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2,3,5,4'-Tetrahydrostilbene-2-o-β-d-glucoside (TSG) is the main active component of Polygonum multiflorum Thunb. It has effects on hypertension. However, the mechanism is unclear. Current research is devoted to exploring the mechanism of TSG improving HHcy-induced hypertension. The mice received a subcutaneous injection of Hcy in the presence or absence of TSG for 4 weeks. Blood pressure (BP) was measured using a noninvasive tail-cuff plethysmography method. Levels of plasma Hcy and endothelin-1 were measured using ELISA. Rat SMA without endothelium was cultured in a serum-free medium in the presence or absence of TSG with or without Hcy. The contractile response to sarafotoxin 6c or endothein-1 was studied using a sensitive myography. The levels of protein were detected using Western blotting. The results showed that TSG lowered HHcy-elevated BP and decreased levels of plasma Hcy and endothelin-1 in mice. Furthermore, the results showed that TSG inhibited Hcy-upregulated ET receptor expression and ET receptor-mediated contractile responses as well as the levels of p-ERK

Increased vascular smooth muscle cell (VSMC) endothelin type B (ETRat superior mesenteric arteries (SMAs) without endothelium were cultured in medium without serum for 24 h. HG with or without mitogen-activated protein kinase (MAPK) signaling pathway inhibitors and downstream nuclear factor-kappaB (NF-κB) inhibitors was coincubated with SMAs. A sensitive myograph detected the contractile responses to sarafotoxin 6c. Western blotting and immunofluorescence staining were used to determine protein expression.HG promoted the expression of VSMC ETIn conclusion, HG upregulated the VSMC ET

OBJECTIVE: To investigate the protective effect of melatonin against myocardial ischemia reperfusion (IR) injury in isolated rat hearts and explore the underlying mechanisms. METHODS: The isolated hearts from 40 male SD rats were randomly divided into 4 groups (n=10): the control group, where the hearts were perfused with KH solution for 175 min; IR group, where the hearts were subjected to global ischemia for 45 min followed by reperfusion for 120 min; IR+melatonin (Mel+IR) group, where melatonin (5 μmol/L) was administered to the hearts 1 min before ischemia and during the first 5 min of reperfusion, followed by 115 min of reperfusion; and IR+2, 3-butanedione monoxime (IR+BDM) group, where the hearts were treated with BDM (20 mmol/L) in the same manner as melatonin treatment. Myocardial injury in the isolated hearts was assessed based on myocardial injury area, caspase-3 activity, and expressions of cytochrome C and cleaved caspase-3 proteins. Cardiac contracture was assessed using HE staining and by detecting lactate dehydrogenase (LDH) activity and the content of cardiac troponin I (cTnI) in the coronary outflow, measurement of left ventricular end-diastolic pressure (LVEDP) and electron microscopy. The content of ATP in the cardiac tissue was also determined. RESULTS: Compared with those in the control group, the isolated hearts in IR group showed significantly larger myocardial injury area and higher caspase-3 activity and the protein expressions of cytochrome C and cleaved caspase-3 with significantly increased LDH activity and cTnI content in the coronary outflow and elevated LVEDP at the end of reperfusion; HE staining showed obvious fractures of the myocardial fibers and the content of ATP was significantly decreased in the cardiac tissue; electron microscopy revealed the development of contraction bands. In the isolated hearts with IR, treatment with Mel or BDM significantly reduced the myocardial injury area, caspase-3 activity, and protein expressions of cytochrome C and cleaved caspase-3, obviously inhibited LDH activity, lowered the content of cTnI and LVEDP, reduced myocardial fiber fracture, and increased ATP content in the cardiac tissue. Both Mel and BDM inhibited the formation of contraction bands in the isolated hearts with IR injury. CONCLUSION: Mel can alleviate myocardial IR injury in isolated rat hearts by inhibiting cardiac contracture, the mechanism of which may involve the upregulation of ATP in the cardiac myocytes to lessen the tear of membrane and reduce cell content leakage.

Ca2+/calmodulin-dependent protein kinase II δ (CaMKIIδ) has been shown to play a vital role in pathological events in myocardial ischemia/reperfusion (IR) injury. Dysregulation of autophagy in cardiomyocytes is implicated in myocardial IR injury. Here, we examined whether CaMKIIδ inhibition could protect against myocardial IR injury through alleviating autophagy dysfunction and evaluated the potential role of CaMKIIδ in Beclin-1-dependent autophagy in ischemia/reperfused hearts. This study was performed using isolated perfused rat hearts and H9c2 cardiac myoblasts. KN-93, but not KN-92, inhibited the phosphorylation of CaMKIIδ at Thr286 and its substrate phospholamban at Thr17 besides the CaMKIIδ activity in myocardial IR. KN-93, but not KN-92 significantly improved post-ischemic cardiac function and reduced cell death. In cultured H9c2 cardiac myoblasts, KN-93 or CaMKIIδ siRNA, but not KN-92, attenuated simulated IR (SIR)-induced cell death. Moreover, CaMKIIδ inhibition could alleviate IR-induced autophagic dysfunction as evidenced in reduced levels of Atg5, p62, and LC3BII in isolated rat hearts and H9c2 cardiac myoblasts. Furthermore, co-treatment with bafilomycin A1, a lysosomal inhibitor, in CaMKII inhibition-treated cells suggested that CaMKII inhibition alleviated autophagic flux. CaMKIIδ inhibition mitigated the phosphorylation of Beclin-1 at Ser90. As expected, Beclin-1 siRNA significantly decreased the levels of Beclin-1 and Beclin-1 phosphorylation accompanied by partial reductions in Atg5, LC3BII, p62, cleaved caspase-3 and cytochrome c. However, Beclin-1 siRNA had little effect on CaMKIIδ phosphorylation. Taken together, these results demonstrated that CaMKIIδ inhibition reduced myocardial IR injury by improving autophagy dysfunction, and that CaMKIIδ-induced autophagy dysfunction partially depended on the phosphorylation of Beclin-1.

To investigate the role of Na+/Ca2+ exchanger (NCX) in myocardial ischemia-reperfusion injury and the underlying mechanisms. Methods: Forty Sprague-Dawley rats were divided into 4 groups randomly: a control group, a KB-R7943 group, an ischemia-reperfusion group (IR group), and an IR plus KB-R7943 group (KB-R7943+IR group). Isolated Sprague Dawley male rat hearts underwent Langendorff perfusion. The ratio of left ventricular developed pressure (LVDP), left ventricular end-diastolic pressure (LVEDP), the infarct size of myocardium, and the lactate dehydrogenase (LDH) activity in the coronary flow was determined. HE staining was used to assess the change of myocardial morphology. Western blot was used to determine the levels of cleaved caspase-3, cytochrome c and the phosphorylation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and the Thr17 site of phospholamban. Results: Compared with the control group, IR group significantly induced an enlarged infarct size, reduction of the ratio of LVDP, up-regulation of cytochrome c, cleaved caspase-3, p-CaMKII and p-phospholamban, and increased in the activity of LDH, the level of LVEDP (P0.01) and the disordered myocardial morphology. These effects were significantly attenuated in the presence of KB-R7943 treatment (10 μmol/L). Conclusion: NCX mediates myocardial ischemia-reperfusion-induced cell apoptosis and necrosis through activation of CaMKII.目的：探讨钠钙交换体(Na+/Ca2+ exchanger，NCX)在离体心脏缺血再灌注损伤中的作用及其可能机制。方法：40只大鼠随机分为4组：正常对照组(Control组)、10 μmol/L KB-R7943药物对照组(KB-R7943组)、缺血再灌注(ischemia reperfusion，IR)组和10 μmol/L KB-R7943干预缺血再灌注组(KB-R7943+IR组)。采用Langendorff灌流装置建立离体心脏缺血再灌注模型；以再灌注末心肌纤维形态的变化、左室发展压(LVDP)比值、左室舒张末压(LVEDP)、冠状动脉流出液中乳酸脱氢酶(lactate dehydrogenase，LDH)活性、心肌梗死面积及细胞色素c和cleaved caspase-3蛋白的变化评价心肌损伤程度；Western印迹检测磷酸化钙/钙调蛋白依赖的蛋白激酶II(Ca2+/calmodulin-dependent protein kinase II，CaMKII)和受磷蛋白(phospholamban，PLN)苏氨酸17位点磷酸化(pThr17)水平的变化。结果：与Control组相比，IR组再灌注末心肌纤维排列紊乱，断裂明显，心肌梗死面积、LVEDP和冠状动脉流出液中LDH活性明显增加，LVDP比值降低，细胞色素c、cleaved caspase-3、磷酸化CaMKII及pThr17-PLN水平均上调(P0.01)；KB-R7943+IR组心肌梗死面积明显减小，LVEDP降低，LVDP比值明显改善，LDH活性明显降低，细胞色素c和cleaved caspase-3蛋白水平明显降低，磷酸化CaMKII和pThr17-PLN水平也明显下调(P0.01)。结论：NCX可通过激活CaMKII启动细胞凋亡和坏死，进而诱导心肌缺血再灌注损伤。.

To investigate the role of melatonin in calcium overload-induced heart injury. Methods: Thirty-two rats were divided into 4 groups: a control group (Control), a melatonin control group (Mel), a calcium overload group (CaP), and a calcium overload plus melatonin group (Mel+CaP). Isolated Sprague Dawley male rat hearts underwent Langendorff perfusion. Left ventricular developed pressure (LVDP) was calculated to evaluate the myocardial performance. Triphenyltetrazolium chloride staining was used to measure the infarct size of myocardium. Lactate dehydrogenase (LDH) activity in the coronary flow was determined. The expressions of caspase-3 and cytochrome c were determined by Western blot. The pathological morphological changes in myocardial fiber were analyzed by HE staining. Results: Compared with the control group, calcium overload significantly induced an enlarged infarct size (P0.01), accompanied by the disordered arrangement of myocardial fiber, up-regulation of cytochrome c and caspase-3 (P0.01), and the increased activity of LDH (P0.01). These effects were significantly attenuated by 10 μmol/L melatonin (P0.01). Conclusion: Melatonin can alleviate calcium overload-induced heart injury.目的：观察褪黑素(melatonin，Mel)对大鼠离体心脏急性钙超载损伤的保护作用。方法：将32只大鼠随机分为4组：对照组(Control)、褪黑素组(Mel)、钙反常组(calcium paradox，CaP)和Mel干预钙反常组(Mel+CaP)。大鼠离体心脏行Langendorff灌流，建立钙反常模型；观察左室发展压、心肌梗死面积、乳酸脱氢酶(lactate dehydrogenase，LDH)活性、细胞色素c及caspase-3蛋白的表达变化；HE染色观察心肌纤维形态学变化。结果：与Control组相比，CaP组心肌几乎全部梗死(P0.01)，冠状动脉流出液中LDH活性显著增加(P0.01)，心肌纤维排列紊乱，细胞色素c和caspase-3蛋白表达均上调(P0.01)；Mel(10 μmol/L)干预钙反常组心脏损伤各项指标均明显降低(P0.01)。结论：Mel可明显减轻大鼠离体心脏急性钙超载诱导的损伤。.

Hyperhomocysteinemia is an independent risk factor for cardiovascular diseases (CVDs). Endothelin type B (ETB) receptors were involved in the pathogenesis of CVDs. However, Sirtuin 1(Sirt1) has potential protect roles for CVDs. The present study was designed to examine the hypothesis that homocysteine up-regulates ETB receptors through down-regulation of Sirt 1. In vitro experiments were performed in rat superior mesenteric artery (SMA). The rat SMA was cultured in serum free medium for 24h in the presence and absence of homocysteine (Hcy) with or without resveroral (Res) (a Sirt 1agonist). In vivo, the rats received subcutaneous injections of Hcy in the presence of or absence of Res for 3 weeks. The contractile responses to sarafotoxin 6c (S6c) (an ETB receptor agonist) were studied using a sensitive myograph. Levels of protein expression were determined using western blotting. The blood pressure of rat was measured via a noninvasive tail-cuff plethysmography method. We observed that Hcy increased the level of ETB receptor protein expression and the ETB receptor-mediated contractile responses induced by S6c, and decreased level of Sirt1 protein expression in SMA without endothelium in vitro. However, these effects were reversed by Res. Moreover, Res also blocked the up-regulation of acetylized p65 induced by Hcy. The in vivo study showed that HHcy down-regulated Sirt 1, and up-regulated acetylized p65 and ETB receptor protein expression, and elevated the blood pressure of rats. However, Res could block these effects. In conclusion, this suggested that Hcy regulated ETB receptor expression through sirt1/nuclear factor-κB signaling pathway.

The gastric mucosa protection effect of three natural plant extracts, Hericium erinaceus (HE), Centella asiatica (CA) and Amomum villosum (AV), were evaluated using the indomethacin damage model. Compared with a single extract, a combination of HE/CA/AV, especially with the ratios of 80:10:10, 45:45:10 and 45:10:45, showed significant synergistic effects for protection of the gastric mucosa with gastric ulcer inhibition rates of 97.8 ± 0.7%, 86.5 ± 2.8% and 86.1 ± 3.6%, respectively. Microscopic appearances of the gastric mucosa were carried out to help confirm the results.

To investigate the changes in aorta morphology and Ca(2+)-activated K(+) (KCa) channel expression in the diabetic rats.A diabetic rat model was established by a single intraperitoneal injection of streptozotocin (30 mg/kg) after a modified high fat and glucose diet for 8 weeks. Pathological changes in the aorta were observed with HE staining, elastic fiber staining, Masson's trichrome staining and immunohistochemistry. Both the mRNA and protein levels of KCa channels in the aorta were measured by RT-PCR and Western blotting.Early atherosclerotic changes were observed in the aorta wall of the diabetic rats. The mRNA and protein levels of KCa1.1 channel α- and β-subunits were significantly decreased, while the expression of KCa3.1 channels was obviously enhanced in the middle layer of the aorta in the diabetic rats.KCa channel switching in smooth muscles may play a role in the development of atherosclerosis in diabetic rats.

To investigate the role of K(Ca)3.1 channel in the proliferation and migration of rat vascular smooth muscle cells of the proliferative phenotype.Rat vascular smooth muscle cells (VSMCs) were cultured with tissue adhesion method. The morphological characteristics of the fist and ninth passages of VSMCs were observed with light and electron microscopy and immunocytochemistry. The expressions of K(Ca)3.1 channel mRNA and protein in the cells were detected using RT-PCR and immunocytochemistry, respectively. MTT and transwell assay were employed to assess the effect of the K(Ca)3.1 channel blocker TRAM-34 on the proliferation and migration of VSMCs.The first and ninth passages of VSMCs showed morphological characteristics of contractile and proliferative phenotypes, respectively. Compared with the first- passage cells, the ninth-passage VSMCs exhibited significantly increased K(Ca)3.1 channel mRNA and protein expressions with enhanced cell proliferation and migration (P0.01), which was inhibited by the application of TRAM-34 (P0.01). TRAM-34 produced no obvious effect on the first-passage VSMCs.Upregulated expression of K(Ca)3.1 channel can promote the proliferation and migration of rat VSMCs of the proliferative phenotype.