Zhengxian Tao

Coexpression of VEGF and angiopoietin-1 promotes angiogenesis and cardiomyocyte proliferation reduces apoptosis in porcine myocardial infarction (MI) heart

VEGF and angiopoietin-1 (Ang1) are two major angiogenic factors being investigated for the treatment of myocardial infarction (MI). Targeting VEGF and Ang1 expression in the ischemic myocardium can increase their local therapeutic effects and reduce possible adverse effects. Adeno-associated viral vectors (AAVs) expressing cardiac-specific and hypoxia-inducible VEGF [AAV-myosin light chain-2v (MLC)VEGF] and Ang1 (AAV-MLCAng1) were coinjected (VEGF/Ang1 group) into six different sites of the porcine myocardium at the peri-infarct zone immediately after ligating the left descending coronary artery. An identical dose of AAV-Cytomegalovirus (CMV)LacZ or saline was injected into control animals. AAV genomes were detected in the liver in addition to the heart. RT-PCR, Western blotting, and ELISA analyses showed that VEGF and Ang1 were predominantly expressed in the myocardium in the infarct core and border of the infarct heart. Gated single-photon emission computed tomography analyses showed that the VEGF/Ang1 group had better cardiac function and myocardial perfusion at 8 wk than at 2 wk after vector injection. Compared with the saline and LacZ controls, the VEGF/Ang1 group expressed higher phosphorylated Akt and Bcl-xL, less Caspase-3 and Bad, and had higher vascular density, more proliferating cardiomyocytes, and less apoptotic cells in the infarct and peri-infarct zones. Thus, cardiac-specific and hypoxia-induced coexpression of VEGF and Ang1 improves the perfusion and function of porcine MI heart through the induction of angiogenesis and cardiomyocyte proliferation, activation of prosurvival pathways, and reduction of cell apoptosis.

Toxic effects of a high dose of non-ionic iodinated contrast media on renal glomerular and aortic endothelial cells in aged rats in vivo

Iodinated contrast media (CM) can induce apoptosis and necrosis of renal tubular cells. The injuries of endothelial cells induced by CM on the systemic condition have not been fully understood. To assess the toxic effects of non-ionic CM on the glomerular and aortic endothelial cells, iopromide and iodixanol, two kinds of representative non-ionic CM, were used for the in vivo study. Sixty aged rats were respectively received the agents or normal sodium intravascularly. No obvious apoptosis and morphological change was detected in the glomerular and aortic endothelial cells apart from renal tubules after CM administration. However, expressions of the nitric oxide synthase (eNOS) in glomerular endothelium were decreased at 12h after CM injection. Furthermore, plasma creatinine and endothelin-1 were increased and plasma nitric oxide (NO) was decreased significantly after CM administration. However, we failed to observe the significant increase of plasma von Willebrand Factor. These results suggest that non-ionic iodinated CM do not induce apoptosis and necrosis of glomerular and aortic endothelial cells in vivo. Decreased eNOS expression and increased plasma endothelin-1 may be involved in non-ionic iodinated CM-induced endothelial dysfunction and kidney injury.

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.

Activation of calcium-sensing receptors is associated with apoptosis in a model of simulated cardiomyocytes ischemia/reperfusion

Objective Calcium-sensing receptors (CaSRs) are G-protein coupled receptors which maintain systemic calcium homeostasis and participate in hormone secretion, activation of ion channels, cell apoptosis, proliferation, and differentiation. Previous studies have shown that CaSRs induce apoptosis in isolated adult rat heart and in normal neonatal rat cardiomyocytes by G-protein-PLC-IP3 signaling transduction. However, little knowledge is presently available concerning the role of CaSRs in the apoptosis induced by ischemia and reperfusion in neonatal cardiomyocytes. Methods Primary neonatal rat ventricular cardiomyocytes were incubated in ischemiamimetic solution for 2 h, and then re-incubated in normal culture medium for 24 h to establish a model of simulated ischemia/reperfusion (I/R). Cardiomyocyte apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The expression of CaSRs mRNA was detected by real-time reverse transcription polymerase chain reaction (RT-PCR). In addition, the expressions of caspase-3 and Bcl-2 were analyzed by western blot. Results The simulated I/R enhanced the expression of CaSRs and cardiomyocyte apoptosis. GdCl3, a specific activator of CaSRs, further increased the expression of CaSRs and cardiomyocyte apoptosis, along with up-regulation of caspase-3 and down-regulation of Bcl-2. Conclusion CaSRs are associated with I/R injury and apoptosis in neonatal rat ventricular cardiomyocytes via suppressing Bcl-2 and promoting caspase-3 expression.

Catheter-Based Intramyocardial Delivery (NavX) of Adenovirus Achieves Safe and Accurate Gene Transfer in Pigs

Background Hepatocyte growth factor (HGF) is one of the major angiogenic factors being studied for the treatment of ischemic heart diseases. Our previous study demonstrated adenovirus-HGF was effective in myocardial ischemia models. The first clinical safety study showed a positive effect in patients with severe and diffused triple coronary disease. Methods 12 Pigs were randomized (1∶1) to receive HGF, which was administered as five injections into the infarcted myocardium, or saline (control group). The injections were guided by EnSite NavX left ventricular electroanatomical mapping. Results The catheter-based injections caused no pericardial effusion, malignant arrhythmia or death. During mapping and injection, alanine aminotransferase, aspartate aminotransferase, blood urea nitrogen, serum creatinine and creatine kinase-MB levels have no significant increase as compared to those before and after the injection in HGF group(P>0.05). HGF group has high HGF expression with Western blot, less myocardial infarct sizes by electroanatomical mapping (HGF group versus after saline group, 5.28±0.55 cm2 versus 9.06±1.06 cm2, P<0.01), better cardiac function with Gated-Single Photon Emission Computed Tomography compared with those in saline group. Histological, strongly increased lectin–positive microvessels and microvessel density were found in the myocardial ischemic regions in HGF group. Conclusion Intramyocardial injection guided by NavX system provides a method of feasible and safe percutaneous gene transfer to myocardial infarct regions.

HGF percutaneous endocardial injection induces cardiomyocyte proliferation and rescues cardiac function in pigs.

Objective To investigate the effect of cardiomyocyte proliferation induced by human hepatocyte growth factor (HGF) in pigs with chronic myocardial infarction (CMI). Methods A steerable, deflectable 7F catheter incorporating a 27-guage needle was advanced percutaneously to the left ventricular myocardium of 18 pigs with CMI. Pigs were randomized (1:1:1) to receive adenoviral vector HGF (total dose, 1×1010 genome copies), which was administered as five injections into the infarcted myocardium (total, 1.0 mL), or saline, or Ad-null (control groups). Injections were guided by Ensite NavX left ventricular electroanatomical mapping. HGF and cyclin proteins were detected by western blot and immunoprecipitation analysis. Histological and immunohistochemical analysis determined proliferating cardiomyocytes. Myocardial perfusion and cardiac function were estimated by Gated-Single Photon Emission Computed Tomography (G-SPECT). Results Western blot analyses showed that HGF were predominantly expressed in the infarct core and border in the myocardium of the infarcted heart. G-SPECT analysis indicated that the HGF group had better cardiac function and myocardial perfusion four weeks after the injection of Ad-HGF than before the injection of Ad-HGF. After treatment there were more proliferating cardiomyocytes in the HGF group compared to either of the control groups. Furthermore, the HGF group myocardial samples expressed higher levels of p-Akt, cyclin A, cyclin E, cyclin D1, cdk2, cdk4 than those in the control groups. Conclusion The over-expression of HGF activates pro-survival pathways, induces cardiomyocyte proliferation, and improves the perfusion and function of the porcine CMI heart.

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.