Ningtian Zhou

Beneficial effects of schisandrin B on the cardiac function in mice model of myocardial infarction.

The fruit of Schisandra chinensis has been used in the traditional Chinese medicine for thousands of years. Accumulating evidence suggests that Schisandrin B (Sch B) has cardioprotection effect on myocardial ischemia in vitro. However, it is unclear whether Sch B has beneficial effects on continuous myocardial ischemia in vivo. The aim of the present study was to investigate whether Sch B could improve cardiac function and attenuate myocardial remodeling after myocardial infarction (MI) in mice. Mice model of MI was established by permanent ligation of the left anterior descending (LAD) coronary artery. Then the MI mice were randomly treated with Sch B or vehicle alone. After treatment for 3 weeks, Sch B could increase survival rate, improve heart function and decrease infarct size compared with vehicle. Moreover, Sch B could down-regulate some inflammatory cytokines, activate eNOS pathway, inhibit cell apoptosis, and enhance cell proliferation. Further in vitro study on H9c2 cells showed similar effects of Sch B on prevention of hypoxia-induced inflammation and cell apoptosis. Taken together, our results demonstrate that Sch B can reduce inflammation, inhibit apoptosis, and improve cardiac function after ischemic injury. It represents a potential novel therapeutic approach for treatment of ischemic heart disease.

Beneficial effects of ginsenoside-Rg1 on ischemia-induced angiogenesis in diabetic mice

Neovascularization and the formation of collateral vessels are often impaired in diabetes mellitus (DM) population compared with non-diabetics. Alterations in vascular endothelial growth factor (VEGF) signaling and endothelial nitric oxide synthase (eNOS) dysfunction have been confirmed to play a crucial role in impaired neovascularization in diabetic mice. Accumulating data have suggested that Rg1, a main component of Panax ginseng, has the ability to promote tubulogenesis of human umbilical vein endothelial cells (HUVECs) in vitro, and that the mechanism involves increased expression level of VEGF as well as increased eNOS activation. Thus, we speculated that Rg1 might also have therapeutic effects on the impairment of neovascularization in diabetic individuals. The aim of the present study was to investigate whether Rg1 could improve angiogenesis in ischemic hindlimb of diabetic mice in vivo. Our data demonstrated that Rg1 treatment resulted in improved angiogenesis in the diabetic ischemic hindlimb, and the potential mechanism might involve increased eNOS activation, upregulated VEGF expression, and inhibited apoptosis. Our results suggest that Rg1 may be used as a novel and useful adjunctive drug for the therapy of peripheral arterial disease in DM.

Beneficial effects of muscone on cardiac remodeling in a mouse model of myocardial infarction.

Musk has been traditionally used in East Asia to alleviate the symptoms of angina pectoris. However, it remains unclear as to whether muscone, the main active ingredient of musk, has any beneficial effects on persistent myocardial ischemia in vivo. The aim of the present study was to investigate whether muscone can improve cardiac function and attenuate myocardial remodeling following myocardial infarction (MI) in mice. Mice were subjected to permanent ligation of the left anterior descending coronary artery to induce MI, and then randomly treated with muscone (2 mg/kg/day) or the vehicle (normal saline) for 3 weeks. Sham-operated mice were used as controls and were also administered the vehicle (normal saline). Treatment with muscone significantly improved cardiac function and exercise tolerance, as evidenced by the decrease in the left ventricular end-systolic diameter, left ventricular end-diastolic diameter, as well as an increase in the left ventricular ejection fraction, left ventricular fractional shortening and time to exhaustion during swimming. Pathological and morphological assessments indicated that treatment with muscone alleviated myocardial fibrosis, collagen deposition and improved the heart weight/body weight ratio. Muscone inhibited the inflammatory response by reducing the expression of transforming growth factor (TGF)-β1, tumor necrosis factor (TNF)-α, interleukin (IL)-1β and nuclear factor (NF)-κB. Treatment with muscone also reduced myocardial apoptosis by enhancing Bcl-2 and suppressing Bax expression. Muscone also induced the phosphorylation of protein kinase B (Akt) and endothelial nitric oxide synthase (eNOS). Our results demonstrate that muscone ameliorates cardiac remodeling and dysfunction induced by MI by exerting anti-fibrotic, anti-inflammatory and anti-apoptotic effects in the ischemic myocardium.

Association of admission serum calcium levels and in-hospital mortality in patients with acute ST-elevated myocardial infarction: an eight-year, single-center study in China.

Objective The relationship between admission serum calcium levels and in-hospital mortality in patients with acute ST-segment elevation myocardial infarction (STEMI) has not been well definitively explored. The objective was to assess the predictive value of serum calcium levels on in-hospital mortality in STEMI patients. Methods From 2003 to 2010, 1431 consecutive STEMI patients admitted to the First Affiliated Hospital of Nanjing Medical University were enrolled in the present study. Patients were stratified according to quartiles of serum calcium from the blood samples collected in the emergency room after admission. Between the aforementioned groups,the baseline characteristics, in-hospital management, and in-hospital mortality were analyzed. The association of serum calcium level with in-hospital mortality was calculated by a multivariable Cox regression analysis. Results Among 1431 included patients, 79% were male and the median age was 65 years (range, 55–74). Patients in the lower quartiles of serum calcium, as compared to the upper quartiles of serum calcium, were older, had more cardiovascular risk factors, lower rate of emergency revascularization,and higher in-hospital mortality. According to univariate Cox proportional analysis, patients with lower serum calcium level (hazard ratio 0.267, 95% confidence interval 0.164–0.433, p<0.001) was associated with higher in-hospital mortality. The result of multivariable Cox proportional hazard regression analyses showed that the Killips class≥3 (HR = 2.192, p = 0.026), aspartate aminotransferase (HR = 1.001, p<0.001), neutrophil count (HR = 1.123, p<0.001), serum calcium level (HR = 0.255, p = 0.001), and emergency revascularization (HR = 0.122, p<0.001) were significantly and independently associated with in-hospital mortality in STEMI patients. Conclusions Serum calcium was an independent predictor for in-hospital mortality in patients with STEMI. This widely available serum biochemical index may be incorporated into the current established risk stratification model of STEMI patients. Further studies are required to determine the actual mechanism and whether patients with hypocalcaemia could benefit from calcium supplement.

Puerarin improves cardiac function through regulation of energy metabolism in Streptozotocin-Nicotinamide induced diabetic mice after myocardial infarction.

It is well recognized that the incidence of heart failure and the risk of death is high in diabetic patients after myocardial infarction (MI). Accumulating evidence showed that puerarin (PUE) has protecting function on both cardiovascular disease and diabetes. The aim of this study is to explore whether puerarin could improve cardiac function in diabetic mice after MI and the underlying mechanism. The left anterior of Streptozotocin (STZ)-Nicotinamide (NA) induced diabetic mice were ligated permanently except for the Shame group. Then the operated mice were randomly treated with PUE or saline. Cardiac function was evaluated by echocardiograph before and at 1, 2, 4 weeks after MI. GLUT4/CD36/p-Akt/PPAR α of the heart was examined after treatment for 4 weeks. The results indicated that PUE significantly increased survival rate, improved cardiac function compared with MI group. Moreover, PUE increased expression and translocation of GLUT4 while attenuated expression and translocation of CD36. Western blot analysis showed that PUE enhanced phosphorylation of Akt and decreased PPAR α. This study demonstrated that PUE improved cardiac function after MI in diabetic mice through regulation of energy metabolism, the possible mechanism responsible for the effect of PUE was increasing the expression and translocation of GLUT4 while attenuating the expression and translocation of CD36.

p27 kip1 haplo-insufficiency improves cardiac function in early-stages of myocardial infarction by protecting myocardium and increasing angiogenesis by promoting IKK activation

p27kip1 (p27) is widely known as a potent cell cycle inhibitor in several organs, especially in the heart. However, its role has not been fully defined during the early phase of myocardial infarction (MI). In this study, we investigated the relationships between p27, vascular endothelial growth factor/hepatocyte growth factor (VEGF/HGF) and NF-κB in post-MI cardiac function repair both in vivo and in the hypoxia/ischemia-induced rat myocardiocyte model. In vivo, haplo-insufficiency of p27 improved cardiac function, diminished the infarct zone, protected myocardiocytes and increased angiogenesis by enhancing the production of VEGF/HGF. In vitro, the presence of conditioned medium from hypoxia/ischemia-induced p27 knockdown myocardiocytes reduced the injury caused by hypoxia/ischemia in myocardiocytes, and this effect was reversed by VEGF/HGF neutralizing antibodies, consistent with the cardioprotection being due to VEGF/HGF secretion. We also observed that p27 bound to IKK and that p27 haplo-insufficiency promoted IKK/p65 activation both in vivo and in vitro, thereby inducing the NF-κB downstream regulator, VEGF/HGF. Furthermore, IKKi and IKK inhibitor negated the effect of VEGF/HGF. Therefore, we conclude that p27 haplo-insufficiency protects against heart injury by VEGF/HGF mediated cardioprotection and increased angiogenesis through promoting IKK activation.

Exogenous HGF prevents cardiomyocytes from apoptosis after hypoxia via up-regulating cell autophagy

Background: Hepatocyte growth factor (HGF) is widely known as a protective factor in ischemic myocardium, however HGF sensitive cellular mechanism remained ill-defined. Autophagy at early stage of hypoxia has been demonstrated to play a role in protecting myocardium both in vivo and vitro. We performed this study to investigate the association between the protective effect of HGF and autophagy. Methods: Ventricular myocytes were isolated from neonatal rat heart (NRVMs). We evaluated cardiomyocytes apoptosis by Hoechst staining and flow cytometry. Autophagy was assessed by transmission electron microscope and mRFP-GFP-LC3 adenovirus infection. Mitochondrial membrane potential was estimated by JC-1 staining. Western blotting and ELISA assay were used to quantify protein concentrations. Results: We found that autophagy in NRVMs increased at early stage after hypoxia and HGF release was consistent with the change of autophagy. Exogenous HGF enhanced autophagy and decreased apoptosis, while neutralizing HGF yielded opposite effects. Besides, inhibition of autophagy increased apoptosis of myocytes. Furthermore, exogenous HGF induced Parkin, the marker of mitochondrial autophagy, indicating increased clearance of injured mitochondria. Conclusions: Our results revealed a potential mechanism in which exogenous HGF prevented NRVMs from apoptosis after hypoxia. Upregulation of Parkin through administration of exogenous HGF may be a potential therapeutic strategy ptotecting myocytes during ischemia.

Necroptosis Induced by Ad-HGF Activates Endogenous C-Kit+ Cardiac Stem Cells and Promotes Cardiomyocyte Proliferation and Angiogenesis in the Infarcted Aged Heart

Background/Aims: The discovery of c-kit+ cardiac stem cells (CSCs) provided us with new therapeutic targets to repair the damaged heart. However, the precise mechanisms regulating CSC proliferation and differentiation in the aged heart remained elusive. Necroptosis, a type of regulated cell death, has recently been shown to occur following myocardial infarction (MI); however, its effect on c-kit+ CSCs remains unknown. We investigated the effects of hepatocyte growth factor (HGF) and necroptosis on the proliferation and differentiation of endogenous c-kit+ CSCs in aged rat hearts following MI. Methods: The c-kit+ CSCs and HGF/p-Met expression levels in neonatal, adult and aged rats were compared using immunofluorescence and Western blotting. Immediately after MI, adenovirus carrying the HGF gene (Ad-HGF) was injected into the left ventricular wall surrounding the infarct areas of the aged rat heart. The proliferation and differentiation of the endogenous c-kit+ CSCs were studied using immunofluorescence. The signalling pathways were analysed via Western blotting and ELISA. Results: HGF/p-Met expression levels and c-kit+ CSC abundance gradually decreased with age. Ad-HGF promoted c-kit+ CSC differentiation into precursor cells of cardiomyocyte, endothelial and smooth muscle cell lineages and enhanced cardiomyocyte proliferation and angiogenesis in aged rats; these effects were reversed by the inhibition of necroptosis. Ad-HGF administration induced necroptosis by increasing the expression of receptor interacting protein kinase (RIP) 1 and receptor interacting protein kinase (RIP) 3 proteins in the infarcted heart. Moreover, Ad-HGF-induced necroptosis increased high-mobility group box 1 protein (HMGB1) levels and enhanced the abundance of c-kit+ cells in the bone marrow, which may partly account for the beneficial effect of necroptosis on the c-kit+ CSCs. Conclusion: Ad-HGF-induced necroptosis facilitated aged heart repair after MI by promoting c-kit+ CSC proliferation and differentiation. These findings may lead to the development of new methods for the treatment of ischaemic heart disease in aged populations.

Cardiac-Specific Expression of the Hepatocyte Growth Factor (HGF) Under the Control of a TnIc Promoter Confers a Heart Protective Effect After Myocardial Infarction (MI)

OBJECTIVE Uncontrolled therapeutic gene expression and neovascularization in non-specific tissues has lowered the safety of gene therapy. The aim of the study was to identify a cardiac-specific promoter to control target gene expression in heart tissue in vitro and in vivo. METHODS Adenovirus vectors containing the firefly luciferase or hepatocyte growth factor (HGF) genes under control of the Troponin I (TnIc) or Cytomegalievirus (CMV) promoters were transfected into cell lines or injected into the left ventricular wall of Sprague Dawley (SD) rats via thoracotomy. Myocardial infarction (MI) was induced immediately before direct injection. In vivo luciferase expression was assessed using a bioluminescence imaging system. Heart function was monitored via echocardiograph intermittently for eight weeks after injection. RESULTS The constitutively active CMV promoter yielded luciferase expression throughout the body while luciferase expression driven by the TnIc promoter was largely restricted to the hypoxic heart. The CMV promoter was more efficient, yielding 100-1000 fold more light output than the TnIc promoter. Four weeks after injection, we observed a significant decline in the ejection fraction (EF) in saline and Ad-Null groups but a 17% increase in the Ad-CMV-HGF group. No change in EF was observed in the Ad-TnIc-HGF group. CONCLUSIONS The adenovirus vector combined with the TnIc promoter largely restricts gene-targeted therapy in the hypoxic heart and prevents heart failure after myocardial infarction.

Diagnostic and prognostic value of minor elevated cardiac troponin levels for percutaneous coronary intervention-related myocardial injury: a prospective, single-center and double-blind study.

Abstract Cardiac troponin-I (cTnI) and -T (cTnT) are sensitive and specific markers of myocardial injury. However, the role of increased cTnI and cTnT in percutaneous coronary intervention (PCI)-related myocardial injury remains controversial. In this prospective, single-center and double-blind study, we aimed to determine the diagnostic and prognostic value of cTnI as well as cTnT (cTns) in PCI-related myocardial injury in a Chinese population. A total of 1,008 patients with stable angina pectoris and non-ST-segment elevation acute coronary syndrome were recruited. The levels of cTnI and cTnT were examined before and after PCI. All patients were followed up for 26±9 months to observe the incidence of major adverse cardiac events (MACEs). Our results showed that post-PCI cTnI and/or cTnT levels were increased to more than the 99th percentile upper reference limit (URL) in 133 (13.2%) patients, among which 22 (2.2%) were more than 5 × 99th percentile URL. By univariate analysis, an elevation in cTns after PCI was not an independent predictor of increased MACEs, HR 1.35 (P  =  0.33, 95%CI: 0.74–2.46). In conclusion, our data demonstrate that the incidence of PCI-related myocardial injury is not common in a Chinese population and minor elevated cTns levels may not be a sensitive prognostic marker for MACEs.