Cardiac tolerance to ischemia in neonatal spontaneously hypertensive rats

Hypertension is the risk factor of serious cardiovascular diseases, such as ischemic heart disease and atherosclerosis. The aim of the present study was to analyze the development of cardiac tolerance to ischemia in neonatal spontaneously hypertensive rats (SHR) and possible protective effect of ischemic preconditioning (IP) or adaptation to intermittent high-altitude hypoxia (IHAH). For this purpose we used 1- and 10-day-old pups of SHR and their normotensive control Wistar Kyoto rats (WKY). Isolated hearts were perfused in the Langendorff mode with Krebs-Henseleit solution at constant pressure, temperature and rate. Cardiac tolerance to ischemia was expressed as a percentage of baseline values of developed force (DF) after global ischemia. IP was induced by three 3-min periods of global ischemia, each separated by 5-min periods of reperfusion. IHAH was simulated in barochamber (8 h/day, 5000 m) from postnatal day 1 to 10. Cardiac tolerance to ischemia in 1-day-old SHR was higher than in WKY. In both strains tolerance decreased after birth, and the difference disappeared. The high cardiac resistance in 1- and 10-day-old SHR and WKY could not be further increased by both IP and adaptation to IHAH. It may be concluded that hearts from newborn SHR are more tolerant to ischemia/reperfusion injury as compared to age-matched WKY; cardiac resistance decreased in both strains during the first ten days, similarly as in Wistar rats.     

Metabolic effects of estrogen substitution in combination with targeted exercise training on the therapy of obesity in ovariectomized Wistar rats

Postmenopausal women tend to have a higher risk in developing obesity and thus metabolic syndrome. Recently we could demonstrate that physical activity and estrogen replacement are effective strategies to prevent the development of nutritional induced obesity in an animal model. The aim of this study was to determine the combined effects of estrogen treatment and exercise training on already established obesity. Therefore ovariectomized (OVX) and sham-operated (SHAM) female Wistar rats were exposed to a high fat diet for ten months. After this induction period obese SHAM and OVX rats either remained sedentary or performed treadmill training for six weeks. In addition OVX rats were treated with 17β-Estradiol (E(2)) alone, or in combination with training. Before and after intervention effects on lipid and glucose metabolism were investigated. Training resulted in SHAM and OVX rats in a significant decrease of body weight, subcutaneous and visceral body fat, size of adipocytes and the serum levels of leptin, cholesterol, low-density lipoprotein and triglycerides. In OVX animals E(2) treatment resulted in similar effects. Often the combination of E(2) treatment and training was most effective. Analysis of the respiratory quotient indicates that SHAM animals had a better fat burning capacity than OVX rats. There was a tendency that training in SHAM animals and E(2) treatment in OVX animals could improve this capacity. Analysis of glucose metabolism revealed that obese SHAM animals had higher glucose tolerance than OVX animals. Training improved glucose tolerance in SHAM and OVX rats, E(2) treatment in OVX rats. The combination of both was most effective. Our results indicate that even after a short intervention period of six weeks E(2) treatment and exercise training improve parameters related to lipid as well as glucose metabolism and energy expenditure in a model of already established obesity. In conclusion a combination of hormone replacement therapy and exercise training could be a very effective strategy to encourage the therapy of diet-induced obesity and its metabolic consequences in postmenopausal women.     

Calorie restriction prevents the development of insulin resistance and impaired insulin signaling in skeletal muscle of ovariectomized rats

Insulin resistance of skeletal muscle glucose transport due to prolonged loss of ovarian function in ovariectomized (OVX) rats is accompanied by other features of the metabolic syndrome and may be confounded by increased calorie consumption. In this study, we investigated the role of calorie consumption in the development of insulin resistance in OVX rats. In addition, we examined the cellular mechanisms underlying skeletal muscle insulin resistance in OVX rats. Female Sprague-Dawley rats were ovariectomized (OVX) or sham-operated (SHAM). OVX rats either had free access to food, pair feeding (PF) with SHAM or received a 35% reduction in food intake (calorie restriction; CR) for 12weeks. Compared with SHAM, ovariectomy induced skeletal muscle insulin resistance, which was associated with decreases (32-70%) in tyrosine phosphorylation of the insulin receptor and insulin receptor substrate-1 (IRS-1), IRS-1 associated p85 subunit of phosphatidylinositol 3-kinase (PI3-kinase), and Akt Ser(473) phosphorylation whereas insulin-stimulated phosphorylation of IRS-1 Ser(307), SAPK/JNK Thr(183)/Tyr(185), and p38 mitogen-activated protein kinase (MAPK) Thr(180)/Tyr(182) was increased (24-62%). PF improved the serum lipid profile but did not restore insulin-stimulated glucose transport, indicating that insulin resistance in OVX rats is a consequence of ovarian hormone deprivation. In contrast, impaired insulin sensitivity and defective insulin signaling were not observed in the skeletal muscle of OVX+CR rats. Therefore, we provide evidence for the first time that CR effectively prevents the development of insulin resistance and impaired insulin signaling in the skeletal muscle of OVX rats.     

Effect of estrogen on calcium-handling proteins, beta-adrenergic receptors, and function in rat heart

Regulation of cellular Ca(2+) cycling is central to myocardial contractile function. Loss of Ca(2+) regulation is associated with cardiac dysfunction and pathology. Estrogen has been shown to modify contractile function and to confer cardioprotection. Therefore, we investigated the effect of estrogen on expression of rat heart myocardial Ca(2+)-handling proteins and beta-adrenergic receptor (beta(1)-AR) and examined functional correlates. Female rats were sham-operated (SHAM) or ovariectomized. Two weeks after ovariectomy rats were injected (i.p.) daily with estradiol benozoate (OVX+EB) or sesame oil (OVX) for 2 weeks. Protein abundance was measured by immunoblotting and mRNA was quantified by real-time RT-PCR. OVX significantly decreased estrogen and progesterone levels and EB replacement returned both estrogen and progesterone to physiological levels. OVX induced a 75% reduction of uterine weight and a gain in body weight. Replacement restored weights to SHAM level. OVX increased and estrogen-replacement normalized abundance of beta(1)-AR and L-type Ca(2+) channel (Cav1.2) protein. OVX decreased sodium-Ca(2+) exchange protein (NCX) and estrogen restored protein abundance to SHAM levels. Sarcoplasmic reticular ATPase (SERCA), phospholamban (PLB), and ryanodine receptor (RyR) abundance was not altered by hormone status. Levels of mRNA encoding for beta(1)-AR, Cav1.2, and NCX were not influenced by OVX or estrogen replacement. OVX had no effect on SERCA and PLB mRNA level but estrogen replacement elicited a significant increase compared to OVX and SHAM. Estrogen-dependent changes in Ca(2+)-handling proteins and beta(1)-AR are theoretically consistent reduced myocellular Ca(2+) load. However, hormone-dependent alterations in protein were not associated with changes in contractile function.     

SHR的胰岛素抗性及骨胳肌的形态学特征

本文研究了自发性高血压大鼠及其对照品系－正常血压大鼠的胰岛素抗性和肌肉组织的形态学差异。结果表明：ＳＨＲ的基础胰岛素水平,糖耐量实验时胰岛素曲线下的面积均高于ＷＫＹ。而基础血糖水平,ＧＴＴ时血糖曲线下的面积两者相似。组织形态学研究表明：ＳＨＲ内胳肌中对胰岛素长一智Ⅰ型纤维比例低于ＷＫＹ。本研究发现：基础胰岛素水平或ＧＴＴ时胰岛素曲线下的面积与血压之间有正相关关系,而基础胰岛素水平与骨胳肌中Ｉ型纤维     

Protective effect of estrogens against oxidative damage to heart and skeletal muscle in vivo and in vitro

Estrogen has been shown to protect skeletal muscle from damage and to exert antioxidant properties. The purpose of the present study was to investigate the antioxidant and protective properties of estrogens in rodent cardiac and skeletal muscle and H9c2 cells. Female Sprague-Dawley rats were separated into three groups, ovariectomized (OVX), ovariectomized with estrogen replacement (OVX + E2), and intact control (SHAM), and were assessed at two time periods, 4 and 8 weeks. Rodents hearts were analyzed for basal and iron-stimulated lipid peroxidation in the absence and presence of beta-estradiol (betaE2) by measuring thiobarbituric acid reactive species (TBARS). Isolated soleus (SOL) and extensor digitorum longus (EDL) were analyzed for creatine kinase (CK) efflux. Using H9c2 cells, the in vitro effects of betaE2 and its isomer alpha-estradiol were investigated under glucose-free/hypoxic conditions. TBARS assay was also performed on the H9c2 in the presence or absence of betaE2. The results indicate that OVX rodent hearts are more susceptible to lipid peroxidation than OVX + E2 hearts. OVX soleus showed higher cumulative efflux of CK than OVX + E2. Furthermore, H9c2 survival during oxidative stress was enhanced when estrogen was present, and both OVX hearts at 4 weeks and H9c2 cells particularly were protected from oxidative damage by estrogens. We conclude that estrogen protects both skeletal and cardiac muscle from damage, and its antioxidant activity can contribute to this protection.     

Effects of acute and 2-Day genistein treatment on cardiac function and ischemic tolerance in ovariectomized rats

Background: Genistein, a naturally occurring isoflavonic phytoestrogen associated with reduced incidence of heart disease, may be a possible alternative treatment for postmenopausal women with heart disease.Objective: This study examined the effects of genistein on in vitro heart function and ischemic tolerance in ovariectomized (OVX) Sprague-Dawley rats.Methods: To examine the acute effects of genistein on cardiac function, isolated working hearts were perfused under aerobic conditions with increasing concentrations of genistein (10–150 µM). A separate group of OVX rats was used to assess ischemic tolerance: treated rats received genistein (250 mg/kg, dissolved in 200 uL dimethyl sulfoxide [DMSO]) injected once daily for 2 days, and control rats received DMSO only. After treatment, hearts were perfused for 30 minutes under aerobic conditions and then subjected to 20 minutes of global no-flow ischemia by clamping the preload and afterload lines, followed by 30 minutes of reperfusion.Results: Genistein was associated with improvements in mechanical function in OVX rat hearts (n = 5) with maximum increases in contractility (259 mm Hg/sec above baseline) and cardiac output (7 mL/min above baseline) observed with 30 μM of genistein (both, P < 0.05). Relative to baseline, genistein-treated hearts (n = 5) also had greater ischemic tolerance than did control hearts (n = 6) and significant improvements in mean (SEM) recovery of contractility (to 75.0% [9.7%] of preischemic function; P < 0.05) and cardiac output (to 48.8% [12.3%] of preischemic function; P < 0.05) after reperfusion. These effects occurred without significant changes in myocardial levels of nonprotein thiols or thiobarbituric acid reactive substances, although a reduction in mean glucose transporter protein 4 content (13.2% [2.7%]; P < 0.05) was observed in genistein-treated hearts. No significant changes in blood pressure were observed with genistein.Conclusions: Despite the lack of significant changes in physical characteristics, 2-day treatment with genistein was associated with significant cardioprotective effects in OVX rats, suggesting a potential therapeutic role in postmenopausal women.     

Obesity and altered glucose metabolism impact HDL composition in CETP transgenic mice: a role for ovarian hormones

Mechanisms underlying changes in HDL composition caused by obesity are poorly defined, partly because mice lack expression of cholesteryl ester transfer protein (CETP), which shuttles triglyceride and cholesteryl ester between lipoproteins. Because menopause is associated with weight gain, altered glucose metabolism, and changes in HDL, we tested the effect of feeding a high-fat diet (HFD) and ovariectomy (OVX) on glucose metabolism and HDL composition in CETP transgenic mice. After OVX, female CETP-expressing mice had accelerated weight gain with HFD-feeding and impaired glucose tolerance by hyperglycemic clamp techniques, compared with OVX mice fed a low-fat diet (LFD). Sham-operated mice (SHAM) did not show HFD-induced weight gain and had less glucose intolerance than OVX mice. Using shotgun HDL proteomics, HFD-feeding in OVX mice had a large effect on HDL composition, including increased levels of apoA2, apoA4, apoC2, and apoC3, proteins involved in TG metabolism. These changes were associated with decreased hepatic expression of SR-B1, ABCA1, and LDL receptor, proteins involved in modulating the lipid content of HDL. In SHAM mice, there were minimal changes in HDL composition with HFD feeding. These studies suggest that the absence of ovarian hormones negatively influences the response to high-fat feeding in terms of glucose tolerance and HDL composition. CETP-expressing mice may represent a useful model to define how metabolic changes affect HDL composition and function.     

Hormonal activation of the cGMP-inhibited low-Km cyclic AMP phosphodiesterase of rat adipocytes from different sites: influence of ovariectomy.

Hormonal activation of the cGMP-inhibited low Km cyclic AMP phosphodiesterase isoenzyme (cGI.PDE) by effectors, acting either through the cAMP-independent (insulin) or through cAMP-dependent (isoproterenol, forskolin ACTH and 8Br-cAMP) mechanisms, were compared in parametrial (PM) and femoral subcutaneous (SC) adipocytes from sham-operated (SHAM) and ovariectomized (OVX) rats. In SHAM rats, the basal cGI.PDE activity was 50% higher in PM than in SC adipocytes. In OVX rats, the cGi.PDE activatory responses to all the effectors tested remained unchanged in SC, but were completely suppressed in PM adipocytes. The mechanism underlying these defective cGI.PDE activatory responses to cAMP-dependent effectors observed in PM adipocytes after OVX seems to involve protein kinase A, since a decreased activation of cGI.PDE by protein kinase A was also found in these cells. Treatment of OVX rats with both estradiol and progesterone reversed the defective cAMP-dependent activation of cGI.PDE, but not the refractoriness of this isoenzyme to insulin activation. Taken together with previous observations from this laboratory on the fat cell adenylate cyclase system (Lacasa et al. (1991) Endocrinology 128, 747-753), these results: (a) demonstrate that the influence of the ovarian status on the key enzymes controlling cAMP metabolism in fat cells depends on the anatomical origin of these cells, and; (b) provide a biochemical explanation to the insensitivity of the SC adipocyte lipolytic system to ovarian hormones.     

Effects of 17β-estradiol and antioxidant administration on oxidative stress and insulin resistance in ovariectomized rats

The prevalence of insulin resistance syndrome increases during menopause with the overproduction of reactive oxygen species and impairment of the free radical scavenger function. Therefore, we investigated the effects of 17β-estradiol (E(2)) and vitamin E, as an antioxidant, on lipid peroxidation and antioxidant levels in the brain cortex and liver of ovariectomized rats as well as on insulin resistance in those rats. Forty female Sprague-Dawley rats, 3?months of age and weighing 231.5?± 9.4 g, were divided into 4 groups: sham, ovariectomized (OVX), OVX treated with E(2) (40 μg/kg subcutaneously), and OVX treated with E(2) and vitamin E (100?mg/kg intraperitoneally). The 4 groups received the appropriate treatment every day for 8?weeks. Levels of glutathione, glutathione peroxidase, superoxide dismutase , catalase, and malondialdehyde in the brain cortex and liver of ovariectomized rats were measured. Also, fasting plasma insulin, glucose, and homeostatis model assessment of insulin resistance (HOMA-IR) were determined. Malondialdehyde increased and antioxidants (glutathione, glutathione peroxidase, catalase, superoxide dismutase) decreased in the brain cortex and liver of OVX rats. Also, fasting glucose, insulin, and HOMA-IR increased in OVX rats. E(2) and E(2) plus vitamin E decreased malondialdehyde and increased antioxidants in the brain cortex and liver of OVX rats. Moreover, they decreased fasting glucose, insulin, and HOMA-IR in ovariectomized rats. This study demonstrates that E(2) and E(2) plus vitamin E supplementation to OVX rats may improve insulin resistance, strengthen the antioxidant system, and reduce lipid peroxidation.     

Influence of ovariectomy on ventricular myocyte contraction in simulated diabetes

We studied the effect of ovariectomy (OVX) on cardiac contraction in myocytes maintained under a 'diabetes-simulated high-glucose' environment. Female rats were ovariectomized or sham operated (SHAM) and kept for 6 weeks. Isolated myocytes were maintained in a diabetes-simulated high [glucose] medium (HG; 25.5 mM) for 24 h before mechanical properties were measured. Contractile indices analyzed included peak shortening (PS), time to PS (TPS), time to 90% relengthening (TR90), maximal velocity of shortening and relengthening (+/- dL/dt), intracellular Ca2+ fura-2 fluorescence intensity and decay rate (tau). Nitric oxide synthase (NOS) activity was also evaluated. OVX myocytes displayed a longer TR(90), slower +/- dL/dt, lower fluorescence intensity and higher tau (slower decay rate) when compared to SHAM myocytes. In the SHAM group, HG exerted diabetes-like contractile dysfunctions, including depressed PS, prolonged TR90, reduced fluorescence intensity, higher tau and enhanced NOS activity when compared to myocytes maintained in low [glucose] medium (5.5 mM). Interestingly, the HG- induced mechanical alterations were significantly exaggerated (TPS, TR90 and tau), reversed (PS and NOS) or lost (+/- dL/dt and fluorescence intensity) in the OVX group. These data suggest that ovarian hormones play a role in the regulation of cardiac contractile function, and may have potentially protective effects against diabetes-associated cardiac dysfunction.     

Cardiac metabolism in mice: tracer method developments and in vivo application revealing profound metabolic inflexibility in diabetes

Studies of cardiac fuel metabolism in mice have been almost exclusively conducted ex vivo. The major aim of this study was to assess in vivo plasma FFA and glucose utilization by the hearts of healthy control (db/+) and diabetic (db/db) mice, based on cardiac uptake of (R)-2-[9,10-(3)H]bromopalmitate ([3H]R-BrP) and 2-deoxy-D-[U-14C]glucose tracers. To obtain quantitative information about the evaluation of cardiac FFA utilization with [3H]R-BrP, simultaneous comparisons of [3H]R-BrP and [14C]palmitate ([14C]P) uptake were first made in isolated perfused working hearts from db/+ mice. It was found that [3H]R-BrP uptake was closely correlated with [14C]P oxidation (r2 = 0.94, P < 0.001). Then, methods for in vivo application of [3H]R-BrP and [14C]2-DG previously developed for application in the rat were specially adapted for use in the mouse. The method yields indexes of cardiac FFA utilization (R(f)*) and clearance (K(f)*), as well as glucose utilization (R(g)'). Finally, in the main part of the study, the ability of the heart to switch between FFA and glucose fuels (metabolic flexibility) was investigated by studying anesthetized, 8-h-fasted control and db/db mice in either the basal state or during glucose infusion. In control mice, glucose infusion raised plasma levels of glucose and insulin, raised R(g)' (+58%), and lowered plasma FFA level (-48%), K(f)* (-45%), and R(f)* (-70%). This apparent reciprocal regulation of glucose and FFA utilization by control hearts illustrates metabolic flexibility for substrate use. By contrast, in the db/db mice, glucose infusion raised glucose levels with no apparent influence on cardiac FFA or glucose utilization. In conclusion, tracer methodology for assessing in vivo tissue-specific plasma FFA and glucose utilization has been adapted for use in mice and reveals a profound loss of metabolic flexibility in the diabetic db/db heart, suggesting a fixed level of FFA oxidation in fasted and glucose-infused states.     

蛋白激酶C对自发性高血压大鼠肥大心肌细胞无负荷收缩功能的调节

在慢性压力超负荷引起心肌肥大过程中,蛋白激酶C(protein kinase C,PKC)的激活起关键性作用,激活的PKC也能调节心肌收缩性能.本文旨在研究自发性高血压大(spontaneously hypertensive rat,SHR)心肌肥大的不同阶段PKC调节心肌收缩性能的特征.采用胶原酶法分离4月龄与10月龄Wistar-Kyoto(WKY)、SHR大鼠的心肌细胞,观测单个心肌细胞无负荷缩短幅值以及在PKC激动剂与抑制剂作用下心肌收缩性能的变化.结果表明:刺激频率从1 Hz增至3 Hz,WKY大鼠心肌细胞无负荷缩短幅值逐渐增加,呈正阶梯效应;4月龄SHR大鼠心肌细胞的缩短幅值较WKY大鼠增强,但在各刺激频率下其缩短幅值基本保持不变;10月龄SHR大鼠心肌细胞的缩短幅值在1 Hz刺激条件下与WKY大鼠无差别,随刺激频率增加,缩短幅值降低,呈负阶梯效应.在PKC激动剂PMA灌流条件下,50、100与200 nmol/L的PMA分别降低WKY大鼠心肌细胞缩短幅值至(69.8±1.9)%、(58.2 2.2)%与(22.7±2.5)%(均P<0.01),呈浓度依赖关系;PMA对4月龄SHR大鼠心肌细胞缩短幅值的降低更明显,分别降至(6.1±0.7)%、(2.4±0.2)%与(12.5±2.6)%(均P<0.01);PMA降低10月龄SHR大鼠心肌细胞缩短幅值至(65.7±1.6)%、(53.9±4.0)%与(16.3±2.0)%(均P<0.01),小于对4月龄SHR大鼠心肌细胞缩短幅值的作用.PKC抑制剂staurosporine增加WKY大鼠心肌细胞缩短幅值,在200 nmol/L的staurosporine灌流条件下,WKY大鼠、4月龄SHR大鼠、10月龄SHR大鼠心肌细胞缩短幅值分别增JJH(63.63±4.53)%、(80.82±4.61)%、(80.97±4.59)%(均P<0.05).结果提示,在SHR大鼠心肌肥大初期,具有负性肌力作用的PKC异构体可能被激活,并参与对心肌收缩性能的调节;而心肌肥大稳定阶段,这些PKC活性可能恢复至正常水平.     

Comparison of local and systemic effects of insulin on myocardial glucose extraction in ischemic heart disease

Physiological increases in circulating insulin level significantly increase myocardial glucose uptake in vivo. To what extent this represents a direct insulin action on the heart or results indirectly from reduction in circulating concentrations of free fatty acids (FFA) is uncertain. To examine this, we measured myocardial glucose, lactate, and FFA extraction in 10 fasting men (ages 49-76 yr) with stable coronary artery disease during sequential intracoronary (10 mU/min, coronary plasma insulin = 140 +/- 20 microU/ml) and intravenous (100 mU/min, systemic plasma insulin = 168 +/- 26 microU/ml) insulin infusion. Basally, hearts extracted 2 +/- 2% of arterial glucose and extracted 27 +/- 6% of FFA. Coronary insulin infusion increased glucose extraction to 5 +/- 3% (P < 0.01 vs. basal) without changing plasma FFA or heart FFA extraction. Conversion to intravenous infusion lowered plasma FFA by approximately 50% and heart FFA extraction by approximately 75%, increasing heart glucose extraction still further to 8 +/- 3% (P < 0. 01 vs. intracoronary). This suggests the increase in myocardial glucose extraction observed in response to an increment in systemic insulin concentration is mediated equally by a reduction in circulating FFA and by direct insulin action on the heart itself. Coronary insulin infusion increased myocardial lactate extraction as well (from 20 +/- 10% to 29 +/- 9%, P < 0.05), suggesting the local action may include stimulation of a metabolic step distal to glucose transport and glycolysis.     

11beta-Hydroxysteroid dehydrogenase in the heart of normotensive and hypertensive rats

Corticosteroids have been shown to play a role in cardiac remodeling, with the possibility of a direct effect of overexpression of 11beta-hydroxysteroid dehydrogenase (11HSD) isoform 2 at the level of the cardiomyocytes. The aim of this study was to examine cardiac steroid metabolism in hypertensive rats with hearts that are hypertrophied and fibrotic and have structural alterations in the coronary circulation. To assess possible alterations of cardiac steroid metabolism the expression and activity of both isoforms of 11beta-hydroxysteroid dehydrogenase (11HSD) were studied in spontaneously hypertensive rats (SHR), their normotensive controls Wistar-Kyoto (WKY), and in Dahl salt-sensitive (DS) and salt-resistant rats (DR) kept on a low- or high-salt diet. Using real-time quantitative RT-PCR and enzyme activity assay we found strain-dependent differences in cardiac metabolism of glucocorticoids. In Dahl rats expression of 11HSD1 and 11HSD2 mRNA was lower in DS than in DR rats and was not influenced by dietary salt intake; 11HSD1 mRNA was expressed at higher level than 11HSD2 mRNA. NADP(+)-dependent cardiac 11HSD activity showed similar distribution as 11HSD1 mRNA-lower activity in DS than in DR rats and no effect of salt intake. In SHR and WKY strains 11HSD2 mRNA expression was significantly higher in WKY than in SHR but no differences were observed in 11HSD1 mRNA abundance and NADP(+)-dependent 11HSD activity. These results show that the heart is able to metabolize glucocorticoids and that this metabolism is strain-dependent but do not support the notion of association between cardiac hypertrophy and changes of 11HSD1 and 11HSD2 expression.     

Myocardium in hypertrophy: Oxygen consumption by isolated cardiac myocytes and working hearts from spontaneously hypertensive rats

Oxygen consumption was measured on suspensions of calcium tolerant myocytes obtained from hearts of Spontaneously Hypertensive Rats (SHR) and normotensive Wistar Kyoto Rats (WKY). Oxygen consumptions of the isolated cells were not significantly different from each other either in the presence or absence of added calcium (1.5 mM). Additionally, there was excellent agreement between the oxygen consumption of the isolated cells and estimates of basal oxygen consumption obtained from linear regression analysis of the relationship between work and myocardial oxygen utilization in isolated perfused working hearts. At any given workload there was no significant difference in oxygen consumption between SHR hearts and WKY hearts. The mechanical performance of the SHR hearts was lower compared to that of the WKY hearts at low preloads. At high preloads and high afterloads the SHR hearts developed higher pressures than did hearts obtained from WKY rats. The data suggest that: (a) basal oxygen consumption of the two hearts are similar and (b) the contractile defects in the SHR heart are not the result of hypoxia.     

Induced pluripotent stem cells' self-renewal and pluripotency is maintained by a bovine granulosa cell line-conditioned medium

Senescence marker protein-30 (SMP30) plays an important role in intracellular Ca²⁺ homeostasis. The aim of the present study was to investigate the effects of estrogens on liver apoptotic damage and changes in SMP30 expression induced by a high saturated fatty acid diet (HSFD). Ovariectomized mice (OVX) and sham-operated mice (SHAM) were randomly divided into five groups: SHAM fed a normal diet (SHAM/ND), SHAM fed HSFD (SHAM/HSFD), OVX fed ND (OVX/ND), OVX fed HSFD (OVX/HSFD) and OVX fed HSFD with 17β-estradiol (E2) supplementation using an implanted slow-release pellet (OVX/HSFD + E2). After 8 weeks, markers of endoplasmic reticulum (ER) stress and apoptosis, and levels of tumor necrosis factor-α (TNFα and SMP30 expression were investigated. Compared with SHAM/ND, OVX/HSFD mice showed significantly increased spliced X-box protein-1 (s-XBP1), phosphorylated eukaryotic initiation factor-2α (p-eIF2α), glucose-regulated protein 78 (GPR78), C/EBP homologous protein (CHOP), cytosolic cytochrome c, caspase-3 activity, and TNFα, and significantly decreased SMP30. These differences in OVX/HSFD mice were restored to the levels of SHAM/ND mice by E2 supplementation. These results suggest that E2 supplementation attenuates HSFD-induced liver apoptotic death in ovariectomized mice by up-regulating SMP30.     

Effect of hypertension on the development of diabetic cardiomyopathy

The effect of hypertension on the progression of diabetic cardiomyopathy was examined by attempting to induce a similar level of diabetes in both spontaneously hypertensive rats (SHR) and Wistar rats. Streptozotocin (STZ) was injected into SHR (45 mg/kg) and Wistar rats (55 mg/kg) before (eight weeks of age) and after (twelve weeks of age) the development of hypertension in the SHR. For both groups of animals, induction of diabetes resulted in depressed weight gain, increased food and fluid consumption, hypoinsulinemia, hyperglycemia, and hypertriglyceridemia. For the rats injected at eight weeks of age, an oral glucose tolerance test (OGTT) demonstrated that although the SHR were significantly less diabetic than Wistar rats, the degree of cardiac dysfunction was equivalent in both strains. These results suggest that hypertension was interacting with the diabetic condition to impair cardiac performance. Injecting SHR at twelve weeks of age increased the severity of diabetes but interestingly did not depress heart function compared with the non-diabetic SHR group. Injecting Wistar rats at this age also increased the severity of diabetes, but unlike the SHR diabetic animals, these rats still had impaired cardiac performance. These results suggest that hypertension exacerbates the cardiac dysfunction seen during diabetes, especially when SHR rats are injected with STZ prior to the elevation of blood pressure. Moreover, in the SHR, the development of LV hypertrophy at the time of STZ injection may have compensated for the damaging effects of diabetes on the myocardium, thereby enabling the heart to perform normally.     

Gender-related distinctions in protein kinase C activity in rat vascular smooth muscle

Gender differences in vascular reactivity have been suggested; however, the cellular mechanisms involved are unclear. We tested the hypothesis that the gender differences in vascular reactivity reflect gender-related, possibly estrogen-mediated, distinctions in the expression and activity of specific protein kinase C (PKC) isoforms in vascular smooth muscle. Aortic strips were isolated from intact and gonadectomized male and female Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). Isometric contraction was measured in endothelium-denuded aortic strips. PKC activity was measured in the cytosolic and particulate fractions, and the amount of PKC was measured using Western blots and isoform-specific anti-PKC antibodies. In intact male WKY rats, phenylephrine (Phe, 10(-5) M) and phorbol 12,13-dibutyrate (PDBu, 10(-6) M) stimulated contraction to 0.37 +/- 0.02 and 0.42 +/- 0.02 g/mg tissue wt, respectively. The basal particulate/cytosolic PKC activity ratio was 0.86 +/- 0.06, and Western blots revealed alpha-, delta-, and zeta-PKC isoforms. Phe and PDBu increased PKC activity and caused significant translocation of alpha- and delta-PKC from the cytosolic to particulate fraction. In intact female WKY rats, basal PKC activity, the amount of alpha-, delta-, and zeta-PKC, the Phe- and PDBu-induced contraction, and PKC activity and translocation of alpha- and delta-PKC were significantly reduced compared with intact male WKY rats. The basal PKC activity, the amount of alpha-, delta-, and zeta-PKC, the Phe and PDBu contraction, and PKC activity and alpha- and delta-PKC translocation were greater in SHR than WKY rats. The reduction in Phe and PDBu contraction and PKC activity in intact females compared with intact males was greater in SHR ( approximately 30%) than WKY rats ( approximately 20%). Phe and PDBu contraction and PKC activity were not significantly different between castrated males and intact males but were greater in ovariectomized (OVX) females than intact females. Treatment of OVX females or castrated males with 17 beta-estradiol, but not 17 alpha-estradiol, subcutaneous implants caused significant reduction in Phe and PDBu contraction and PKC activity that was greater in SHR than WKY rats. Phe and PDBu contraction and PKC activity in OVX females or castrated males treated with 17 beta-estradiol plus the estrogen receptor antagonist ICI-182,780 were not significantly different from untreated OVX females or castrated males. Thus a gender-related reduction in vascular smooth muscle contraction in female WKY rats with intact gonads compared with males is associated with reduction in the expression and activity of vascular alpha-, delta-, and zeta-PKC. The gender differences in vascular smooth muscle contraction and PKC activity are augmented in the SHR and are possibly mediated by estrogen.     

A Comparison of the Effects of Estrogen and Cimicifuga racemosa on the Lacrimal Gland and Submandibular Gland in Ovariectomized Rats

This study aims to observe the effects of estradiol and Cimicifuga racemosa on the lacrimal gland and submandibular gland of ovariectomized rats. We randomly divided 20 adult female SD rats into four groups—a sham-operated group (SHAM), ovariectomized (OVX) group, ovariectomized group treated with estradiol (OVX+ E), and ovariectomized group treated with the isopropanolic extract of Cimicifuga racemosa (OVX+ iCR). The SHAM group and OVX group used distilled water to instead the drugs. Two weeks after ovariectomy, the estradiol and iCR were administered for 4 weeks. Next, we used H&E staining and electron microscopy to observe any histological changes in the lacrimal and submandibular glands and immunohistochemical staining to observe the expressions of cleaved caspase-3 (Casp-3) and Cu-Zn SOD (superoxide dismutase). The H&E staining find that both drugs can prevent the cells of area from shrinkage in the two kinds of gland. But under the electron microscopy, estradiol and iCR have different efficacy. Estradiol is more effective at protecting mitochondria in lacrimal gland acinar cells than iCR, and iCR is more effective at suppressing endoplasmic reticulum expansion than estradiol. Both estradiol and iCR have a similar protective function on mitochondria in the submandibular gland. The protective function of the two glands may inhibit apoptosis by suppressing the expression of Casp-3. In addition, iCR increases the expression of Cu-Zn SOD in duct system of submandibular gland. The results suggest that both estradiol and iCR confer a protective effect on the lacrimal and submandibular glands of ovariectomized rats via different mechanisms.