bluuebunny/crossref_metadata_2025 huggingface dataset:
Myocardial ischemia-reperfusion damage impacts occurrence of ventricular fibrillation in dogs
Authors: Xing Dezhi , Martins James B.
Published: December 2017

To define the relationship between ischemia-reperfusion-induced myocardial damage (IRD) and the occurrence of ventricular tachycardia (VT) and fibrillation (VF), we studied 23 dogs with a three-dimensional activation mapping system. Left anterior descending (LAD) coronary artery occlusion and reperfusion were performed while recording electrograms during VF and atrial pacing. Prior nonischemic sites showing IRD, defined as at least 10% loss of electrogram voltage after reperfusion, had the longest ventricular effective refractory periods (ERPs). IRD sites also occurred more frequently in dogs with reperfusion VF (44 ± 2 sites, P < 0.01) compared with dogs with VT (18 ± 5 sites) and no VT (16 ± 3 sites). In dogs ( n= 3) with 3 h of reperfusion, 95% of IRD sites still had lower voltage than those recorded during occlusion. Activation mapping of the first eight complexes of VF had Purkinje or endocardial focal origin in 57%, and complexes originated from IRD sites in 28%. In contrast, dogs with only reperfusion VT also had Purkinje or endocardial focal origin in 79%, but only 5% ( P < 0.01 vs. VF dogs) of the sites of origin had IRD. Therefore, dogs with reperfusion VF had more IRD sites where the ERP was longest, and more focal ventricular complexes originated from IRD sites, indicating that IRD may be one important factor in the occurrence of VF during reperfusion.
Importance of ventilation in modulating interaction between sympathetic drive and cardiovascular variability
Authors: Van De Borne Philippe , Montano Nicola , Narkiewicz Krzysztof , Degaute Jean P. , Malliani Alberto , Pagani Massimo , Somers Virend K.
Published: December 2017

Chemoreflex stimulation elicits both hyperventilation and sympathetic activation, each of which may have different influences on oscillatory characteristics of cardiovascular variability. We examined the influence of hyperventilation on the interactions between changes in R-R interval (RR) and muscle sympathetic nerve activity (MSNA) and changes in neurocirculatory variability, in 14 healthy subjects. We performed spectral analysis of RR and MSNA variability during each of the following interventions: 1) controlled breathing, 2) maximal end-expiratory apnea, 3) isocapnic voluntary hyperventilation, and 4) hypercapnia-induced hyperventilation. MSNA increased from 100% during controlled breathing to 170 ± 25% during apnea ( P = 0.02). RR was unchanged, but normalized low-frequency (LF) variability of both RR and MSNA increased markedly ( P < 0.001). During isocapnic hyperventilation, minute ventilation increased to 20.2 ± 1.4 l/min ( P < 0.0001). During hypercapnic hyperventilation, minute ventilation also increased (to 19.7 ± 1.7 l/min) as did end-tidal CO2 (both P < 0.0001). MSNA remained unchanged during isocapnic hyperventilation (104 ± 7%) but increased to 241 ± 49% during hypercapnic hyperventilation ( P < 0.01). RR decreased during both isocapnic and hypercapnic hyperventilation ( P < 0.05). However, normalized LF variability of RR and of MSNA decreased ( P < 0.05) during both isocapnic and hypercapnic hyperventilation, despite the tachycardia and heightened sympathetic nerve traffic. In conclusion, marked respiratory oscillations in autonomic drive induced by hyperventilation may induce dissociation between RR, MSNA, and neurocirculatory variability, perhaps by suppressing central genesis and/or inhibiting transmission of LF cardiovascular rhythms.
Na+/H+ exchange inhibition reduces hypertrophy and heart failure after myocardial infarction in rats
Authors: Kusumoto Keiji , Haist James V. , Karmazyn Morris
Published: December 2017

We investigated the effect of sodium/hydrogen exchange inhibition (NHE-1) on hypertrophy and heart failure after coronary artery ligation (CAL) in the rat. Animals were subjected to occlusion (or sham) of the left main coronary artery and immediately administered a control diet or one consisting of the NHE-1 inhibitor cariporide for 13–15 wk. Hearts were separated by small [≤30% of left ventricle (LV)] and large (>30% of LV) infarcts. CAL depressed change in left ventricular increase in pressure over time (LV +dP/d t) in small and large infarct groups by 18.8% ( P < 0.05) and 34% ( P < 0.01), respectively, whereas comparative values for the cariporide groups were 8.7% (not significant) and 23.1% ( P < 0.01), respectively. LV end-diastolic pressure was increased by 1,225% in the control large infarct group but was significantly reduced to 447% with cariporide. Cariporide also significantly reduced the degree of LV dilation in animals with large infarcts. Hypertrophy, defined by tissue weights and cell size, was reduced by cariporide, and shortening of surviving myocytes was preserved. Infarct sizes were unaffected by cariporide, and the drug had no influence on either blood pressure or the depressed inotropic response of infarcted hearts to dobutamine. These results suggest an important role for NHE-1 in the progression of heart failure after myocardial infarction.
Mechanisms of coronary angiogenesis in response to stretch: role of VEGF and TGF-β
Authors: Zheng Wei , Seftor Elisabeth A. , Meininger Cynthia J. , Hendrix Mary J. C. , Tomanek Robert J.
Published: December 2017

To test the hypotheses that cyclic stretch of 1) cardiac myocytes produces factors that trigger angiogenic events in coronary microvascular endothelial cells (CMEC) and 2) CMEC enhances the expression of growth factors, cardiac myocytes and CMEC were subjected to cyclic stretch in a Flexercell Strain Unit. Vascular endothelial growth factor (VEGF) but not basic fibroblast growth factor mRNA and protein levels increased approximately twofold in myocytes after 1 h of stretch. CMEC DNA synthesis increased approximately twofold when conditioned medium from stretched myocytes or VEGF protein was added, and addition of VEGF neutralizing antibody blocked the increase. CMEC migration and tube formation increased with the addition of conditioned media but were markedly attenuated by VEGF neutralizing antibody. Myocyte tumor growth factor-β (TGF-β) increased 2.5-fold after 1 h of stretch, and the addition of TGF-β neutralizing antibodies inhibited the stretch-induced upregulation of VEGF. Stretch of CMEC increased VEGF mRNA in these cells (determined by Northern blot and RT-PCR) and increased the levels of VEGF protein (determined by ELISA analysis) in the conditioned media. Therefore, cyclic stretch of cardiac myocytes and CMEC appears to be an important primary stimulus for coronary angiogenesis through both paracrine and autocrine VEGF pathways. These data indicate that 1) CMEC DNA synthesis, migration, and tube formation are increased in response to VEGF secreted from stretched cardiac myocytes; 2) VEGF in CMEC subjected to stretch is upregulated and secreted; and 3) TGF-β signaling may regulate VEGF expression in cardiac myocytes.
Activation of purified cardiac ryanodine receptors by dihydropyridine agonists
Authors: Sagawa Toshio , Nishio Manabu , Sagawa Kazuko , Kelly James E. , Lokuta Andrew J. , Tsai John , Kan Edward , Wasserstrom J. Andrew
Published: December 2017

Prior observations have raised the possibility that dihydropyridine (DHP) agonists directly affect the sarcoplasmic reticulum (SR) cardiac Ca2+release channel [i.e., ryanodine receptor (RyR)]. In single-channel recordings of purified canine cardiac RyR, both DHP agonists (−)-BAY K 8644 and (+)-SDZ202-791 increased the open probability of the RyR when added to the cytoplasmic face of the channel. Importantly, the DHP antagonists nifedipine and (−)-SDZ202-791 had no competitive blocking effects either alone or after channel activation with agonist. Thus there is a stereospecific effect of SDZ202-791, such that the agonist activates the channel, whereas the antagonist has little effect on channel activity. Further experiments showed that DHP agonists changed RyR activation by suppressing Ca2+-induced inactivation of the channel. We concluded that DHP agonists can also influence RyR single-channel activity directly at a unique allosteric site located on the cytoplasmic face of the channel. Similar results were obtained in human purified cardiac RyR. An implication of these data is that RyR activation by DHP agonists is likely to cause a loss of Ca2+from the SR and to contribute to the negative inotropic effects of these agents reported by other investigators. Our results support this notion that the negative inotropic effects of DHP agonists result in part from direct alteration in the activity of RyRs.
Role of leukocytes in uterine hypoperfusion and fetal growth retardation induced by ischemia-reperfusion
Authors: Miyakoshi Kei , Ishimoto Hitoshi , Nishimura Osamu , Tanigaki Shinji , Tanaka Mamoru , Miyazaki Toyohiko , Natori Michiya , Yoshimura Yasunori
Published: December 2017

We investigated leukocyte involvement in uterine hypoperfusion and intrauterine fetal growth retardation (IUGR) induced by ischemia-reperfusion (I/R) in Sprague-Dawley rats. On day 17 of gestation, leukocyte accumulation in the uterus and placenta subjected to 30 min of ischemia, followed by reperfusion, was assessed by measuring myeloperoxidase (MPO) activity. Uterine MPO activity was significantly higher after 1 h of reperfusion than it was before ischemia ( P < 0.05), without any increase in placental MPO activity. Immunohistochemical staining showed leukocyte accumulation in the uterus subjected to I/R. The effects of treatment with monoclonal antibodies against CD11a (WT1) and CD18 (WT3) at a dose of 0.8 mg/kg on uterine blood flow and IUGR were investigated. Laser-Doppler flowmetry demonstrated that uterine hypoperfusion at 2 h after ischemia (blood flow, −51.7 ± 1.2%; P < 0.01) was inhibited by WT1 and WT3 treatment. I/R-induced IUGR at full term ( P < 0.05 vs. nonischemic horn) was prevented by WT1 and WT3 treatment on day 17. These results indicate that leukocyte accumulation may play an important role in the pathogenesis of uterine hypoperfusion and IUGR induced by I/R in pregnant rats.
Signal transduction of flumazenil-induced preconditioning in myocytes
Authors: Yao Zhenhai , McPherson Bradley C. , Liu Huiping , Shao Zuohui , Li Changqing , Qin Yimin , Vanden Hoek Terry L. , Becker Lance B. , Schumacker Paul T.
Published: December 2017

The objective of this study was to examine the role of oxygen radicals, protein kinase C (PKC), and ATP-sensitive K+(KATP) channels in mediating flumazenil-produced preconditioning. Chick cardiomyocyte death was quantified using propidium iodide, and oxygen radical generation was assessed using 2′,7′-dichlorofluorescin oxidation. Preconditioning was initiated with 10 min of ischemia followed by 10 min of reoxygenation. Alternatively, flumazenil was infused for 10 min and removed 10 min before ischemia. Flumazenil (10 μM) and preconditioning increased oxygen radicals [1,693 ± 101 ( n = 3) and 1,567 ± 98 ( n = 3), respectively, vs. 345 ± 53 ( n = 3) in control] and reduced cell death similarly [22 ± 3% ( n = 5) and 18 ± 2% ( n = 6), respectively, vs. controls 49 ± 5% ( n = 8)]. Protection and increased oxygen radicals by flumazenil were abolished by pretreatment with the antioxidant thiol reductant 2-mercaptopropionyl glycine (800 μM; 52 ± 10%, n = 6). Specific PKC inhibitors Go-6976 (0.1 μM) and chelerythrine (2 μM), given during ischemia and reoxygenation, blocked flumazenil-produced protection (47 ± 5%, n = 6). The PKC activator phorbol 12-myristate 13-acetate (0.2 μM), given during ischemia and reoxygenation, reduced cell death similarly to that with flumazenil [17 ± 4% ( n = 6) and 22 ± 3% ( n = 5)]. Finally, 5-hydroxydecanoate (1 mM), a selective mitochondrial KATPchannel antagonist given during ischemia and reoxygenation, abolished the protection of flumazenil and phorbol 12-myristate 13-acetate. Thus flumazenil mimics preconditioning to reduce cell death in cardiomyocytes. Oxygen radicals activate mitochondrial KATPchannels via PKC during the process.
Determinants and interindividual variation of R-R interval dynamics in healthy middle-aged subjects
Authors: Pikkujämsä Sirkku M. , Mäkikallio Timo H. , Airaksinen K. E. Juhani , Huikuri Heikki V.
Published: December 2017

Determinants and intersubject variations of fractal and complexity measures of R-R interval variability were studied in a random population of 200 healthy middle-aged women (age 51 ± 6 yr) and 189 men (age 50 ± 6 yr) during controlled conditions in the supine and sitting positions. The short-term fractal exponent (α1) was lower in women than men in both the supine (1.18 ± 0.20 vs. 1.12 ± 0.17, P < 0.01) and sitting position ( P < 0.001). Approximate entropy (ApEn), a measure of complexity, was higher in women in the sitting position (1.16 ± 0.17 vs. 1.07 ± 0.19, P < 0.001), but no gender-related differences were observed in ApEn in the supine position. Fractal and complexity measures were not related to any other demographic, laboratory, or lifestyle factors. Intersubject variations in a fractal measure, α1 (e.g., 1.15 ± 0.20 in the supine position, z value 1.24, not significant), and in a complexity measure, ApEn (e.g., 1.14 ± 0.18 in the supine position, z value 1.44, not significant), were generally smaller and more normally distributed than the variations in the traditional measures of heart rate variability (e.g., standard deviation of R-R intervals 49 ± 21 ms in the supine position, z value 2.53, P < 0.001). These results in a large random population sample show that healthy subjects express relatively little interindividual variation in the fractal and complexity measures of heart rate behavior and, unlike the traditional measures of heart rate variability, they are not related to lifestyle, metabolic, or demographic variables. However, subtle gender-related differences are also present in fractal and complexity measures of heart rate behavior.
Calmodulin levels are dynamically regulated in living vascular smooth muscle cells
Authors: Hulvershorn Justin , Gallant Cynthia , Wang C.-L. Albert , Dessy Chantal , Morgan Kathleen G.
Published: December 2017

The total unbound calmodulin (i.e., not bound to target proteins) level in living smooth muscle cells from the ferret portal vein was monitored with a low-affinity, calmodulin-binding peptide tagged with an environmentally sensitive fluorophore. GS17C, a previously characterized peptide, from the calmodulin-binding domain of caldesmon was tagged with iodoacetyl nitrobenz-2-oxa-1,3-diazole (NBD) or, as a negative control, with iodoacetylfluorescein isothiocyanate. Increases in NBD-GS17C fluorescence were detected by using confocal microscopy when chemically loaded cells were stimulated with solutions of elevated [K+] or the calcium ionophore 4-bromoA-23187 to elicit increases in intracellular Ca2+concentration ([Ca2+]i) quantified by fura 2. Increases in peptide fluorescence were detected in response to a phorbol ester in the absence of changes in [Ca2+]i. These changes were blocked by the addition of the calmodulin antagonist calmidazolium. These results suggest that the total unbound intracellular calmodulin levels may be sufficient to regulate the activity of caldesmon and, furthermore, that phosphorylation of protein kinase C substrates may increase the level of available calmodulin in living smooth muscle cells.
AE anion exchangers in atrial tumor cells
Authors: Papageorgiou Panos , Shmukler Boris E. , Stuart-Tilley Alan K. , Jiang Lianwei , Alper Seth L.
Published: December 2017

Intracellular pH homeostasis and intracellular Cl−concentration in cardiac myocytes are regulated by anion exchange mechanisms. In physiological extracellular Cl−concentrations, Cl−/HCO[Formula: see text] exchange promotes intracellular acidification and Cl−loading sensitive to inhibition by stilbene disulfonates. We investigated the expression of AE anion exchangers in the AT-1 mouse atrial tumor cell line. Cultured AT-1 cells exhibited a substantial basal Na+-independent Cl−/HCO[Formula: see text] (but not Cl−/OH−) exchange activity that was inhibited by DIDS but not by dibenzamidostilbene disulfonic acid (DBDS). AT-1 cell Cl−/HCO[Formula: see text] activity was stimulated two- to threefold by extracellular ATP and ANG II. AE mRNAs detected by RT-PCR in AT-1 cells included brain AE3 (bAE3), cardiac AE3 (cAE3), AE2a, AE2b, AE2c1, AE2c2, and erythroid AE1 (eAE1), but not kidney AE1 (kAE1). Cultured AT-1 cells expressed AE2, cAE3, and bAE3 polypeptides, which were detected by immunoblot and immunocytochemistry. An AE1-like epitope was detected by immunocytochemistry but not by immunoblot. Both bAE3 and cAE3 were present in intact AT-1 tumors. Cultured AT-1 cells provide a useful system for the study of mediators and regulators of Cl−/HCO[Formula: see text] exchange activity in an atrial cell type.
Overexpression of cardiac I-κBα prevents endotoxin-induced myocardial dysfunction
Authors: Haudek Sandra B. , Spencer Erika , Bryant Debora D. , White D. Jean , Maass David , Horton Jureta W. , Chen Zhijian J. , Giroir Brett P.
Published: December 2017

Nuclear factor-κ B (NF-κB) is an inducible transcription factor that regulates expression of many genes, such as tumor necrosis factor-α (TNF-α), which may contribute to myocardial dysfunction. We investigated whether cardiac NF-κB activation is involved in the development of myocardial dysfunction after lipopolysaccharide (LPS) challenge. Mice were intraperitoneally injected with LPS, and the hearts were harvested and assayed for NF-κB translocation. After LPS challenge, NF-κB activation was detected within 30 min and remained for 8 h. In transgenic mice constitutively overexpressing a nondegradable form of I-κBα (I-κBαΔN) in cardiomyocytes, myocardial NF-κB translocation was prevented after LPS challenge. Myocytes isolated from these transgenics secreted significantly less TNF-α than did wild-type cardiomyocytes after LPS stimulation. When whole hearts were excised, perfused in a Langendorff preparation, and challenged with endotoxin, I-κBαΔN transgenic hearts displayed normal cardiac function, whereas profound contractile dysfunction was observed in wild-type hearts. These data indicate that myocardial NF-κB translocates within minutes after LPS administration. Inhibition of myocyte NF-κB activation by overexpression of myocyte I-κBα is sufficient to block cardiac TNF-α production and prevent cardiac dysfunction after LPS challenge.
Increased uncoupling proteins and decreased efficiency in palmitate-perfused hyperthyroid rat heart
Authors: Boehm Ernest A. , Jones Barney E. , Radda George K. , Veech Richard L. , Clarke Kieran
Published: December 2017

The physiological role of mitochondrial uncoupling proteins (UCPs) in heart and skeletal muscle is unknown, as is whether mitochondrial uncoupling of oxidative phosphorylation by fatty acids occurs in vivo. In this study, we found that UCP2 and UCP3 protein content, determined using Western blotting, was increased by 32 and 48%, respectively, in hyperthyroid rat heart mitochondria. Oligomycin-insensitive respiration rate, a measure of mitochondrial uncoupling, was increased in all mitochondria in the presence of palmitate: 36% in controls and 71 and 100% with 0.8 and 0.9 mM palmitate, respectively, in hyperthyroid rat heart mitochondria. In the isolated working heart, 0.4 mM palmitate significantly lowered cardiac output by 36% and cardiac efficiency by 38% in the hyperthyroid rat heart. Thus increased mitochondrial UCPs in the hyperthyroid rat heart were associated with increased uncoupling and decreased myocardial efficiency in the presence of palmitate. In conclusion, a physiological effect of UCPs on fatty acid oxidation has been found in heart at the mitochondrial and whole organ level.
Role of nitric oxide in the cerebrovascular and thermoregulatory response to interleukin-1β
Authors: Monroy Manuel , Kuluz John W. , He Dansha , Dietrich W. Dalton , Schleien Charles L.
Published: December 2017

Central administration of interleukin-1β (IL-1β) increases cerebral blood flow (CBF) and body temperature, in part, through the production of prostaglandins. In previous studies, the temporal relationship between these effects of IL-1β have not been measured. In this study, we hypothesized that the increase in CBF occurs before any change in brain or body temperature and that the cerebrovascular and thermoregulatory effects of IL-1β would be attenuated by inhibiting the production of nitric oxide (NO). Adult male rats received 100 ng intracerebroventricular (icv) injection of IL-1β, and cortical CBF (cCBF) was measured by laser-Doppler in the contralateral cerebral cortex. A central injection of IL-1β caused a rapid increase in cCBF to 133 ± 12% of baseline within 15 min and to an average of 137 ± 12% for the remainder of the 3-h experiment. Brain and rectal temperature increased by 0.4 ± 0.2 and 0.5 ± 0.2°C, but not until 45 min after IL-1β administration. Pretreatment with N ω-nitro-l-arginine methyl ester (l-NAME; 5 mg/kg iv) completely prevented the changes in cCBF and brain and rectal temperature induced by IL-1β.l-Arginine (150 mg/kg iv) partially reversed the effects ofl-NAME and resulted in increases in both cCBF and temperature. These findings suggest that the vasodilatory effects of IL-1β in the cerebral vasculature are independent of temperature and that NO plays a major role in both the cerebrovascular and thermoregulatory effects of centrally administered IL-1β.
Platelet hyperactivity and abnormal Ca2+homeostasis in diabetes mellitus
Authors: Li Yun , Woo Vincent , Bose Ratna
Published: December 2017

We sought to determine the mechanisms for hyperactivity and abnormal platelet Ca2+homeostasis in diabetes. The glycosylated Hb (HbA1c) level was used as an index of glycemic control. Human platelets were loaded with Ca- green-fura red, and cytosolic Ca2+([Ca2+]i) and aggregation were simultaneously measured. In the first series of experiments, the platelets from diabetic and normal subjects were compared for the ability to release Ca2+or to promote Ca2+influx. A potent and relatively specific inhibitor of Na+/Ca2+exchange, 5-(4-chlorobenzyl)-2′,4′-dimethylbenzamil (CB-DMB), increased the second phase of thrombin-induced Ca2+response, suggesting that the Na+/Ca2+exchanger works in the forward mode to mediate Ca2+efflux. In contrast, in the platelets from diabetics, CB-DMB decreased the Ca2+response, indicating that the Na+/Ca2+exchanger works in the reverse mode to mediate Ca2+influx. In the second series of experiments we evaluated the direct effect of hyperglycemia on platelets in vitro. We found that thrombin- and collagen-induced increases in [Ca2+]iand aggregation were not acutely affected by high glucose concentrations of 45 mM. However, when the platelet-rich plasma was incubated with a high glucose concentration at 37°C for 24 h, the second phase after thrombin activation was inhibited by CB-DMB. In addition, collagen-stimulated [Ca2+]iresponse and aggregation were also increased. Thus in diabetes the direction and activity of the Na+/Ca2+exchanger is changed, which may be one of the mechanisms for the increased platelet [Ca2+]iand hyperactivity. Prolonged hyperglycemia in vitro can induce similar changes, suggesting hyperglycemia per se may be the factor responsible for the platelet hyperactivity in diabetes.
Nonnoradrenergic mechanism of reflex cutaneous vasoconstriction in men
Authors: Stephens Dan P. , Aoki Ken , Kosiba Wojciech A. , Johnson John M.
Published: December 2017

We tested for a nonnoradrenergic mechanism of reflex cutaneous vasoconstriction with whole body progressive cooling in seven men. Forearm sites (<1 cm2) were pretreated with: 1) yohimbine (Yoh; 5 mM id) to antagonize α-adrenergic receptors, 2) Yoh plus propranolol (5 mM Yoh-1 mM PR id) to block α- and β-adrenergic receptors, 3) iontophoretic application of bretylium tosylate (BT) to block all sympathetic vasoconstrictor nerve effects, or 4) intradermal saline. Skin blood flow was measured by laser Doppler flowmetry and arterial pressure by finger photoplethysmography; cutaneous vascular conductance (CVC) was indexed as the ratio of the two. Whole body skin temperature (TSK) was controlled at 34°C (water-perfused suit) for 10 min and then lowered to 31°C over 15 min. During cooling, vasoconstriction was blocked at BT sites ( P > 0.05). CVC at saline sites fell significantly beginning at TSK of 33.4 ± 0.01°C ( P <0.05). CVC at Yoh-PR sites was significantly reduced beginning at TSK of 33.0 ± 0.01°C ( P < 0.05). After cooling, iontophoretic application of norepinephrine (NE) confirmed blockade of adrenergic receptors by Yoh-PR. Because the effects of NE were blocked at sites showing significant reflex vasoconstriction, a nonnoradrenergic mechanism in human skin is indicated, probably via a sympathetic cotransmitter.
Regulation of B2-kinin receptors by glucose in vascular smooth muscle cells
Authors: Christopher Julie , Velarde Victoria , Zhang Da , Mayfield Douglas , Mayfield Ronald K. , Jaffa Ayad A.
Published: December 2017

The development of vascular disease is accelerated in hyperglycemic states. Vascular injury plays a pivotal role in the progression of atherosclerotic vascular disease in diabetes, which is characterized by increased vascular smooth muscle cell (VSMC) proliferation and extracellular matrix accumulation. We previously reported that diabetes alters the activity of the kallikrein-kinin system and results in the upregulation of kinin receptors in the vessel wall. To determine whether glucose can directly influence the regulation of kinin receptors, the independent effect of high glucose (25 mM) on B2-kinin receptors (B2KR) in VSMC was examined. A threefold increase in B2KR protein levels and a 40% increase in B2KR surface receptors were observed after treatment with high glucose after 24 h. The mRNA levels of B2KR were also significantly increased by high glucose as early as 4 h later. To elucidate the cellular mechanisms by which glucose regulates B2KR, we examined the role of protein kinase C (PKC). High glucose increased total PKC activity and resulted in the translocation of conventional PKC isoforms (β1and β2), novel (ε), and atypical (ζ) PKC isoforms into the membrane. Inhibition of PKC activity prevented the increase in B2KR levels induced by ambient high glucose. These findings provide the first evidence that glucose regulates the expression of B2receptors in VSMC and provide a rationale to further study the interaction between glucose and kinins on the pathogenesis of atherosclerotic vascular disease in diabetes.
Aortic depressor nerve unmyelinated fibers in spontaneously hypertensive rats
Authors: Fazan Valéria Paula Sassoli , Salgado Helio Cesar , Barreira Amilton Antunes
Published: December 2017

The objective of the present study was to compare the morphology of the unmyelinated fibers in the aortic depressor nerves (ADN) of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). In anesthetized rats, the ADN was identified by its spontaneous activity synchronous with the arterial pulses. Thin sections of the proximal and distal segments of the ADN were analyzed by electron microscopy, and a morphometric study of the unmyelinated fibers and Schwann cells was performed. The proximal segments of WKY and SHR ADN contain an average of 335 ± 68 and 130 ± 14 unmyelinated fibers, respectively ( P < 0.05), and the distal segments contain an average of 337 ± 46 and 242 ± 77 unmyelinated fibers, respectively ( P < 0.05). The distribution of the diameters of unmyelinated fibers was unimodal for both strains, with the histogram from the SHR significantly shifted to the left. Because the unmyelinated fibers play a role in the tonic inhibition of the medullary vasomotor centers, especially in the presence of hypertension, the morphological differences observed in the ADN from SHR may account, at least in part, for the blunted baroreflex of SHR.
Assembly and activation of HK-PK complex on endothelial cells results in bradykinin liberation and NO formation
Authors: Zhao Yongqiang , Qiu Qianyao , Mahdi Fakhri , Shariat-Madar Zia , Røjkjær Rasmus , Schmaier Alvin H.
Published: December 2017

Prekallikrein (PK) activation on human umbilical endothelial cells (HUVEC) presumably leads to bradykinin liberation. On HUVEC, PK activation requires the presence of cell-bound high-molecular-weight kininogen (HK) and Zn2+. We examined the Zn2+requirement for HK binding to and the consequences of PK activation on endothelial cells. Optimal HK binding (14 pmol/106HUVEC) is seen with no added Zn2+in HEPES-Tyrode buffer containing gelatin versus 16–32 μM added Zn2+in the same buffer containing bovine serum albumin. The affinity and number of HK binding sites on HUVEC are a dissociation constant of 9.6 ± 1.8 nM and a maximal binding of 1.08 ± 0.26 × 107sites/cell (means ± SD). PK is activated to kallikrein by an antipain-sensitive mechanism in the presence of HK and Zn2+on HUVEC, human microvascular endothelial cells, umbilical artery smooth muscle cells, and bovine pulmonary artery endothelial cells. Simultaneous with kallikrein formation, bradykinin (5.0 or 10.3 pmol/106HUVEC in the absence or presence of lisinopril, respectively) is liberated from cell-bound HK. Liberated bradykinin stimulates the endothelial cell bradykinin B2 receptor to form nitric oxide. Assembly and activation of PK on endothelial cells modulates their physiological activities.
Oxygen supply and oxidative phosphorylation limitation in rat myocardium in situ
Authors: Kreutzer Ulrike , Mekhamer Yousry , Chung Youngran , Jue Thomas
Published: December 2017

The1H-NMR signal of the proximal histidyl-NδH of deoxymyoglobin is detectable in the in situ rat myocardium and can reflect the intracellular Po2. Under basal normoxic conditions, the cellular Po2is sufficient to saturate myoglobin (Mb). No proximal histidyl signal of Mb is detectable. On ligation of the left anterior descending coronary artery, the Mb signal at 78 parts/million (ppm) appears, along with a peak shoulder assigned to the corresponding signal of Hb. During dopamine infusion up to 80 μg · kg−1· min−1, both the heart rate-pressure product (RPP) and myocardial oxygen consumption (MV˙o2) increase by about a factor of 2. Coronary flow increases by 84%, and O2extraction (arteriovenous O2difference) rises by 31%. Despite the increased respiration and work, no deoxymyoglobin signal is detected, implying that the intracellular O2level still saturates MbO2, well above the Po2at 50% saturation of Mb. The phosphocreatine (PCr) level decreases, however, during dopamine stimulation, and the ratio of the change in Piover PCr (ΔPi/PCr) increases by 0.19. Infusion of either pyruvate, as the primary substrate, or dichloroacetate, a pyruvate dehydrogenase activator, abolishes the change in ΔPi/PCr. Intracellular O2supply does not limit MV˙o2, and the role of ADP in regulating respiration in rat myocardium in vivo remains an open question.
Endothelial microtubule disruption blocks flow-dependent dilation of arterioles
Authors: Sun Dong , Huang An , Sharma Sansar , Koller Akos , Kaley Gabor
Published: December 2017

The cytoskeleton is believed to have an important role in the structural and functional integrity of endothelial cells. The role of the endothelial cytoskeleton, specifically microtubules, in the mediation of flow-induced dilation of arterioles has not yet been studied. Thus the aim of our study was to investigate the role of microtubules in the endothelial mechanotransduction of flow-induced dilation of isolated gracilis arterioles of the rat. The active diameter of arterioles at a constant perfusion pressure (80 mmHg) was ∼63 μm, whereas their passive diameter (Ca2+-free solution) was ∼119 μm. At a constant pressure, increases in flow of the perfusate solution (from 0 to 10 and from 10 to 20 μl/min) elicited increases in diameter up to ∼95 μm (∼53% increase). Intraluminal administration of nocodazole at concentrations of 5 × 10−9and 5 × 10−8M had no discernible effects on the structure of endothelial microtubules or on flow-induced dilation, whereas it disassembled microtubules and eliminated flow-induced dilation at a concentration of 5 × 10−7M. At this higher concentration, however, the basal diameter and dilations to acetylcholine (10−8M), sodium nitroprusside (10−7M), arachidonic acid (5 × 10−6M), and prostaglandin E2(10−8M) were unaffected. Colchicine (5 × 10−7M) also disassembled microtubules and eliminated flow-induced dilation. We concluded that, in isolated arterioles, the integrity of the endothelial cytoskeleton is essential for the transduction of the shear stress signal that results in the release of endothelial factors evoking dilation.
Oxidative burst and NO generation as initial response to ischemia in flow-adapted endothelial cells
Authors: Manevich Yefim , Al-Mehdi Abu , Muzykantov Vladimir , Fisher Aron B.
Published: December 2017

Shear stress modulates endothelial physiology, yet the effect(s) of flow cessation is poorly understood. The initial metabolic responses of flow-adapted bovine pulmonary artery endothelial cells to the abrupt cessation of flow (simulated ischemia) was evaluated using a perfusion chamber designed for continuous spectroscopy. Plasma membrane potential, production of reactive O2species (ROS), and intracellular Ca2+and nitric oxide (NO) levels were measured with fluorescent probes. Within 15 s after flow cessation, flow-adapted cells, but not cells cultured under static conditions, showed plasma membrane depolarization and an oxidative burst with generation of ROS that was inhibited by diphenyleneiodonium. EGTA-inhibitable elevation of intracellular Ca2+and NO were observed at ∼30 and 60 s after flow cessation, respectively. NO generation was decreased in the presence of inhibitors of NO synthase and calmodulin. Thus flow-adapted endothelial cells sense the altered hemodynamics associated with flow cessation and respond by plasma membrane depolarization, activation of NADPH oxidase, Ca2+influx, and activation of Ca2+/calmodulin-dependent NO synthase. This signaling response is unrelated to cellular anoxia.
Relating myocardial laminar architecture to shear strain and muscle fiber orientation
Authors: Arts T. , Costa K. D. , Covell J. W. , McCulloch A. D.
Published: December 2017

Cardiac myofibers are organized into laminar sheets about four cells thick. Recently, it has been suggested that these layers coincide with the plane of maximum shear during systole. In general, there are two such planes, which are oriented at ±45° to the main principal strain axes. These planes do not necessarily contain the fiber axis. In the present study, we explicitly added the constraint that the sheet planes should also contain the muscle fiber axis. In a mathematical analysis of previously measured three-dimensional transmural systolic strain distributions in six dogs, we computed the planes of maximum shear, adding the latter constraint by using the also-measured muscle fiber axis. Generally, for such planes two solutions were found, suggesting that two populations of sheet orientation may exist. The angles at which the predicted sheets intersected transmural tissue slices, cut along left ventricular short- or long-axis planes, were strikingly similar to experimentally measured values. In conclusion, sheets coincide with planes of maximum systolic shear subject to the constraint that the muscle fiber axis is contained in this plane. Sheet orientation is not a unique function of the transmural location but occurs in two distinct populations.
PKC-β is not necessary for cardiac hypertrophy
Authors: Roman Brian B. , Geenen David L. , Leitges Michael , Buttrick Peter M.
Published: December 2017

Studies in human and rodent models have shown that activation of protein kinase C-β (PKC-β) is associated with the development of pathological hypertrophy, suggesting that ablation of the PKC-β pathway might prevent or reverse cardiac hypertrophy. To explore this, we studied mice with targeted disruption of the PKC-β gene (knockout, KO). There were no detectable differences in expression or distribution of other PKC isoforms between the KO and control hearts as determined by Western blot analysis. Baseline hemodynamics were measured using a closed-chest preparation and there were no differences in heart rate and arterial or left ventricular pressure. Mice were subjected to two independent hypertrophic stimuli: phenylephrine (Phe) at 20 mg · kg−1 · day−1 sq infusion for 3 days, and aortic banding (AoB) for 7 days. KO animals demonstrated an increase in heart weight-to-body weight ratio (Phe, 4.3 ± 0.6 to 6.1 ± 0.4; AoB, 4.0 ± 0.1 to 5.8 ± 0.7) as well as ventricular upregulation of atrial natriuretic factor mRNA analogous to those seen in control animals. These results demonstrate that PKC-β expression is not necessary for the development of cardiac hypertrophy nor does its absence attenuate the hypertrophic response.
Effects of body temperature during exercise training on myocardial adaptations
Authors: Harris M. Brennan , Starnes Joseph W.
Published: December 2017

This study determined the role of body temperature during chronic exercise on myocardial stress proteins and antioxidant enzymes as well as functional recovery after an ischemic insult. Male Sprague-Dawley rats were exercised for 3, 6, or 9 wk in a 23°C room (3WK, 6WK, and 9WK, respectively) or in a 4–8°C environment with wetted fur (3WKC, 6WKC, and 9WKC, respectively). The colder room prevented elevations in core temperature. During weeks 3–9 the animals ran 5 days/wk up a 6% grade at 20 m/min for 60 min. Myocardial heat shock protein 70 (HSP 70) increased 12.3-fold ( P < 0.05) in 9WK versus sedentary (SED) rats but was unchanged in the cold-room runners. Compared with SED rats, αB-crystallin was 90% higher in 9WKC animals, HSP 90 was 50% higher in 3WKC and 6WKC animals, and catalase was 23% higher in 3WK animals ( P < 0.05 for all). Cytosolic superoxide dismutase increased and mitochondrial SOD decreased ( P < 0.05) in 3WK and 6WK rats compared with 3WKC and 6WKC rats. Antioxidant enzymes returned to SED values in all runners by 9 wk. No differences were observed among any of the groups for glucose-regulated protein 75, heme oxygenase-1, or glutathione peroxidase. Mechanical recovery of isolated working hearts after 22.5 min of global ischemia was enhanced in 9WK ( P < 0.05) but not in 9WKC rats. We conclude that exercise training results in dynamic changes in cardioprotective proteins over time which are influenced by core temperature. In addition, cardioprotection resulting from chronic exercise appears to be due to increased HSP 70.
Overexpression of Bcl-2 attenuates apoptosis and protects against myocardial I/R injury in transgenic mice
Authors: Chen Zhongyi , Chua Chu Chang , Ho Ye-Shih , Hamdy Ronald C. , Chua Balvin H. L.
Published: December 2017

To test whether the antiapoptotic protein Bcl-2 prevents apoptosis and injury of cardiomyocytes after ischemia-reperfusion (I/R), we generated a line of transgenic mice that carried a human Bcl-2 transgene under the control of a mouse α-myosin heavy chain promoter. High levels of human Bcl-2 transcripts and 26-kDa Bcl-2 protein were expressed in the hearts of transgenic mice. Functional recovery of the transgenic hearts significantly improved when they were perfused as Langendorff preparations. This protection was accompanied by a threefold decrease in lactate dehydrogenase (LDH) released from the transgenic hearts. The transgenic mice were subjected to 50 min of ligation of the left descending anterior coronary artery followed by reperfusion. The infarct sizes, expressed as a percentage of the area at risk, were significantly smaller in the transgenic mice than in the nontransgenic mice (36.6 ± 5 vs 69.9 ± 7.3%, respectively). In hearts subjected to 30 min of coronary artery occlusion followed by 3 h of reperfusion, Bcl-2 transgenic hearts had significantly fewer terminal deoxynucleodidyl-transferase nick-end labeling-positive or in situ oligo ligation-positive myocytes and a less prominent DNA fragmentation pattern. Our results demonstrate that overexpression of Bcl-2 renders the heart more resistant to apoptosis and I/R injury.
Anesthetic inhibition in ischemic and nonischemic murine heart: comparison with conscious echocardiographic approach
Authors: Takuma Shin , Suehiro Kotaro , Cardinale Carol , Hozumi Takeshi , Yano Hideaki , Shimizu Juichiro , Mullis-Jansson Samantha , Sciacca Robert , Wang Jie , Burkhoff Daniel , Di Tullio Marco R. , Homma Shunichi
Published: December 2017

It is well known that the level of anesthesia obtained by intraperitoneal injection is variable and may alter cardiac function. In this study, we compared the effects of different anesthetics on cardiac function with the conscious state using high-resolution two-dimensional echocardiography in nonischemic and ischemic mice. Eighty-four mice were tested before and after surgery with ligation of the coronary artery. All 84 mice were studied in the conscious state and under high-dose intraperitoneal anesthesia. Twenty-two of 84 mice were studied under low-dose intraperitoneal anesthesia. Another 22 mice were also studied under gas anesthesia and spontaneous breathing. Experiments in the conscious state were performed by two investigators before the administration of anesthesia: one investigator held the animal and the transducer and the other operated the ultrasound equipment. Left ventricular systolic function was measured, and measurements obtained after surgery were compared with infarcted areas assessed by histological staining. Results showed that both high- and low-dose intraperitoneal anesthesia significantly reduced heart rates and left ventricular contractility in both pre- and postsurgical mice as opposed to conscious mice ( P < 0.01). There were significantly higher correlation coefficients between mean fractional area change (FAC) and infarcted area in conscious state compared with high-dose intraperitoneal anesthesia ( P< 0.05). The correlation coefficient between FAC and infarcted area during gas anesthesia was also significantly higher compared with high-dose intraperitoneal anesthesia ( P < 0.05). In conclusion, conscious experiments or the use of gas anesthesia is preferred for echocardiographic assessment of cardiac function in mice because intraperitoneal injection significantly induces a significant reduction in heart rate and left ventricular systolic function.
NO formation in nucleus tractus solitarii attenuates pressor response evoked by skeletal muscle afferents
Authors: Li Jianhua , Potts Jeffrey T.
Published: December 2017

We have previously shown that static muscle contraction induces the expression of c-Fos protein in neurons of the nucleus tractus solitarii (NTS) and that some of these cells were codistributed with neuronal NADPH-diaphorase [nitric oxide (NO) synthase]-positive fibers. In the present study, we sought to determine the role of NO in the NTS in mediating the cardiovascular responses elicited by skeletal muscle afferent fibers. Static contraction of the triceps surae muscle was induced by electrical stimulation of the L7 and S1 ventral roots in anesthetized cats. Muscle contraction during microdialysis of artificial extracellular fluid increased mean arterial pressure (MAP) and heart rate (HR) 51 ± 9 mmHg and 18 ± 3 beats/min, respectively. Microdialysis ofl-arginine (10 mM) into the NTS to locally increase NO formation attenuated the increases in MAP (30 ± 7 mmHg, P < 0.05) and HR (14 ± 2 beats/min, P > 0.05) during contraction. Microdialysis ofd-arginine (10 mM) did not alter the cardiovascular responses evoked by muscle contraction. Microdialysis of N G-nitro-l-arginine methyl ester (2 mM) during contraction attenuated the effects ofl-arginine on the reflex cardiovascular responses. These findings demonstrate that an increase in NO formation in the NTS attenuates the pressor response to static muscle contraction, indicating that the NO system plays a role in mediating the cardiovascular responses to static muscle contraction in the NTS.
Properties of smooth muscle hyperpolarization and relaxation to K+ in the rat isolated mesenteric artery
Authors: Dora Kim A. , Garland Christopher J.
Published: December 2017

Smooth muscle membrane potential and tension in rat isolated small mesenteric arteries (inner diameter 100–200 μm) were measured simultaneously to investigate whether the intensity of smooth muscle stimulation and the endothelium influence responses to exogenous K+. Variable smooth muscle depolarization and contraction were stimulated by titration with 0.1–10 μM phenylephrine. Raising external K+ to 10.8 mM evoked correlated, sustained hyperpolarization and relaxation, both of which were inhibited as the smooth muscle depolarized and contracted to around −38 mV and 10 mN, respectively. At these higher levels of stimulation, raising the K+ concentration to 13.8 mM still hyperpolarized and relaxed the smooth muscle. Relaxation to endothelium-derived hyperpolarizing factor, released by ACh, was not altered by the level of stimulation. In endothelium-denuded arteries, the concentration-relaxation curve to K+ was shifted to the right but was not depressed. In denuded arteries, relaxation to K+ was unaffected by the extent of prior stimulation and was blocked with 0.1 mM ouabain but not with 30 μM Ba2+. The ability of K+ to stimulate simultaneous hyperpolarization and relaxation in the mesenteric artery is consistent with a role as an endothelium-derived hyperpolarizing factor activating inwardly rectifying K+ channels on the endothelium and Na+-K+-ATPase on the smooth muscle cells.
EDHF mediates flow-induced dilation in skeletal muscle arterioles of female eNOS-KO mice
Authors: Huang An , Sun Dong , Carroll Mairead A. , Jiang Houli , Smith Carolyn J. , Connetta Joseph A. , Falck John R. , Shesely Edward G. , Koller Akos , Kaley Gabor
Published: December 2017

Vasodilation to increases in flow was studied in isolated gracilis muscle arterioles of female endothelial nitric oxide synthase (eNOS)-knockout (KO) and female wild-type (WT) mice. Dilation to flow (0–10 μl/min) was similar in the two groups, yet calculated wall shear stress was significantly greater in arterioles of eNOS-KO than in arterioles of WT mice. Indomethacin, which inhibited flow-induced dilation in vessels of WT mice by ∼40%, did not affect the responses of eNOS-KO mice, whereas miconazole and 6-(2-proparglyoxyphenyl)hexanoic acid (PPOH) abolished the responses. Basal release of epoxyeicosatrienonic acids from arterioles was inhibited by PPOH. Iberiotoxin eliminated flow-induced dilation in arterioles of eNOS-KO mice but had no effect on arterioles of WT mice. In WT mice, neither Nω-nitro-l-arginine methyl ester nor miconazole alone affected flow-induced dilation. Combination of both inhibitors inhibited the responses by ∼50%. 1 H-[1,2,4]oxadiazolo[4,3- a]quinoxalin-1-one (ODQ) alone inhibited flow-induced dilation by ∼49%. ODQ + indomethacin eliminated the responses. Thus, in arterioles of female WT mice, nitric oxide and prostaglandins mediate flow-induced dilation. When eNOS is inhibited, endothelium-derived hyperpolarizing factor substitutes for nitric oxide. In female eNOS-KO mice, metabolites of cytochrome P-450, via activation of large-conductance Ca2+-activated K+channels of smooth muscle, mediate entirely the arteriolar dilation to flow.