Wen-Zhu Ma

VEGF165 and angiopoietin-1 decreased myocardium infarct size through phosphatidylinositol-3 kinase and Bcl-2 pathways

Angiogenic growth factors, vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang1) could decrease myocardial infarct size, which was assumed to be related with newly formed capillaries. We doubted that these capillaries could do this solely and the potential protective mechanisms of VEGF and Ang1 on myocardium need to be evaluated. Three types of adenoviruses encoding human VEGF165 (Ad-VEGF165), human angiopoietin-1 (Ad-Ang1) and green fluorescent protein (Ad-GFP, as a parallel control) were constructed. Experiments were taken both in vitro and in vivo. As in vitro, the antiapoptosis effect of VEGF165, Ang1 and VEGF165+Ang1 on cardiac myoblasts was observed, which seemed to be related with the activation of phosphatidylinositol-3 kinase and Bcl-2 pathways. As in vivo, adenoviruses were intramyocardially injected immediately after the ligation of the left anterior descending coronay arteries in rats. The results showed positive effect of VEGF165, Ang1 and VEGF165+Ang1 on decreasing the myocardial infarct size at the 7th day. Myocardial PI-3K activity and Bcl-2 expression were elevated relatively at the 3rd day. The protective effect of VEGF165 and Ang1 on the myocardium may broaden their functional research and contribute to their clinical use in the future.

A novel approach to transplanting bone marrow stem cells to repair human myocardial infarction: delivery via a noninfarct-relative artery.

Bone marrow stem cells are able to repair infarcted human myocardium following intracoronary transplantation via the infarct-relative artery. However, traditional reperfusion strategies fail to open the artery in some patients, making effective delivery impossible. Our previous study demonstrated a safe and efficient approach to delivering bone marrow stem cells via a noninfarcted artery in an animal myocardial infarction model. The objective of the present study was to evaluate the safety and feasibility of autologous bone marrow mesenchymal stem cell transplantation via such an approach in patients with acute myocardial infarction (AMI). Sixteen patients with anterior AMI who had successfully undergone percutaneous coronary intervention (PCI) were enrolled in this pilot, randomized study. Three weeks after PCI, cultured bone marrow mesenchymal stem cells were injected into the myocardium via either the infarct-relative artery (left anterior descending branch artery, LAD) or a noninfarct-relative artery (right coronary artery, RCA). The safety and feasibility of the cell infusion were evaluated during the procedure and during 6 months of follow-up. In addition, 2D echocardiography, technetium-99m methoxyisobutylisonitrile (99mTc-MIBI) and 18F-deoxyglucose single photon emission computed tomography were employed to examine cardiac function, myocardial perfusion, and viable cardiomyocytes, respectively, at day 4 after PCI and 6 months after the cell infusion. There were no arrhythmia and any other side-effects, including infections, allergic reactions or adverse clinical events, during, immediately after, or 6 months after cell transplantation. Cardiac function and myocardial perfusion had improved 6 months after PCI/bone marrow stem cells transplantation. Viable cardiomyocytes metabolism was detected in the infarcted areas in both groups after the cell infusion, as demonstrated by 18F-deoxyglucose. Intracoronary infusion of autologous bone marrow mesenchymal stem cells via a noninfarct-relative artery appears safe and feasible in the treatment of patients with AMI.

Experimental study of bone marrow-derived mesenchymal stem cells combined with hepatocyte growth factor transplantation via noninfarct-relative artery in acute myocardial infarction

We investigated the impact of bone marrow-derived mesenchymal stem cells (BM-MSCs) alone or in combination with hepatocyte growth factor (HGF) transplantation via noninfarct-relative artery in a swine myocardial infarction (MI) model. Donor BM-MSCs were derived in vitro from swine auto-bone marrow cultures labeled by bromodeoxyuridine (BrdU) incorporation. Host MI swine model was created by ligating the distal left anterior descending artery. After 4 weeks, age-matched male MI swines were used for the transplantation. Male MI swines were transfused via noninfarct-relative artery with vehicle (control, n=6) or BrdU-labeled BM-MSCs (5 × 106) alone (MSCs, n=6) or BrdU-labeled BM-MSCs (5 × 106) combined with HGF (4 × 109 PFU) (MSCs+HGF, n=6). To evaluate the collateral artery growth (Rentrop) and cardiac perfusion in these animals, gate cardiac perfusion imaging and coronary angiography were performed before and 4 weeks after transplantation, respectively. To assess the contribution of donor-originated cells in stimulation of cardiomyocyte regeneration and angiogenesis, immunohistochemistry for BrdU and α-smooth muscle actin (α-SMA) and quantitative image analysis were performed at 4 weeks after transplantation. The results are as follows: (1) BrdU-positive cells were detected in host myocardium in both MSCs and MSCs+HGF groups, but not in the vehicle group. Most BrdU-positive cells expressed myosin heavy chain β. (2) α-SMA-positive arteriole densities in the infarcted border area and infarcted area were increased significantly in both transplantation groups compared with the vehicle group. (3) Gate cardiac perfusion imaging demonstrated that the cardiac perfusion was significantly improved in transplantation groups compared with the vehicle group. (4) Ejection fraction and α-SMA-positive arteriole densities were increased significantly in both transplantation groups compared with the vehicle group. However, there was no difference in ejection fraction and α-SMA-positive arteriole densities between the MSCs group and the MSCs+HGF group. Growth of collateral arteries was not detected by coronary angiography in all three groups. In conclusion, the current study indicates that BM-MSCs transplantation via noninfarct-relative artery stimulates cardiomyocyte regeneration and angiogenesis and improves cardiac function, but does not stimulate collateral artery growth. BM-MSCs transplantation combined with HGF therapy is not superior to BM-MSCs alone transplantation. BM-MSCs transplantation via noninfarct-relative artery may be an alternative for those patients who cannot be transplanted via infarct-relative artery in clinical practice.

Testosterone is negatively associated with the severity of coronary atherosclerosis in men.

This study aimed to determine whether plasma testosterone is associated with the severity of coronary atherosclerosis in a group of 803 men who underwent elective coronary angiography. Testosterone levels were measured in 803 male patients who were categorized into three groups according to testosterone level tertiles. All patients underwent elective coronary angiography, and the severity of coronary artery disease (CAD) was determined by the Gensini score. Moreover, patients were classified into two groups according to Gensini scores (score ≤26 and score >26) using the median values as cutoff points. The plasma testosterone levels were measured by an ELISA kit. The level of testosterone was negatively associated with the Gensini score (r=-0.188; P=0.000). A multiple linear regression analysis revealed that testosterone was an independent risk factor for the Gensini score (β=-0.110; P=0.002) after adjusting for confounding covariates. In a multivariate logistic regression model, the severity of CAD was shown to be significantly lower in the third tertile (highest) of testosterone compared to the first tertile (lowest) of testosterone (odds ratio (OR)=0.465; 95% confidence interval (CI): 0.327-0.662; P=0.000). In this study, patients with lower testosterone levels had higher Gensini scores in a group of 803 men who underwent elective coronary angiography. Additional studies are needed to clarify the direction of causality and possible underlying mechanisms.

HEPATOCYTE GROWTH FACTOR PLAYS A CRITICAL ROLE IN THE REGULATION OF CYTOKINE PRODUCTION AND INDUCTION OF ENDOTHELIAL PROGENITOR CELL MOBILIZATION: A PILOT GENE THERAPY STUDY IN PATIENTS WITH CORONARY HEART DISEASE

1 There is growing evidence of the beneficial effects of hepatocyte growth factor (HGF) in myocardial infarction, heart failure and occlusive peripheral arterial disease. The aim of the present study was to evaluate the effects of intracoronary administration of an adenovirus vector encoding the human HGF gene (Ad‐HGF) on serum levels of cytokines and mobilization of CD34+ and CD117+ cells in patients with coronary heart disease. 2 Twenty‐one patients with severe coronary artery disease were recruited to the study: 11 patients received both a stent and administration of Ad‐HGF; the remaining 10 patients received a stent alone and served as the control group. Blood samples were obtained from the femoral vein before and then 6 and 24 h, 3 and 6 days and 2 weeks after treatment for the isolation of serum and peripheral blood mononuclear cells. Intracoronary administration of Ad‐HGF in patients with coronary heart disease resulted in high levels of HGF gene expression, as well as its receptor c‐met, compared with the control group, as demonstrated by real‐time reverse transcription–polymerase chain reaction. In addition, serum levels of HGF, vascular endothelial growth factor, monocyte chemoattractant protein‐1 and interleukin (IL)‐10 were increased and serum levels of IL‐8 were decreased in patients administered Ad‐HGF compared with the control group. The percentage of CD34+ and CD117+ cells in the peripheral blood increased in patients administered Ad‐HGF. 3 In conclusion, HGF gene therapy may play an important role in the regulation of cytokines and the induction of endothelial progenitor cell mobilization in patients with coronary heart disease.

Induction of collateral artery growth and improvement of post-infarct heart function by hepatocyte growth factor gene transfer

AbstractAim:To study the effect of adenovirus5-mediated human hepatocyte growth factor (Ad5-HGF) transfer on post-infarct heart failure in a swine model.Methods:Twelve young Suzhong swine were randomly divided into 2 groups: the Ad5-HGF group (n=6) and the null-Ad5 group (n=6). Four weeks after left anterior descending coronary artery (LAD) ligation, Ad5-HGF was transferred into the myocardium via the right coronary artery. Coronary angiography and gated cardiac perfusion imaging were performed at the end of 4 and 7 weeks after LAD ligation, respectively, to evaluate collateral artery growth and cardiac perfusion. Then all animals were killed, the expression of HGF and α-smooth muscle actin (α-SMA) were evaluated by enzyme-linked immunosorbent assay and immunohistochemistry.Results:Compared with the null-Ad5 group, higher expression of human HGF was observed in the myocardium in the Ad5-HGF group (109.3±7.8 vs 6.2±2.6, t=30.685, P<0.01). The left ventricular ejection fraction was higher in the Ad5-HGF group than in the null-Ad5group (43.9±4.3 vs 30.4±2.8, t=6.514, P<0.01). From the 4th week to the 7th week after operation, left ventricular end systolic volume (42.1±3.0 vs 31.0±4.9, t=12.800, P<0.01) and left ventricular end diastolic volume (62.2±4.2 vs 55.0±4.8 t=13.207, P<0.01) were improved in the Ad5-HGF group. Cardiac perfusion was significantly improved in the Ad5-HGF group. In the Ad5-HGF group, growth of collateral arteries was obviously greater (average rank sum 9.17 vs 3.83, n=6, u=-2.687, P<0.01), and the number of α-SMA+ vessels/mm2 was significantly greater (56.1±4.2 vs 16.4±3.5, t=17.731, P<0.01) than in the null-Ad5 group.Conclusion:High expression levels of human HGF were observed in the myocardium because of non-infarct-related vessel transfer. HGF can increase the number of functional arterioles and improve collateral artery growth. HGF can improve cardiac perfusion and heart function.

Association of SNP Rs6903956 on Chromosome 6p24.1 with Angiographical Characteristics of Coronary Atherosclerosis in a Chinese Population

Objective To explore the association between rs6903956 and severity of coronary artery disease (CAD) in a Chinese population. Methods A cohort of 1075 consecutive patients who underwent coronary arteriography for suspected or known coronary atherosclerosis was enrolled in our study. Coronary atherosclerosis severity was defined by Gensinis Score System and counts of diseased vessels. Results Gensini score frequencies and counts of diseased vessels differed among GG, AG, AA genotype groups at the rs6903956 locus (p = 0.025 for Gensini score frequencies vs. p = 0.024 for counts of diseased vessels, respectively). A univariate logistic regression analysis revealed that the genotype distribution of this SNP was associated significantly with angiographical characteristics of coronary atherosclerosis risk (p = 0.030, odds ratio (OR) = 1.444, 95% confidence interval (CI) = 1.036∼2.013 for AG vs. GG; p = 0.021, OR = 5.896, 95% CI = 1.299∼26.750 for AA vs. GG and p = 0.007, OR = 1.564, 95% CI = 1.132∼2.162 for combined (AG+AA) vs. GG). A multivariate logistic regression analysis indicated that the genotype distribution of the rs6903956 polymorphism be associated significantly with the angiographical characteristics of coronary atherosclerosis risk (p = 0.004, OR = 1.578, 95% CI = 1.155∼2.154 for GG vs. AG vs. AA; p = 0.013, OR = 1.541, 95% CI = 1.097∼2.163 for GG vs. GA+ AA). A stratification analysis revealed that male subjects and smoking subjects had a higher frequency of the rs6903956 heterozygous mutant among higher Gensini score subjects than among lower Gensini score subjects (p = 0.023, OR = 1.579, 95% CI = 1.064∼2.344 for male subgroup; p = 0.005, OR = 2.075, 95% CI = 1.249∼3.448 for smoking subgroup). Conclusions Allele A is a risk factor for CAD and the G-to-A allele substitution may underlie the association between rs6903956 and CAD.

Renin-Angiotensin-Aldosterone System Gene Polymorphisms and Coronary Artery Disease: Detection of Gene-Gene and Gene-Environment Interactions

Objective: The objective of this study was to explore the association between coronary artery disease and genetic polymorphisms of the renin–angiotensin–aldosterone system (RAAS) pathway. In addition, we examined the interactions between demographic and lifestyle risk factors (environmental factors including age, sex, smoking status, alcohol intake) and RAAS polymorphisms on disease risk. Methods: A total of 1089 subjects who underwent coronary angiography were enrolled in this study. Eight RAAS polymorphisms were genotyped in this population: the G2350A (rs4343) polymorphism in exon 17 of the angiotensin converting enzyme (ACE) gene, 1166A→C (rs5186) and 573C/T (rs5182) in the angiotensin II type 1 receptor (AGTR1) gene, the -344C→T transversion (rs1799998) in the aldosterone synthase (CYP11B2) gene, and the G-217A (rs5049), G-6A (rs5051), M235T (rs699; T4072C), and T174M (rs4762; C3889T) polymorphisms in the angiotensinogen (AGT) gene. Subjects with coronary heart disease were defined as those with at least 50% stenosis in at least one major coronary artery, and, the severity of coronary atherosclerosis was defined by the Gensini scoring system. Results: Compared to the subjects with AA genotype, the subjects with AG + GG genotype of rs1799998 had significant lower gensini score (p=0.029). After adjusting for age, gender, cigarette smoking, and alcohol intake status, the AG genotype (OR 0.717 95%CI 0.541–0.950, p=0.021) and the AG + GG genotype (OR 0.730 95%CI 0.559–0.954, p=0.021) distributions of rs1799998 were significantly different between the cases and controls compare to the AA genotype. Subjects with three at-risk loci had increased risk of coronary artery disease compared to subjects carrying 0 or 1 risk-associated polymorphism (OR [95% CI]:1.579 [1.077–2.316], p=0. 019), and the significance of the association was not reduced after adjusting for age, sex, cigarette smoking, or alcohol intake (adjusted OR [95% CI]: 1.673 [1.116–2.507], p=0.013). The results of multifactor-dimensionality reduction analysis revealed an interaction effect of CYP11B2 -344C→T, age, and smoking status on the risk of coronary heart disease (training OR [95% CI]: 3.7685 [2.8463–4.9895], p<0.0001; testing OR [95% CI]: 2.7583 [1.2038–6.3203], p=0.015). Conclusions: Subjects who carried the G allele of the rs1799998 polymorphism significantly associated with coronary heart disease and severity of coronary atherosclerosis estimated by the Gensini score in the whole population of the study. And, multiple RAAS gene polymorphisms are associated with coronary artery disease. The interaction of the CYP11B2 -344C→T polymorphism (rs1799998), age, and smoking status is also associated with enhanced risk of coronary artery disease.

Interaction between microRNA expression and classical risk factors in the risk of coronary heart disease.

The aim of this study was to identify the synergistic effect of microRNA expression with classical risk factors of coronary heart disease (CHD) and to explore their diagnostic value for coronary stenotic lesions in subjects with CHD. Plasma samples were obtained from 66 subjects with CHD and from 58 control individuals. A quantitative reverse-transcription PCR (RT-qPCR) assay was conducted to confirm the relative expressions of the known CHD-related miRNAs. The severity of coronary atherosclerosis was based on the Gensini scoring system. The expression of miR-125b in plasma of the CHD group was lower than that of the non-CHD group (0.14 ± 0.09 vs. 0.18 ± 0.10, p = 0.055), and the miR-125b levels significantly decreased following an increasing Gensini score (P = 0.037). Spearman correlation analyses indicated the Gensini score was negatively associated with miR-125b (r = −0.215, p = 0.017). Of all the miRNAs, miR-125b showed the lowest AUC (0.405; 95% CI: 0.305 ~ 0.506, p = 0.070). We found several synergistic effects between miR-125b and classical risk factors, such as age, sex, CR, FBG and HDL-C; the proportion of CHD attributable to the interaction of miR-125b and age was as high as 80%. Therefore, miR-125b was shown to play an important role in individual’s susceptibility to developing CHD.

Proinsulin Is an Independent Predictor of the Angiographical Characteristics of Coronary Atherosclerosis

Objective: The objective of this survey was to study the association between circulating proinsulin level and the severity of coronary atherosclerosis. Methods: The study population consisted of 1,039 consecutive patients (775 males and 264 females) who underwent coronary angiography for suspected or known coronary atherosclerosis. The patients’ anthropometric and plasma measurements including body mass index, blood pressure, blood lipid, blood glucose, and proinsulin level were performed. The severity of coronary atherosclerosis was defined by the Gensini’s score system. Results: When proinsulin level was examined as a categorical variable classified by quartile values, subjects with a high proinsulin level had significantly higher values of Gensini’s score than those with a low proinsulin level (p = 0.000). The Spearman correlation analysis suggests that the Gensini’s score was significantly correlated with proinsulin level (r = –0.177, p = 0.000). Multiple stepwise linear regression analysis demonstrated that glucose (β= 0.136, p = 0.000), age (β= 0.189, p = 0.000), proinsulin (β= 0.135, p=0.000), SBP (mm Hg) (β= –0.061, p = 0.043), fasting low-density lipoprotein cholesterol (mmol/l) (β= 0.141, p = 0.003), and total cholesterol (β= –0.105, p = 0.029) are significantly independently associated with the Gensini’s score. Conclusions: Proinsulin is a strong and statistically highly significant pre- dictor of coronary atherosclerosis independent of the other major risk factors including age, body mass index, blood pressure, fasting blood glucose, and blood lipid. The exact mechanisms need further study.