bluuebunny/crossref_metadata_2025 huggingface dataset:
Nutritional Interventions in Skeletal Muscle Atrophy
Authors: Crissey Jacqueline Marie , Farrar Roger P.
Published: June 2021

Skeletal muscle atrophy occurs with prolonged bed rest, limb immobilization, microgravity, physical inactivity, and advancing age. This study evaluated the efficacy of a carbohydrate‐protein (+ leucine) supplement (Pacific Health Laboratory) to reduce muscle mass loss and maintain muscle physiological function in a rat atrophy model of hindlimb unloading (HU). Rats were fed high (CPH) and low (CPL) doses of the supplement or vehicle (HOH) (n=10/group) and were suspended by their tails to remove all weight‐bearing function on their hindlimbs for 28 days. The supplement (5 ml/kg) or vehicle (H2O) was administered twice daily via gavage, excluding the naïve control (CON) group (n=10); all rats were fed rat chow ad libitum. On day 28 in‐situ force measurements of the left lateral gastrocnemius were recorded in all HU groups (CPL, CPH, and HOH). Statistical analyses utilized ANOVA and the Dunnett's method. Muscle weights of the soleus (−36%), plantaris (−16%), and gastrocnemius (−21%) were significantly reduced in HOH, CPL, and CPH rats after 28 days of HU (p<0.05 compared to CON). The nutritional treatment (CPL and CPH) did not reduce muscle mass loss induced by HU. Specific in‐situ tetanic tension (maximal force/cross‐sectional area) of the left lateral gastrocnemius muscle was significantly less in the HOH group (17.2 N/cm2) compared to CPL (20.2 N/cm2, p<0.05) and CON groups (20.4 N/cm2, p<0.05). The specific tension of the CPL group did not differ significantly from the CON group. The CPH group's specific tension (19.2 N/cm2) did not significantly differ from either the HOH or CON groups. This carbohydrate‐protein (+ leucine) nutritional supplement was able to maintain muscle function (specific tension) but not muscle mass. Pacific Health Cooperation and The University of Texas at Austin supported this research.
The effects of Xin shRNA with cardiotoxin injury on skeletal muscle in vivo
Authors: Labatia Rita , Hawke Thomas J
Published: June 2021

Xin, an actin binding protein, has been implicated in the regulation of satellite cells during skeletal muscle regeneration. It is believed to exert effects on the cytoskeleton through interactions with other structural proteins like filamin c, f‐actin and desmin. In order to investigate the role of Xin in muscle regeneration following injury, we injected Xin shRNA adenovirus (XinAd) into the right tibialis anterior (R‐TA) of male C57B6 mice while the left tibialis anterior (L‐TA) served as a control. Muscle injury in both legs was induced 4 days later using a cardiotoxin injection. As expected, XinAd reduced endogenous Xin expression by 67.8 ± 7.4% at 0 days. Surprisingly, at 3 days Xin Ad injected muscle displayed 29.3 ± 8.1% increase in filamin c, 12.6 ± 5.6% increase in f‐actin and a 41.2 ± 12.9% decrease in desmin expression compared to the L‐TA. Histochemically, the treatment and control tissues illustrated similar signs of degradation and regeneration at all time points. Therefore, while silencing Xin modifies actin‐associated protein expression patterns, it appears that other factors may compensate for its disappearance in terms of whole muscle regeneration.
TWEAK augments matrix metallanoproteinease‐9 expression in skeletal muscle cells through the activation of p38 mitogen‐activated protein kinase and nuclear factor‐kappa B signaling pathways
Authors: Kumar Ashok , Tyagi Suresh C. , Srivastava Daya S.
Published: June 2021

Skeletal muscle atrophy is a debilitating complication of several chronic disease states such as diabetes, sepsis, chronic heart failure, and cancer. We have recently identified TNF‐related weak inducer of apoptosis (TWEAK) as a novel and potent skeletal‐muscle wasting cytokine. We hypothesize that one of the mechanisms by which TWEAK induces skeletal muscle myopathy is through the increased production of extracellular matrix (ECM)‐degrading enzymes. To test this hypothesis, we have performed a genome‐wide microarray analysis using RNA isolated from control and TWEAK‐treated cultured C2C12 myotubes. Among others, we have identified matrix metallanoproteinease‐9 (MMP‐9), as one of the highly up‐regulated ECM protease in TWEAK‐treated C2C12 myotubes. The increased expression of MMP‐9 in myotubes in response to TWEAK was confirmed by quantitative PCR. Treatment of myotubes with TWEAK dose‐dependently increased the production of MMP‐9 in culture supernatants. TWEAK activated mitogen‐activated protein kinase (MAPK) and nuclear‐factor‐kappa B (NF‐κB) signaling pathways in myotubes. Furthermore, inhibition of p38MAPK or NF‐κB blocked the TWEAK‐induced MMP‐9 production. Collectively, our data suggest that p38 MAPK and NF‐κB constitute the biochemical signaling pathways that lead to the increased production of MMP‐9 in myotubes in response to TWEAK. This work was supported by RO1 grant AG029623.
Effect of IGF‐I and rapamycin on diaphragm muscle atrogenes with malnutrition
Authors: Fournier Mario , Da Xiaoyu , Lewis Michael I
Published: June 2021

We recently reported that the addition of rapamycin to IGF‐I decreased P‐Akt in the diaphragm (DIA) with acute nutritional deprivation (ND). As Akt impacts on pathways mediating proteolysis, the aim was to evaluate the influence of IGF‐I and rapamycin on rat DIA atrogenes with ND. Groups: 1) Controls (CTL); 2) ND (20% of food intake for 4 days); 3) ND+IGF‐I; and 4) ND+IGF‐I+rapamycin. DIA mRNA was analyzed by real‐time PCR. Foxo1 and 3a increased 2.3 and 2.9‐fold with ND, while Foxo4 decreased by 46%. IGF‐I reduced Foxo1 and 3a to CTL levels, while Foxo4 increased 2.5‐fold. Rapamycin negated all IGF‐I effects. ND increased MAFbx/Atrogin1 and MuRF1 by 2.7 and 19‐fold, with no change in E3‐[alpha]II. IGF‐I decreased these atrogenes to near CTL levels. Rapamycin returned these E3 ligases to ND levels. ND increased caspase 3 (important in myofibrillar disassembly) 4.8‐fold. IGF‐I decreased it to 1.5‐fold CTL. Rapamycin increased abundance to ND levels. ND increased myostatin (negative regulator of muscle mass) 2.4‐fold. IGF‐I reduced it to 0.78 CTL levels while rapamycin negated IGF‐I effects. We conclude that IGF‐I significantly reduces atrogenes and their regulators with ND. These positive effects were reversed with rapamycin, an mTOR inhibitor. We speculate that influences on mTOR feedback negatively on Akt and/or other regulators of pathways mediating protein degradation. Funded by NIH HL‐071227
The muscle carnosine and taurine content of elderly non‐obese korean males with impaired glucose tolerance
Authors: Harris Roger C , Kim Hyo J , Jones Glenys , Lee Jon S , Kim Chang K , Wise John A
Published: June 2021

Carnosine (β‐Alanylhistidine) (Carn) occurs in high concentrations in muscle where it contributes to H+ buffering during exercise. Synthesis is limited by the availability of β‐Alanine synthesised in the liver from Uracil and transported to muscle fibres, but is enhanced by the inclusion of β‐Ala (or a β‐Ala source such as Carn) in the diet. Whilst the transport of β‐Ala into muscle is still not fully understood there is evidence that this may be linked to the action of insulin. Taurine (Tau), a beta‐sulfonic amino acid which shares the same transporter as β‐Ala, was also investigated in this study.

PURPOSE

To investigate if muscle Carn or Tau are altered in patients with impaired glucose tolerance (IGT).

METHOD

A single muscle biopsy of the vastus lateralis was obtained from 8 elderly non‐obese Korean males (58–67 y) with IGT. ~2mg of freeze‐dried powdered muscle was analysed for Carn and Tau by HPLC. Results were compared to muscle values of 10 young (19–25 y) active Korean males determined at the same time by the same method.

RESULTS

The mean muscle Carn and Tau contents of IGT subjects (21.5±3.8 and 44.9±12.0 mmol.kg−1 dm) were very similar to contents of younger subjects (22.1±2.8 and 42.8±10.3 mmol.kg−1 dm). Carn showed the same negative correlation to Tau in IGT as in younger subjects (a relationship previously observed in other subjects).

CONCLUSION

Muscle Carn and Tau were unaffected by IGT or age in Korean non‐obese males.
Ca2+‐ and Sr2+‐activated contractile characteristics of permeabilized muscle fibers from dystrophic dogs
Authors: Lynch Gordon S , Murphy Kate T , Bizario Joao CS , Costa Maria CR , Pinto Mildre L , van der Poel Chris
Published: June 2021

The Golden Retriever muscular dystrophy (GRMD) dog has a mutation in the dystrophin gene, suffers muscle wasting and weakness, and dies prematurely from cardiac or pulmonary failure. It is a relevant model for understanding the underlying mechanisms and investigating potential therapeutic approaches for Duchenne muscular dystrophy (DMD). Unfortunately, few GRMD colonies exist worldwide and there is a lack of information regarding aspects of muscle fiber function between dystrophic and healthy dogs. Our aim was to identify Ca2+‐ and Sr2+‐activated contractile characteristics of single permeabilized fibers from the biceps femoris muscles of 2‐year old GRMD dogs. Single fibers were attached to a force transducer, activated in Ca2+‐ and Sr2+‐buffered solutions to generate force‐pCa(pSr) relations, and then analyzed for myosin heavy chain and troponin‐C isoform composition. Specific force was ~25% lower in dystrophic dogs compared with litter mate controls (215 ± 18 kN/m2, n = 15 vs 160 ± 9 kN/m2, n = 15), but fiber sensitivity to Ca2+ was unchanged. A high proportion (~65%) of fibers from dystrophic dogs were ‘hybrid’ fibers as evident from their biphasic force‐pSr relations and combination of slow and fast troponin‐C isoforms. These functional assessments provide important information about the progression of muscular dystrophy in the GRMD model and its relevance to DMD.

Supported by the Muscular Dystrophy Association (USA)
Eccentric contractions protect skeletal muscle from temperature‐induced contracture in Y522S RyR1 knock‐in mice
Authors: Ingalls Christopher Paul , Corona Benjamin T. , Rouviere Clement
Published: June 2021

Mice with a malignant hyperthermia mutation (Y522S) of the ryanodine receptor (RyR1) can exhibit muscle contracture and die when exposed to elevated temperature. When stressed by elevated temperature (T), Ca2+ leak from the Y522S RyR1 and cellular accumulation is thought to produce reactive nitrogen species that modifies RyR1 resulting in sustained Ca2+ release and muscle contracture. Eccentric contractions are known to reduce muscle strength by impairing voltage‐gated Ca2+ release from RyR1. The purpose of this study was to determine if eccentric contraction‐induced injury would reduce contractures when EDL muscle from Y522S mice is incubated at 35°C. Anterior crural muscles from wild‐type (WT) and Y522S mice were injured by performing 150 eccentric contractions in vivo. EDL muscle function was assessed via force‐frequency (F‐F) experiments at 30 and 35°C 3 and 14 d post‐injury. At 30°C, uninjured EDL from Y522S mice exhibited stable resting force during the F‐F experiment, and peak force at 300 Hz (Po) was not different than WT mice. At 35°C, increases in resting force in Y522S mice at 3 (38%) and 14 (152%) d after injury were less than increases in uninjured muscle (322%). T‐induced decreases in Po in Y522S mice at 3 (34%) and 14 (36%) d were less than in uninjured muscle (65%). Eccentric contractions attenuate T‐induced contractures and reductions in active force in Y522S mice.

Supported by NIH grant AR41802
Changes in junctophilin 1 contribute to strength deficits after eccentric contraction‐induced muscle injury
Authors: Corona Benjamin , Wilson Andrea K , Rouviere Clement , Hamilton Susan L , Ingalls Christopher P
Published: June 2021

Performance of eccentric contractions (ECCs) disrupts excitation‐contraction (E‐C) coupling and induces strength deficits. E‐C uncoupling may result from disturbance to the SR / t‐tubule triad structure. Junctophilin 1 (JP1) is a triadic protein that is thought to be required for normal voltage‐gated SR Ca++ release. We hypothesize that ECCs alter tibialis anterior (TA) muscle JP1 protein content and that functional recovery from ECCs corresponds to recovery of JP1. To test the hypothesis we compared strength and JP1 changes after ECCs in wild type (WT; n =12) and malignant hyperthermia susceptible (Y522S; n =12) mice that recover function after ECCs faster than WT mice. The left anterior crural muscles (primarily TA muscle) of Y522S and WT mice performed 150 ECCs and a pre‐ and 3 or 14d post‐injury torque‐frequency test, in vivo. JP1: tubulin protein content was determined in injured and uninjured TA muscles. 3d post‐injury, torque and JP1 were reduced similarly in both WT and Y522S mice. 14d post‐injury, torque and JP1 were recovered in Y522S mice, but were still reduced in WT mice. Regression analysis indicated that reductions of JP1 are strongly associated with torque deficits (r = 0.70). These results indicate that JP1 is reduced following ECCs and suggest that recovery of JP1 contributes to the restoration of prolonged torque deficits in TA muscle.

‐Supported by NIH grant R01 AR41802 (S.L. Hamilton)
Myonuclear apoptosis occurs in regenerative process following eccentric contractions in rats
Authors: Sudo Mizuki , Kano Yutaka
Published: June 2021

Eccentric contractions (ECC) induce myofibrillar collapse, edema and inflammation. Although apoptosis of myonuclei following ECC are activated at inflammatory phase, the apoptosis response of regenerative phase remains to be elucidated. The aim of present study was to determine the inflammatory and regenerative phase of the apoptosis responses induced by ECC. In anesthetized rats, the tibialis anterior muscles were subjected to 40 times‐repeated ECC, which was evoked by surface electric stimulation (100Hz, 10V) with mechanical muscle stretch. The apoptosis was examined control, 1, 3, 7 and 14 d after ECC (n = 4). Identified TUNEL‐positive myonuclei were assessed by triple labeling with dystrophin staining, TUNEL assay and DAPI. At 1 and 3 d, focal edema and necrotic fibers invaded by mononuclear phagocytes were recognized. The regenerated fibers with central nucleus were detected at 7 and 14 d. TUNEL‐positive myonuclei were significantly increased at 7 and 14 d compared with control (7 d:7.0 ± 1.5%, 14 d:3.8 ± 1.1%, P < 0.05, respectively, % of total myonuclei, mean ± SE). Further we found that myonuclear apoptosis was located in subsarcolemma rather than central nucleus at 7 and 14 d. In conclusion, apoptosis responses in myocyte generate not only inflammatory phase but also regenerative process after ECC.
TRPA1 and TRPV3 Agonists Increase Intracellular Ca2+ Levels in Native Cerebral Artery Endothelial Cells
Authors: Earley Scott , Gonzales Albert L. , Crnich Rachael
Published: June 2021

Increases in endothelial cell [Ca2+] initiate the production of vasodilatory factors such as nitric oxide, prostaglandin, and epoxyeicosatrienoic acids. The molecular identities of the ion channels responsible for endothelial cell Ca2+ entry are not well defined. We examined the hypothesis that members of the transient receptor potential (TRP) superfamily of cation channels serve this function. Using ratiometric Ca2+ imaging, we examined the effects of the TRPA1 agonist allyl isothiocyanate (AI) and the combined TRPA1/TRPV3 agonists eugenol (EG) and carvacrol (CV) on the intracellular [Ca2+] of freshly dispersed native endothelial cells from rat cerebral arteries. All three agonists stimulated increases in endothelial cell [Ca2+], however, CV was much more potent (EC50 = 34 μM) than either AI (EC50 = 400 μM) or EG (EC50 = 2.3 mM). None of these agonists stimulated increases in endothelial cell [Ca2+] when extracellular Ca2+ was not present in the bathing solution, suggesting that Ca2+ influx is required for this response. Additional experiments demonstrated that CV and AI both caused concentration‐dependent dilation of intact, pre‐constricted cerebral arteries. Our findings indicate that stimulation of TRPA1 and/or TRPV3 activity elicits increases in endothelial cell [Ca2+] and cerebral artery dilation, possibly by promoting the production of vasodilatory factors. Supported by AHA0535226N.
Lysophosphatidic acid upregulates vascular endothelial growth factor‐C expression in human endothelial cells and enhances lymphangiogenesis
Authors: Lin Chi‐Iou , Chen Chiung‐Nien , Huang Miao‐Tzu , Lee Shyh‐Jye , Lee Hsinyu
Published: June 2021

Lysophosphatidic acid (LPA) is a small‐molecular‐weight lipid growth factor which binds to G‐protein‐coupled receptors and activates a variety of physiological functions. Previous studies demonstrated that LPA enhances cell proliferation and migration, activation of proteases, and expression of inflammatory‐related genes in human endothelial cells. Furthermore, LPA regulates expression of angiogenic factors such as VEGF‐A in cancer cells which leads to tumor angiogenesis. These results suggested that LPA is an important regulator of vessel functions; however, the role of LPA on lymphatic vessel formation and lymphangiogenesis has not been studied. VEGF‐C has been shown to regulate the development and growth of the lymphatic system. In this study, we showed that LPA upregulated VEGF‐C mRNA expression in human umbilical vascular endothelial cells and subsequent endothelial cell tube formation in vitro and in vivo. These enhancement effects were LPA1‐ and LPA3‐dependent and required COX‐2, EGFR transactivation and activation of NF‐κB. LPA‐induced endothelial tubes expressed the lymphatic marker, Prox‐1. Our results suggest that LPA regulates lymphangiogenesis through the induction of VEGF‐C expression in endothelial cells. This study provides some basic information that LPA might be a novel target for therapeutics against lymphangiogenesis and tumor metastasis.
Effect of fatty acids on (ECV‐304) endothelial cell function
Authors: Curi Rui , Portioli Silva Erica Paula , de Lima Thais Martins , Reis Silveira Leonardo dos
Published: June 2021

Endothelium inflammation is the key step for atheroma formation. Plasma fatty acid (FA) levels are increased in conditions (i.e. obesity and type‐II diabetes) in which there are high occurrence of atherosclerosis. Up to now, however, there is no report concerning the direct effect of FA on endothelium function such as production of NO and ROS, release of pro‐inflammatory cytokines, and endothelial cell death. The purpose of this study was to compare the effects of highly atherogenic (saturated) with poorly atherogenic (mono and polyunsaturated) FAs on endothelium inflammation. We investigated the effect of various FAs [palmitic (16:0)‐ PA, stearic (18:0)‐ SA, oleic (C18:1)‐ OA, linoleic (18:2)‐ LA, eicosapentaenoic (20:5)‐ EPA, and docosahexaenoic (22:6)‐ DHA acids] on ECV‐304 endothelial cell line in culture. The following parameters were investigated: NFκB activation, release of pro‐inflammatory cytokines (IL‐6, IL‐8, TNF‐α), production of NO and ROS and cell death. Evidence was obtained that saturated and unsaturated fatty acids exerted toxic effects on ECV‐304 cells. However, the cytotoxic effects of PA and SA on endothelial cells were markedly more pronounced than those of OA, LA, EPA and DHA. EPA, SA and PA exerted an inhibitory effect on NFκB activation. Production of inflammatory cytokines, IL‐6 and TNF‐α, was not changed by treatment with all FA tested. PA, SA, DHA and EPA caused a marked decrease of IL‐8 production. These findings may consist the basic mechanisms that underlay the high occurrence of atherosclerosis in populations under saturated fat‐rich diet. This research was supported by FAPESP, CNPq and CAPES
IL‐20 causes tube formation and spheroid sprouting in human lymphatic endothelial cells
Authors: Dissing Steen , Hammer Troels , Ditlev Sisse B , Hübschmann Martin V , Hansen Anker J , Tritsaris Katerina
Published: June 2021

IL‐20 is an arteriogenic cytokine that remodels collateral networks in vivo and in particular plays a role in cellular organization (Tritsaris et al., PNAS, 104, 15364–15369, 2007). We investigated its role in lymphangiogenesis using an immortalized lymphatic endothelial cell line, hTERT‐HDLEC, that express lymphatic markers and responds to VEGF‐C by forming tube like structures in matrigel. Upon stimulation of hTERT‐HDLEC with IL‐20 we found a rapid, transient increase in the intracellular, free Ca2+ concentration which was also observed with VEGF‐A but not with VEGF‐C. We also found that IL‐20 activates Akt and Erk1/2 phosphorylations within 10–60 minutes. In addition, the mTOR complex which is regulated by Akt signaling was phosphorylated within 5–30 min. In matrigels IL‐20 caused tube formations as well as morphological changes resulting in elongated cell structures. We inserted hTERT‐HDLEC into a collagen bed with 2 % FCS and found that both IL‐20 and VEGF‐C caused formation of spheroid sprouting within 24 hrs. These data show that IL‐20 activates signaling processes underlying changes in morphology and cell proliferation. Thus, IL‐20 is a cytokine that has the potential of activating or modulating the formation of lymphatic vessels. Its role in vivo could be in concert with other lymphangiogenic factors such as VEGF‐C and PDGF‐BB.
Replicative senescence of vascular endothelial cells isolated from coronary patients is worsened by oxidative stress associated with risk factors for cardiovascular disease
Authors: Voghel Guillaume , Thorin‐Trescases Nathalie , Farhat Nada , Mamarbachi Maya , Villeneuve Louis , Perrault Louis P , Carrier Michel , Thorin Eric
Published: June 2021

The p53‐p21 pathway can trigger senescence either by telomere shortening (replicative senescence via ATM) or following exposure to oxidative stress and DNA damages (stress‐induced senescence, SIS, via caveolin‐1). Our objective was to determine the main pathway in endothelial cells (EC) from coronary patients exposed to risk factors for cardiovascular diseases (CVD). EC were isolated from human internal mammary artery discarded during CABG (n = 56) and cultured in vitro until senescence. We quantified senescence by cytochemical detection of senescence‐associated b‐galactosidase, telomere length (RFL) by Southern blotting, gene expression of p53, p21, ATM and caveolin‐1 (Cav‐1) by qPCR and levels of lipid peroxidation (HNE) by immunofluorescence. Senescent EC displayed both telomeric instability markers (RFL shortening and increase in ATM, p<0.05) and oxidative stress markers (HNE and Cav‐1, p<0.05). mRNA levels of p53 were directly associated with the DNA damage response protein ATM and p21 (p<0.05) but not with Cav‐1 or HNE (p>0.05), suggesting that senescence is not exclusively directed by SIS. In conclusion, our data suggest that in human EC exposed to risk factors for CVD, there is a significant interplay between replicative senescence and SIS due to a state of cumulative cell damages. This study was supported by the Canadian Institutes of Health Research and the Fonds de la Recherche en Santé du Québec.
Hypoxic Vasodilation in Human Skeletal Muscle: contributions of nitric oxide and vasodilating prostaglandins
Authors: Markwald Rachel R , Kirby Brett S , Carlson Rick E , Voyles Wyatt F , Dinenno Frank A
Published: June 2021

We tested the hypothesis that nitric oxide (NO) and prostaglandins (PGs) contribute independently to hypoxic vasodilation, and that combined inhibition would reveal a potential compensatory interaction between these two dilators in the regulation of vascular tone. In 12 healthy adults, we measured forearm blood flow (Doppler ultrasound) and calculated forearm vascular conductance (FVC) responses to 5 minutes of steady‐state isocapnic hypoxia (O2 saturation ∼85%). All trials were performed during α‐ and β‐adrenergic blockade (via brachial artery catheter) to eliminate sympathoadrenal influences on vascular tone. Hypoxic vasodilator responses were determined under control conditions, during independent NO synthase (NOS; L‐NAME) or cyclooxygenase inhibition (COX; ketorolac), and finally during combined NOS and COX inhibition. Drug order was counterbalanced across subjects. In both groups, acute hypoxia caused significant forearm vasodilation under control conditions (ΔFVC ∼25%; P<0.05), and this was not influenced by prior inhibition of NO or PGs. In contrast, combined inhibition of NO and PGs in both groups reduced hypoxic vasodilation by ∼90%. Our findings indicate that NO and PGs are not independently obligatory for regulating vascular tone during hypoxia, but that together they play a significant role in hypoxic vasodilation in human skeletal muscle.

Supported by AG‐022337, HL‐087952
Hereditability Defect in Red‐Cell K in Hypertension
Authors: Delgado‐Almeida Antonio , Delgado Maria Carolina
Published: June 2021

Background:

Following our description of an hereditability of a decreased erythrocyte K [K]er independent of cell Na, in essential hypertension (EH), a long‐term study of RBC (Ki, Nai, H2O) content, plasma (Na, K, Cl, ionized Ca++), trans‐tubular K gradient (TTKG), total body K (TBK) and body water distribution by Bio‐Impedance Analysis was carried out in 185 hypertensives (47±19 y, 57 females; BP 167±11/101±7 mm Hg).

Results:

All cases had low Ki (85.3±6 mmol/l vs 96.2±4 mmol/l in normal, p<0.001) despite normal (82%) or mild elevated Nai (7.2±1.3 mmol/l vs 6.2±1.1 mmol/l in normal, p=NS), despite similar urinary Na excretion (107±22 vs 101±35 mmol/l). Patients with lowest Ki (78.3±5 mmol/l) had lower urinary K 21±15 mmol/l), TTKG (4.2±0.7) and higher 12‐h urinary volume (≥1.3 ml/min) than those with higher Ki (86.5±6mmol/l, p<0.002), urinary K (39±11mmol/l, p 0.004), TTKG (7.2±1.3) but lower 12‐h urinary volume (≤0.85 ml/min). Reversal of this abnormal Ki content in hypertensives was associated with decline and better control of BP (82%), fasting plasma glucose and regression of ST‐T alterations related to LVH hypertrophy (71%) or coronary artery disease (61%). Such body K disturbances, recorded at different month periods, unrelated to age, sex, race or dietary K intake in the hypertensives, and present in half (48%) of their normotensive offspring were implying a defective K mechanism in erythrocyte‐progenitor cells.

Conclusion:

These observations strongly suggest a critical role of RBC in normal body K homeostasis that is disturbed in EH.
Role of Renal NAD(P)H Oxidase in Dahl Salt‐Sensitive Hypertension
Authors: Manning R. Davis , Tian Niu , Moore Rebecca S , Pryor Janelle S , Stockstill Rachel
Published: June 2021

The goal of this study was to determine the role of NAD(P)H oxidase in Dahl S hypertension. Rats were subjected to a 5‐week high or low Na diet with and without inhibition of NAD(P)H oxidase (apocynin [apo], 100mg/kg/d). The results after 5 weeks are shown below (values are mean ± SEM; † P<0.05 comparing Hi Na and Hi Na+apo groups; * P<0.05 compared to Low Na group). Compared to low Na intake, high Na intake caused marked increases in renal mRNA expression of NAD(P)H oxidase, arterial pressure, proteinuria and decreased renal hemodynamics. Apo treatment in Dahl S rats on high Na intake decreased renal NAD(P)H oxidase expression and arterial pressure, improved renal dysfunction, and attenuated renal injury. In conclusion, the renal NAD(P)H oxidase pathway may play an important role in causing renal oxidative stress and in the progressive increases in arterial pressure and renal damage and decreases in renal hemodynamics that occur in Dahl salt‐sensitive hypertension. Support NIH 51971.

Treatment

Hi Na

Hi Na+Apo

Low Na

Low Na+Apo

NADPH gp 91 mRNA (fold)

9.0±0.6† *

4.3± 0.8 *

1.0±0.2

1.7±0.3

NADPH p22 mRNA (fold)

5.8±1.8† *

2.1± 0.3

1.0±0.3

1.6±0.3

NADPH p47 mRNA (fold)

9.4±0.6† *

3.5± 0.6 *

1.0±0.2

1.5±0.2

NADPH p67 mRNA (fold)

7.8±1.2† *

2.9± 0.5

1.0 ± 0.2

1.2±0.2

Mean arterial pressure (mmHg)

183±3 † *

162± 4

103±2.4

102±1.5

GFR (ml/min)

1.9±0.1† *

2.8±0.1 *

3.3±0.25

3.3±0.1

Renal plasma flow (ml/min)

6.8± .5 † *

8.5± .3 *

11.6±.8

10.7± .5

Renal vascular resistance

53± 5 † *

36± 1 *

17±2

18±1

Proteinuria (mg/day)

174±12† *

137± 13

35± 6

37±7
Agonistic autoantibodies to the angiotensin II type 1 receptor increases Angiotensin II‐induced ET‐1 production
Authors: Roberts Lyndsay , Dukes Matthew P , Fournier Lillian , Dechend Ralf , Wallukat Gerd , LaMarca Babbette
Published: June 2021

Endothelial dysfunction plays an important role in development of hypertension during pregnancy. Endothelin‐1 (ET‐1) and agonistic autoantibodies to the angiotensin II (AngII) type I receptor (AT1‐AA) may be important links between endothelial dysfunction and hypertension during preeclampsia. Although it is clear that activation of the Ang II type 1 receptor (AT1R) increases endothelial cell production of ET‐1, it is unknown whether interaction of the AT1‐AA with the AT1R enhances AngII‐induced ET‐1 production. Therefore, we tested the hypothesis that binding of the AT1‐AA to the AT1R increased ET‐1 secretion from endothelial cells. To achieve this goal we examined the effect of AngII on ET‐1 production in the presence and absence of the AT1‐AA. HUVEC were pretreated with serum free media, AngII (10‐7mol/L), 1:100 AT1‐AA isolated from preeclamptic women, or AT1‐AA+AngII for six hours. ET‐1 was measured by ELISA and normalized to total protein/flask. Basal ET‐1 production increased from 19.3 +/− 1.7 to 45.4 +/− 14 pg/ml with AngII and to 32.9 +/− 2.3 pg/ml with the AT1‐AA alone. Interestingly, ET‐1 production increased markedly from 19.3 +/− 1.7 to 297.6 +/− 50 pg/ml in response to AngII in the presence of the AT1‐AA. This exaggerated response was abolished with an AT1R antagonist or an AT1‐AA specific blocking peptide. These findings support the hypothesis that AT1‐AA increases AngII‐induced ET‐1 production.
Leptin icv blunts the renal sympatho‐inhibition of volume expansion (VE) in normal and fat fed Wistar rats
Authors: Johns Edward J. , Devoy Louise , Huang Chunlong
Published: June 2021

This study investigated the impact of leptin icv on the renal sympatho‐inhibition arising from stimulation of the cardiopulmonary reflex in normal and high fat fed rats. Male Wistar rats, at 7 weeks of age, were fed either a normal (3.4% fat, n=6) or a fat diet (45% fat, n=4) for 4 weeks. The rats were anaesthetised and cannulae placed in the femoral artery and vein, electrodes sealed onto the left renal nerve and a cannula implanted into the right lateral cerebroventricle. VE, 30 min of saline iv at 0.25% of bwt/min, was performed before and 2 h after leptin was given icv at 5μg+10μg/h. Means ±SEM were subjected to ANOVA with significance at P<0.05. In normal Wistar rats, VE had no effect on blood pressure (BP), at 80±5mmHg, or heart rate (HR), at 384±23 bpm, but renal sympathetic nerve activity (RSNA), at 4.0±0.8 mV/s, fell by 51% (P<0.05). After 2h of leptin icv, VE decreased RSNA by 29% (P<0.05), which was less (P<0.05) than before leptin. In fat fed rats, BP was comparable to those on a normal diet but HR, at 433±9 bpm, and RSNA, at 11.4 mV/s, were significantly higher (both P<0.05), while VE reduced RSNA by 17.2% (P<0.05), which was smaller (P<0.05) than that in rats on a normal diet. Leptin icv did not change the response patterns of BP, HR and RSNA to VE.

The data showed that leptin within brain appeared to exert a tonic inhibition on the cardiopulmonary mediated suppression of RSNA.
Renal Tissue Oxygenation with Renal Arterial Stenosis
Authors: Warner Lizette , Gomez Sabas , Haas John A , Bolterman Rodney , Bentley Michael D , Joyner Michael J , Lerman Lilach O , Romero Juan Carlos
Published: June 2021

Functional renal oxygen consumption, VrO2 is due to tubular reabsorption of sodium (TNa) in the proximal (∼67%) and distal(∼30%) nephrons, resulting in compartments (cortex/medulla) with distinct VrO2/TNa relationships. A progressive drop in renal blood flow (RBF) with renal arterial stenosis (RAS), disrupts O2 delivery, vital for reabsorption. The overall objective for the study is to determine the intra‐renal regional alterations in tissue oxygenation (PtO2) and theVrO2/TNa relationship in acute RAS. The hypothesis is RAS is accompanied by intra‐renal regional changes in oxygenation. Unilateral acute RAS was induced in 8 pigs by an extra‐vascular balloon. RBF and PtO2 were measured continuously with an ultrasound flow probe during baseline, RBF autoregulation and 3 sequential decreases in RBF, recovery, and furosemide administration. VrO2 was determined using Fick arterio‐venous differences. There were concomitant decreases in VrO2, and PtO2 throughout the occlusion periods. Medullary PtO2 changes were 35% more pronounced than cortical. This study suggests medullary decreases in PtO2 are more aggravated than cortical changes with reductions in RBF, accentuating the medulla's vulnerability to progressive RAS. Mild to moderately severe obstructions seem to preserve the tubular transport costs.

Funding provided by NIH Grant (HL16496‐32), 2007 APS Porter Physiology Fellowship and the Mayo Foundation.
Factors influencing the contribution of myocardial laminar sheets to ventricular wall thickening
Authors: Coppola Benjamin , Covell James , Omens Jeffrey
Published: June 2021

The myocardium is composed of laminar sheets of myofibers (oriented at angle β from radial), which shear during systole and account for >50% of wall thickening (E33). The sheet strains (Ess, Enn, Esn) are related to wall thickening by E33 = Ess*cos2β + Enn*sin2β + 2*Esn*cosβ*sinβ. We hypothesized that the contribution of each term to wall thickening was determined primarily by anatomic structure, and independent of load. To test this hypothesis, we imaged transmural columns of beads implanted into the left ventricle of open‐chest dogs (n=15) using biplane cineradiography. Strains were expressed in terms of a histologically measured fiber‐sheet coordinate system. At 20% depth, the sheet extension (Ess) term was the primary contributor to end‐systolic wall thickening, with an average of 68%. At 52 and 81% depths, the sheet shear (Esn) term was the predominant factor, contributing 49 and 48% respectively. This transmural trend held during diastolic wall thinning (EDP from 3 to 18 mmHg), with Ess term = 78%, and Esn term = 79 and 127% corresponding to the 3 depths. Increasing afterload (LV pressure from 119 to 169 mmHg) also gave a similar transmural result. We conclude that the transmural deformation of the myocardial wall is a property of the local anatomical orientation, with laminar structures contribution similarly during diastole and systole with varying loads. Supported by NIH grant HL32583.
Continuous Cardiac Output Monitoring by Analysis of Peripheral Blood Pressure
Authors: Lee Kichang , Arai Tatsuya , Cohen Richard J.
Published: June 2021

The purpose of this study was to evaluate our novel technique for continuous cardiac output monitoring using mathematical analysis of peripheral blood pressure waveforms. As a gold standard measure of cardiac output, we used a surgically implanted Transonics aortic flow probe, which provides an instantaneous flow signal, around ascending aortic root of 9 Yorkshire swine (∼35 kg) during a deep plane of anesthesia with isoflurane. Two micromanometer‐tipped catheters and two external fluid‐filled catheter blood pressure transducers assessed blood pressure. Heart rate, arterial pressure, and cardiac output were changed simultaneously or independently by infusion of vaso‐active drugs (hetastarch, dobutamine, isoproterenol, nitroglycerine, phenylephrine, and esmolol), or hemorrhage. Significant changes in heart rate, mean blood pressure, and cardiac output were observed and used to test algorithms for monitoring cardiac output. The estimated cardiac output error was low (15%) and highly correlated with the measured cardiac output (R2= 0.83). Our new technique accurately estimates continuous cardiac output by mathematical analysis of the measured arterial blood pressure waveforms in the peripheral arterial tree.
Heme Oxygenase‐1 Attenuates Isoproterenol‐induced Cardiac Hypertrophy in vivo
Authors: Kinobe Robert T , Simpson Jeremy A , Brunt Keith R , Hall Sean R , Ballantyne Laurel L , Ogunyankin Kofo O , Ward Christopher , Melo Luis G
Published: June 2021

Heme oxygenase‐1 (HO‐1) is an oxidative stress responsive gene that is implicated in protection against many pathophysiological processes including ischemia reperfusion injury and heart failure. As elevated activity of the sympathetic nerve system is a common feature of heart failure, the present study was designed to investigate the role of HO‐1 in cardiac hypertrophy during chronic β‐adrenoceptor stimulation. Isoproterenol was administered to wild type, non‐transgenic or transgenic cardiac specific HO‐1 overexpressing (α‐MHC‐HO‐1) FVB mice for 15 or 30 days at a rate of 30 μg/g/day. Chronic isoproterenol infusion caused a significant increase in left ventricular cardiac mass in the non‐transgenic but not transgenic mice (mg/g heart to body weight ratio: sham 4.9 ± 0.4, isoproterenol 6.4 ± 1.3; and sham 5.9 ± 0.7, isoproterenol 5.8 ± 0.5 for non‐transgenic and transgenic mice, respectively). Isoproterenol‐induced cardiac hypertrophy was accompanied by an increase in ventricular end diastolic pressure (P < 0.05), myocardial fibrosis and superoxide generation as measured by NADH oxidase activity. These data suggest that HO‐1 alleviates oxidative stress and cardiac hypertrophy under excessive β‐adrenergic stimulation, and that overexpression of HO‐1 is beneficial for sympatho‐excitatory cardiac hypertrophy and failure in mice. This work was supported by the Heart and Stroke Foundation of Canada.
The role of the endocannabinoid system in cirrhotic cardiomyopathy
Authors: Batkai Sandor , Mukhopadhyay Partha , Pacher Pal , Kunos George
Published: June 2021

Advanced liver cirrhosis is associated with hyperdynamic circulation: hypotension, decreased peripheral resistance and cardiac dysfunction termed cirrhotic cardiomyopathy. Previous studies have revealed the role of endocannabinoids and CB1 receptors in the development of generalized hypotension in models of liver cirrhosis, and CB1 receptors have been also implicated in the decreased β‐adrenergic responsiveness of cirrhotic heart tissue. Here we document the cardiac contractile dysfunction in vivo in liver cirrhosis and explore the role of the endocannabinoid system in its development. Rats with CCl4‐induced cirrhosis developed low blood pressure, tachycardia and decreased cardiac contractility. CB1 antagonist AM251 acutely increased blood pressure as well as both load‐dependent and ‐independent indices of systolic function, whereas no such changes were elicited by AM251 in controls. Furthermore, tissue levels of the endocannabinoid anandamide increased in the heart of cirrhotic compared to control rats without any change in 2‐AG levels. CB1 receptor expression in the heart was unaffected by cirrhosis. Thus, activation of cardiac CB1 receptors by endogenous anandamide contributes to the reduced cardiac contractility in liver cirrhosis, and CB1 receptor antagonists may be used to improve contractile function in cirrhotic cardiomyopathy and, possibly, in other forms of heart failure.
Cardiac Function in Transgenic Mice with Calcineurin Inhibition
Authors: Gao Shumin , Gelpi Ricardo , Hong Chull , Shen Youtang , Zhai Peiyong , Dhar Sunil K. , Vatner Dorothy E. , Vatner Stephen F. , Sadoshima Junichi
Published: June 2021

The goal of this study was to determine whether left ventricular (LV) function is altered in Zaki4 transgenic (TG) mice, which inhibit calcineurin, at baseline and with chronic pressure overload (aortic banding). LV function was measured by pulse wave and tissue Doppler. At baseline, LV ejection fraction (EF) was not different in TG and wild type (WT) mice. However, diastolic function was already impaired in TG, as reflected by mitral inflow E/A ratio 1.46±0.07 VS 1.22±0.06, P<0.05. After 2‐weeks of aortic banding, LV/BW in TG mice was significantly less than that in WT mice (3.3±0.1 VS 4.0±0.2, P<0.05) due to calcineurin inhibition. However, not only E/A ratio, but also deceleration time (DT ms), IVRT (ms), pulsed Doppler Tei index and tissue Doppler Tei index were significantly impaired in TG, and LVEF was also decreased, P<0.05, TG 66±1% VS WT 70±1%. Thus, while inhibition of calcineurin in TG mice prevents the development of LV hypertrophy, it impairs LV diastolic function, which is evident even without pressure overload. With the stress of chronic pressure overload, the diastolic dysfunction is intensified and systolic dysfunction appears.
Improvement of cardiac contractile function by proteasome inhibition after pressure overload
Authors: Hedhli Nadia , Liu Huasheng , Madura Kiran , Hong Chull , Vatner Stephen F , Depre Christophe
Published: June 2021

The goal of this study was to determine if proteasome inhibition (PI) could regress pre‐existing hypertrophy and collagen accumulation. Mice were submitted to transverse aortic banding (B) for 3 weeks and compared to shams (S). Two weeks after B, mice were treated daily with the proteasome inhibitor epoxomicin (E, 0.5 mg/kg) or with the vehicle (V). After 3 weeks B, B‐V showed an increase in proteasome activity (PA: 347±19in B‐V vs.114±7 in S, P<0.05), and left ventricle/tibial length (LV/TL: 10.2±0.56 in B‐V vs.6.2±0.3 in S, P<0.05). B‐V mice developed signs of heart failure shown by a decreased ejection fraction (EF: 50±4 in B‐V vs. 75±2 in S P<0.05). Compared to the B‐V, B‐E mice showed significantly lower PA (85±5, p<0.05 vs. B‐V) and LV/TL (8.4±0.3, P<0.05 vs. B‐V), and a better EF (62±4, P<0.05 vs. B‐V). Because pressure overload activates the expression of collagen and matrix metalloproteases (MMP), we tested whether E would prevent this activation. Transcripts of collagen type 1 and MMP2 increased 4‐fold in B‐V versus sham (P<0.05), which was abolished in B‐E. As a result, accumulation of interstitial collagen after B was lower in B‐E versus B‐V (1.6±0.8 in B‐E vs. 9.0±0.5 in B‐V, P<0.05). Therefore, PI started after B rescues ventricular function and decreases myocyte hypertrophy and collagen accumulation.
High altitude physiology research and training platforms used by American physiologists during World War II: innovative altitude chambers and high flying bomber aircraft
Authors: Dean Jay B.
Published: June 2021

America's fledgling aviation medicine program had only 3 man‐rated chambers in 1940, located at Wright Field's Aero Medical Lab (AML: H. Armstrong, D.B. Dill & F.G. Hall), Mayo Clinic (W. Boothby & W.R. Lovelace), and Harvard School of Public Health (C. Drinker). This poor state of affairs was best exemplified when members of the Committee on Aviation Medicine inquired about using the elevators in the Empire State Building for physiological studies of “rapid” decompression. Eventually, research facilities for rapid decompression were established in the Physiology Department at The Ohio State University (Dr. F.A. Hitchcock). Numerous altitude chambers were set up across the US in the ensuing months, which were used to study the physiology of O2 want, decompression sickness, and explosive decompression. A Boeing B‐17 bomber (Nemesis of Aeroembolism), assigned to Wright Field's AML, flew above 40,000 feet to test flight gear and provide a research platform for physiological studies. The high altitude research program was complemented by an indoctrination program that reached its zenith in 1945: >200 physiologists, using 65 altitude chambers at 45 air fields, taught ∼58,000 men/month how to protect themselves during high altitude flight. The success of America's aviation medicine program contributed significantly to the allied victory in the air war and laid the foundation for aerospace medicine.
Purification and identification of positive regulators binding to a Novel element in the c‐jun promoter
Authors: Jiang Daifeng , Zhou YanWen , Jarrett Harry W.
Published: June 2021

A putative response element, GAGCCTC, was investigated to regulate c‐jun expression in this study. Electrophoretic mobility shift assays demonstrate that sequence‐specific binding proteins form a specific complex with this element in HEK293 cells. Additionally, unlabeled consensus AP‐1 element DNA, but not a similar NF‐jun element DNA, competes with complex formation. Mutations of this element decrease c‐jun promoter reporter activity by nearly 5‐fold in HEK293 cells. A new, two‐step oligonucleotide trapping technique was developed to purify the element binding proteins. LC‐Nanospray‐ESI‐MS/MS identification and western blotting analysis show that the purified complex contains Ku80, Ku70 and c‐jun, which was further confirmed by antibody supershift and immunoprecipitation analysis, and by southwestern blot and UV crosslinking analysis in vitro as well as chromatin immunoprecipitation in vivo. c‐Jun promoter activity decreases after Ku80 was significantly decreased by its siRNA, and c‐jun expression decreased at both the mRNA and protein levels. A mutant Ku80 with normal amino acid sequence but immune to the siRNA recovers c‐jun promoter activity from siRNA inhibition. Similarly, Ku70 wild type transfection can also up‐regulate c‐jun promoter activity. Thus, we speculate that Ku80‐Ku70‐c‐jun trimer activates c‐jun expression by binding to this GAGCCTC element in the c‐jun promoter.
Repair of CAG/CTG repeat loop in human cells
Authors: Li Guo‐Min , Hou Caixia , Chan Nelson , Gu Liya
Published: June 2021

Trinucleotide repeat (TNR) expansion is the genetic basis for certain neurological and neurodegenerative disorders, including Huntington's disease. Since TNR sequences, e.g., (CAG)n, form a loop or hairpin structure, DNA loop repair is considered a major mechanism for TNR stability. However, the molecular mechanisms of DNA loop repair and protein components involved in this pathway are poorly understood. Using a biochemical approach, where human nuclear extracts were examined for their ability to process a plasmid DNA containing a (CAG)25 or (CTG) 25 loop, we show here that repair of CAG or CTG loop in human cells occurs in a nick‐ or loop‐directed manner. The repair is initiated by an incision at the top of the loop, followed by additional incision and excision, limited repair DNA synthesis, and ligation. Several biochemical mutants derived from fractionations allow us to demonstrate that PCNA, RPA, and FEN1 are essential components involved in TNR loop repair. We also show that MutSβ, a mismatch recognition protein that was previously thought to promote CAG expansion in transgenic mouse model by binding to the CAG loop and inhibiting its repair, has no inhibitory effect on TNR loop repair in human extracts, suggesting that the transgenic animal model may not reflect TNR instability in human cells. Our findings provide molecular insight into the mechanism by which cells maintain TNR stability.
Illuminating the elementary steps of helicase translocation and unwinding via single molecule analysis
Authors: Ha Taekjip
Published: June 2021

The step sizes of DNA translocation and unwinding by helicases have been a subject of significant biophysics interest for some time. In principle, up to 10 base pairs (bp) of DNA can be unwound using the energy liberated by one ATP hydrolyzed. Translocation on single stranded DNA (ssDNA)has even less constraints. Past structural studies suggested the smallest step size of 1 nt translocation per ATP but functional studies suggested larger physical or kinetic step sizes ranging from 4 to 23 nt/bp translocation/unwinding. We have used single molecule fluorescence techniques to deduce the ssDNA translocation step size of PcrA helicase and unwinding step size of hepatitus C virus NS3 helicase. The former was made possible by the highly regular cycles of repetitive ssDNA looping coupled with PcrA translocation. In NS3, we found kinetic evidence that each 3 bp unwinding step requires approximately 3 hidden steps, presumably due to 1 ATP hydrolysis each. Our results reconcile the apparent discrepancies between the structural and biochemical data, and may prove general for many other helicases.