C3b and factor H (CFH) and negatively regulates complement by accelerating factor I-mediated inactivation of C3b in the presence of cofactors, CFH or C4b binding protein. By promoting activation of the plasma procarboxypeptidase B, thrombomodulin also accelerates the inactivation of anaphylatoxins C3a and C5a. Cultured cells expressing thrombomodulin variants associated with atypical hemolytic-uremic syndrome had diminished capacity to inactivate C3b and to activate procarboxypeptidase B and were thus less protected from activated complement. CONCLUSIONS: Mutations that impair the function of thrombomodulin occur in about 5% of patients with atypical hemolytic-uremic syndrome.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_019

Neonatal onset atypical hemolytic uremic syndrome successfully treated with eculizumab. BACKGROUND: Atypical hemolytic uremic syndrome (aHUS) is characterized by the triad of microangiopathic hemolytic anemia, thrombocytopenia, and renal impairment. Neonatal cases are extremely uncommon. Plasma therapy is the first choice therapy in patients with aHUS based on the belief of an underlying complement dysregulation. Alternatively, eculizumab, which targets complement 5, is used to block complement activation. CASE-DIAGNOSIS/TREATMENT: Sudden onset macroscopic hematuria, hypertension, and bruises over the entire body were noted in a 5 day-old newborn. Investigations revealed hemolytic anemia, thrombocytopenia, renal impairment, and a low serum C3, leading to the diagnosis of aHUS. Fresh frozen plasma (FFP) infusions and peritoneal dialysis for acute kidney injury were initiated. This approach yielded full renal and hematological remission. The patient was discharged with FFP infusions, but subsequently developed three life-threatening disease recurrences at 1, 3, and 6 months of age. The last relapse presented with uncontrolled hypertension and impaired renal function while the patient was receiving FFP infusions. After the first dose of eculizumab, his renal and hematological parameters returned to normal and his blood pressure normalized. Genetic screening of the CFH gene revealed a novel homozygous p. Tyr1177Cys mutation. CONCLUSION: Eculizumab can be considered as an alternative to plasma therapy in the treatment of specific patients with aHUS, even in infants.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_020

Hemolytic-uremic syndrome associated with gemcitabine: a case report and review of literature. CONTEXT: Hemolytic uremic syndrome is a rare condition compromising the clinical triad of acute renal failure, microangiopathic hemolytic anemia, and thrombocytopenia. Hemolytic uremic syndrome may be associated with a variety of etiologies, and chemotherapeutic agents have also been reported to be associated with hemolytic uremic syndrome, including mitomycin, cisplatin, bleomycin, and most recently gemcitabine. CASE REPORT: A 72-year-old Caucasian male treated with four cycles of gemcitabine at 1,000 mg/m2 developed clinical and laboratory findings compatible with hemolytic uremic syndrome. He developed microangiopathic hemolysis, rapidly declining renal function with proteinuria and hematuria, and renal biopsy revealed thrombotic microangiopathy. Hemodialysis, plasmapheresis, and corticosteroid therapy were utilized but the process ultimately was irreversible. CONCLUSION: With multiple reports of hemolytic uremic syndrome complicating gemcitabine therapy, it is imperative that clinicians heighten their awareness of this potentially lethal complication.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_021

Hemolytic-uremic syndrome associated with gemcitabine: a case report and review of literature. CONTEXT: Hemolytic uremic syndrome is a rare condition compromising the clinical triad of acute renal failure, microangiopathic hemolytic anemia, and thrombocytopenia. Hemolytic uremic syndrome may be associated with a variety of etiologies, and chemotherapeutic agents have also been reported to be associated with hemolytic uremic syndrome, including mitomycin, cisplatin, bleomycin, and most recently gemcitabine. CASE REPORT: A 72-year-old Caucasian male treated with four cycles of gemcitabine at 1,000 mg/m2 developed clinical and laboratory findings compatible with hemolytic uremic syndrome. He developed microangiopathic hemolysis, rapidly declining renal function with proteinuria and hematuria, and renal biopsy revealed thrombotic microangiopathy. Hemodialysis, plasmapheresis, and corticosteroid therapy were utilized but the process ultimately was irreversible. CONCLUSION: With multiple reports of hemolytic uremic syndrome complicating gemcitabine therapy, it is imperative that clinicians heighten their awareness of this potentially lethal complication.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_022

Hemolytic uremic syndrome in children in Puerto Rico: a rare disease with atypical features. Hemolytic Uremic Syndrome (HUS) consists ofa triad of acquired hemolytic anemia, thrombocytopenia, and renal failure that occurs acutely in otherwise healthy individuals. HUS may be divided into two broad categories, typical, preceded by a diarrheal prodrome, and atypical. The clinical symptoms of HUS as well as its course, prognosis, and response to treatment appear to be significantly influenced by a number of factors, including age at onset, type and severity of underlying infections, and/or systemic diseases. A retrospective case series review of five patients diagnosed with Hemolytic Uremic Syndrome at the Pediatric University Hospital in Puerto Rico between 1997-2007 was performed. The study showed that the incidence of HUS in children in Puerto Rico is lower than other countries. However, the majority of cases have an atypical presentation, which places our patients at higher risk for life-threatening complications.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_023

Hemolytic uremic syndrome in children in Puerto Rico: a rare disease with atypical features. Hemolytic Uremic Syndrome (HUS) consists ofa triad of acquired hemolytic anemia, thrombocytopenia, and renal failure that occurs acutely in otherwise healthy individuals. HUS may be divided into two broad categories, typical, preceded by a diarrheal prodrome, and atypical. The clinical symptoms of HUS as well as its course, prognosis, and response to treatment appear to be significantly influenced by a number of factors, including age at onset, type and severity of underlying infections, and/or systemic diseases. A retrospective case series review of five patients diagnosed with Hemolytic Uremic Syndrome at the Pediatric University Hospital in Puerto Rico between 1997-2007 was performed. The study showed that the incidence of HUS in children in Puerto Rico is lower than other countries. However, the majority of cases have an atypical presentation, which places our patients at higher risk for life-threatening complications.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_024

[Reversible cerebral changes in hemolytic-uremic syndrome]. The hemolytic-uremic syndrome is a pathology characterized by a triad consisting of acute renal failure, microangiopathic hemolytic anemia and thrombocytopenia, with complications of the central nervous system arising in a considerable number of cases. Altered cranial computerized tomography examinations usually reveal cerebral infarctions. We present here two cases in which diffuse hypodensity was observed in the white matter in addition to the infarcts. This hypodensity was reversible after resolution of the acute phase of the disease, as is also the case for the alterations described in uremic encephalopathy and in hypertensive encephalopathy of other etiologies.

List Hemolytic Uremic Syndrome Triad.

56be0da3ef6e394741000007_025