Ji-Peng Zhou

4-HNE increases intracellular ADMA levels in cultured HUVECs: evidence for miR-21-dependent mechanisms.

Objective To investigate whether 4-hydroxynonenal (4-HNE) regulates asymmetric dimethylarginine (ADMA) metabolism through pathway independent of direct adduct formation with ADMA metabolizing enzyme and the involvement of microRNA (miRNA) miR-21 in human umbilical venous endothelial cells (HUVECs). Methods Cultured HUVECs were treated with 4-HNE (at concentrations of 1, 5, and 10 µM, respectively) or 1‰ DMSO (vehicle control) for 24 h. MiR-21 inhibitor (final concentration of 100 nM) was transfected at 1 h before 4-HNE treatment. HUVECs were also transfected with miR-21 (at concentrations of 50 nM and 100 nM) and cultured for 12, 24, and 48 h, respectively. DDAH mRNA and miR-21 expression in the HUVECs were determined by semi-quantitative real time PCR. DDAH1 and DDAH2 protein expression were analyzed by Western blot. ADMA in the cell medium and cell lysates were analyzed by ELISA. ADMA metabolizing activity of the cell lysates was also determined. Results MiR-21 decreased DDAH1 and DDAH2 expression and ADMA metabolic activity significantly, while increased intracellular ADMA accumulation significantly in HUVECs. 10 µM 4-HNE treatment for 24 h increased the expression of miR-21 and intracellular ADMA concentration, decreased the expression of DDAH1/2 mRNA and protein, decreased ADMA metabolizing activity of the cell lysates significantly. MiR-21 inhibitor reversed the inhibitory effects of 4-HNE on DDAH1 expression completely, and partially reversed the changes in ADMA metabolizing activity and intracellular ADMA accumulation challenged by 10 µM 4-HNE. Conclusion 4-HNE down-regulates DDAH1 expression and increases intracellular ADMA accumulation in HUVECs through a miR-21-dependent mechanism.

Correlations of DDAH1 Transcript Variants with Human Endothelial Asymmetric Dimethylarginine Metabolizing Activity

BACKGROUND Dimethylarginine dimethylaminohydrolases 1 (DDAH1) is the major enzyme responsible for inactivation of asymmetric dimethylarginine (ADMA). This study seeks to clarify the correlations between mRNA expression levels of DDAH1 transcript variants and the relationship with ADMA metabolizing activity in human. METHODS The mRNA expression levels of DDAH1 transcript variants in primarily cultured human umbilical vein endothelial cells (HUVECs) and peripheral blood mononuclear cells (PBMCs) from healthy control subjects and patients suffering from both acute ischemic stroke (AIS) and acute myocardial infarction (AMI) were determined by real-time polymerase chain reaction. ADMA metabolizing activity of the cell lysates from HUVECs was determined by enzyme-linked immunosorbent assay. RESULTS A novel DDAH1 transcript variant DDAH1-V3 was identified. DDAH1-V3 mRNA expression correlated significantly with that of both -V2 (R = 0.811; P = 0.000008) and -V1 (R = 0.454; P = 0.04) in HUVECs. In PBMCs from healthy subjects, significant correlation was observed only between DDAH1-V2 and -V3 (R = 0.571; P = 0.001; n = 36). Delta threshold cycle (DCT) values for both DDAH1-V2 and -V3 transcripts were increased significantly in PBMCs from AIS patients (P < 0.05, respectively). In PBMCs from patients suffering from both AIS and AMI, positive pairwise correlations between mRNA levels of DDAH1 transcripts were also observed as analyzed by partial correlation analysis (P < 0.05, respectively). However, only mRNA expression level of the DDAH1-V1 transcript correlated significantly with intracellular ADMA metabolizing activity in HUVECs (R = 0.805; P=0.002). CONCLUSIONS This study demonstrated that although there are positive correlations between mRNA expression levels of DDAH1 transcript variants, only the DDAH1-V1 transcript is responsible for ADMA metabolism, and transcript specific primers are recommended to determine DDAH1 mRNA expression.

MiR-15b-5p Regulates Collateral Artery Formation by Targeting AKT3 (Protein Kinase B-3)Highlights

Objective— To identify circulating microRNAs that are differentially expressed in severe coronary heart disease with well or poorly developed collateral arteries and to investigate their mechanisms of action in vivo and in vitro. Approach and Results— In our study, we identified a circulating microRNA, miR-15b-5p, with low expression that, nevertheless, characterized patients with sufficient coronary collateral artery function. Moreover, in murine hindlimb ischemia model, in situ hybridization identified that miR-15b-5p was specifically expressed in vascular endothelial cells of adductors in sham group and was remarkably downregulated after femoral artery ligation. Overexpressed miR-15b-5p significantly inhibited arteriogenesis and angiogenesis in mice. In vitro, both under basal and vascular endothelial growth factor stimulation, loss-of-function or gain-of-function studies suggested that miR-15b-5p significantly promoted or depressed the migration and proliferation of endothelial cells. We identified AKT3 (protein kinase B-3) as a direct target of miR-15b-5p. Interestingly, AKT3 deficiency by injection with Chol-AKT3-siRNA obviously suppressed arteriogenesis and the recovery of blood perfusion after femoral ligation in mice. Conclusions— These results indicate that circulating miR-15b-5p is a suitable biomarker for discriminating between patients with well-developed or poorly developed collaterals. Moreover, miR-15b-5p is a key regulator of arteriogenesis and angiogenesis, which may represent a potential therapeutic target for ischemic disease.

Plasma Omentin-1 Level as a Predictor of Good Coronary Collateral Circulation

Aims: Coronary collateral circulation (CCC) is crucial during an acute ischemic attack. Evidences showed that omentin-1 exhibited remarkable antiatherogenic effects and ischemia-induced revascularization. The aim of this study was to investigate the relationship between plasma omentin-1 levels and CCC in patients with ≥ 90% angiography-proven coronary occlusion. Methods: 142 patients with ≥ 90% luminal diameter stenosis in at least one major epicardial coronary artery were recruited. Among them, 79 patients with Rentrop 0–1 grade were classified into the poor CCC group and 63 patients with Rentrop 2–3 grade were included into the good CCC group. The association between plasma omentin-1 levels and CCC status was assessed. Results: Plasma omentin-1 level was significantly higher in patients with good CCC than those with poor CCC (566.57 ± 26.90 vs. 492.38 ± 19.70 ng/mL, p = 0.024). Besides, omentin-1 was positively correlated with total cholesterol (TC), high-density lipoprotein, and gensini score but inversely with hyperlipidemia and body mass index (all p values < 0.05). Multivariate regression analysis indicated that omentin-1 [odds ratio (OR) = 1.002, 95% confidence interval (CI): 1.000 – 1.004, p = 0.041)], TC, the number of the diseased vessels, a higher frequency of left circumflex artery and right coronary artery, chronic total occlusion, and gensini score remained as the independent predictors of good CCC. Conclusion: Higher plasma omentin-1 level was associated with better CCC development. Our findings suggest that omentin-1 may be an alternative marker for adequate CCC in patients with ≥ 90% coronary occlusion.

Basigin rs8259 Polymorphism Confers Decreased Risk of Chronic Heart Failure in a Chinese Population

Left ventricular remodeling is an essential risk factor contributing to the pathogenesis of chronic heart failure (CHF). Basigin (BSG) promotes cardiovascular inflammation and myocardial remodeling processes by induction of extracellular matrix metalloproteinases and inflammatory cytokines. BSG rs8259 polymorphism was associated with BSG expression and risk of acute coronary syndrome. Therefore, we investigated whether rs8259 polymorphism contributes to risk and prognosis of CHF in Chinese patients. In total 922 adult patients with CHF and 1107 matched healthy controls were enrolled. BSG rs8259 polymorphism was genotyped using PCR-restriction fragment length polymorphism. Whole blood BSG mRNA expression data from Genotype-Tissue Expression project was accessed. Evaluation of follow-up data was performed in only 15.2% (140) of the patients with CHF. BSG rs8259 TT genotype was associated with a decreased risk of CHF (OR = 0.83, 95% CI = 0.72–0.96, p = 0.010), especially in patients with hypertension (OR = 0.80, 95% CI = 0.68–0.95, p = 0.011) and coronary heart disease (OR = 0.81, 95% CI = 0.69–0.96, p = 0.013) after adjustment for multiple cardiovascular risk factors. Rs8259 T allele was associated with decreased BSG mRNA in whole blood from 338 healthy normal donors (p = 1.31 × 10−6). However, rs8259 polymorphism failed to exhibit an association with cardiovascular mortality (p = 0.283). BSG rs8259 polymorphism may contribute to decreased risk of CHF in a Chinese Han population.

Association of CKIP-1 P21A polymorphism with risk of chronic heart failure in a Chinese population

Pathological cardiac hypertrophy is an independent risk factor for chronic heart failure. Casein kinase-2 interacting protein-1 (CKIP-1) can inhibit pathological cardiac hypertrophy. Therefore, we investigated whether CKIP-1 nonsynonymous polymorphism rs2306235 (Pro21Ala) contributes to risk and prognosis of chronic heart failure in a Chinese population.A total of 923 adult patients with chronic heart failure and 1020 age- and gender-matched healthy controls were recruited. CKIP-1 rs2306235 polymorphism was genotyped using PCR-restriction fragment length polymorphism. Additional follow-up data for 140 chronic heart failure patients was evaluated. The rs2306235 G allele was associated with an increased risk of chronic heart failure (OR = 1.38, 95% CI = 1.09-1.75, p = 0.007), especially in patients with hypertension (OR = 1.45, 95% CI = 1.09-1.75, p = 0.006) and coronary heart disease (OR = 1.41, 95% CI = 1.09-1.83, p = 0.010) after adjustment for multiple cardiovascular risk factors. However, rs2306235 polymorphism was not associated with cardiovascular mortality in chronic heart failure (p = 0.875). CKIP-1 rs2306235 polymorphism may be a risk factor for chronic heart failure in a Chinese Han population.

Association of NPR3 polymorphism with risk of essential hypertension in a Chinese population

Essential hypertension (EH) is a common disease exhibiting large individual difference in occurrence, development and treatment response. Genetic factors are implicated in the development and progression of EH. This study aimed to explore the association between NPR3 single nucleotide polymorphism rs2270915 (A/G, Asn521Asp) and the risk of EH in a Chinese Han population by a case‐control study.