Jing-Song Ou

MicroRNA-21 Regulates Vascular Smooth Muscle Cell Function via Targeting Tropomyosin 1 in Arteriosclerosis Obliterans of Lower Extremities

Objective —The goal of this study was to determine the expression signature and the potential role of microRNAs in human arteries with arteriosclerosis obliterans (ASO). Methods and Results —The expression profiles of microRNAs in human arteries with ASO and in normal control arteries were determined by quantitative reverse transcription–polymerase chain reaction array. Among the 617 detected microRNAs, multiple microRNAs were aberrantly expressed in arteries with ASO. Some of these dysregulated microRNAs were further verified by quantitative reverse transcription–polymerase chain reaction. Among them, microRNA-21 (miR-21) was mainly located in arterial smooth muscle cells (ASMCs) and was increased by more than 7-fold in ASO that was related to hypoxia inducible factor 1-&agr;. In cultured human ASMCs, cell proliferation and migration were significantly decreased by inhibition of miR-21. 3′-Untranslated region luciferase assay confirmed that tropomyosin 1 was a target of miR-21 that was involved in miR-21-mediated cellular effects, such as cell shape modulation. Conclusion —The results suggest that miR-21 is able to regulate ASMC function by targeting tropomyosin 1. The hypoxia inducible factor-1 &agr;/miR-21/tropomyosin 1 pathway may play a critical role in the pathogenesis of ASO. These findings might provide a new therapeutic target for human ASO.

Simvastatin Reduces Myocardial Injury Undergoing Noncoronary Artery Cardiac Surgery A Randomized Controlled Trial

Objective—Myocardial injury during cardiac surgery is a major cause of perioperative morbidity and mortality. We determined whether perioperative statin therapy is cardioprotective in patients undergoing noncoronary artery cardiac surgery and the potential mechanisms. Methods and Results—One hundred fifty-one patients undergoing noncoronary artery cardiac surgery were randomly assigned to either a statin group (n=77) or a control group (n=74). Simvastatin (20 mg) was administered preoperatively and postoperatively. Plasma were analyzed for troponin T, isoenzyme of creatine kinase, C-reaction protein, interleukin-6, interleukin-8, creatinine, and blood urea nitrogen. Cardiac echocardiography was performed. Endothelial nitric oxide synthase (eNOS), Akt, p38, heat shock protein 90, caveolin-1, and nitric oxide (NO) in the heart were detected. Simvastatin significantly reduced plasma troponin T, isoenzyme of creatine kinase, C-reaction protein, blood urea nitrogen, creatinine, interleukin-6, interleukin-8, and the requirement of inotropic postoperatively. Simvastatin increased NO production, the expression of eNOS and phosphorylation at serine1177, phosphorylation of Akt, expression of heat shock protein 90, heat shock protein 90 association with eNOS and decreased eNOS phosphorylation at threonine 495, phosphorylation of p38, and expression of caveolin-1. Simvastatin also improved cardiac function postoperatively. Conclusion—Perioperative statin therapy can improve cardiac function and renal function by reducing myocardial injury and inflammatory response through activating Akt-eNOS and attenuating p38 signaling pathways in patients undergoing noncoronary artery cardiac surgery. Clinical Trial Registration—URL: http://www.clinicaltrials.gov. Unique identifier: NCT01178710.

Anticoagulation Regimens During Pregnancy in Patients With Mechanical Heart Valves: A Systematic Review and Meta-analysis

BACKGROUND Managing anticoagulation in pregnant women with mechanical heart valves remains challenging. Our aim was to evaluate the effectiveness and safety of 4 regimens in these women. METHODS Relevant studies published before June 2015 were collected in several databases and analyzed with RevMan version 5.3 and SPSS version 19.0. Four regimens were defined as follows: a regimen of a vitamin K antagonist (VKA) throughout pregnancy; a heparin (H)/VKA regimen using VKAs except for unfractionated heparin (UFH) or low molecular weight heparin (LMWH) during 6-12 weeks of pregnancy; a LMWH regimen of adjusted LMWH doses throughout pregnancy; and a UFH regimen of adjusted UFH doses throughout pregnancy. The low warfarin dose in the VKA regimen was defined as 5 mg/d or less. RESULTS Fifty-one studies comprising 2113 pregnancies in 1538 women were included. The rate of fetal wastage was significantly higher in the high warfarin dose subgroup than in the low dose one. Compared with the H/VKA regimen, the rate of maternal major thromboembolic event in the low-dose VKA regimen group was significantly lower, although the fetal outcomes were similar. Compared with the H/VKA regimen, the rate of fetal wastage in the LMWH regimen group was significantly lower, and the maternal outcomes were similar. The UFH regimen presented the worst maternal and fetal outcomes. CONCLUSIONS In the absence of large prospective trials, this meta-analysis showed that the VKA regimen should be best for pregnant women with a low warfarin dose, and the H/VKA regimen might be reasonable for those with a high warfarin dose. The LMWH regimen could be used for those who refuse VKA.

C-reactive protein can upregulate VEGF expression to promote ADSC-induced angiogenesis by activating HIF-1α via CD64/PI3k/Akt and MAPK/ERK signaling pathways

BackgroundProliferation of the vasa vasorum has been implicated in the pathogenesis of atherosclerosis, and the vasa vasorum is closely associated with resident stem cells within the vasculature. C-reactive protein (CRP) is positively correlated with cardiovascular disease risk, and our previous study demonstrated that it induces inflammatory reactions of perivascular adipose tissue by targeting adipocytes.MethodsHere we investigated whether CRP affected the proliferation and proangiogenic paracrine activity of adipose-derived stem cells (ADSCs), which may contribute to vasa vasorum angiogenesis.ResultsWe found that CRP did not affect ADSC apoptosis, cell cycle, or proliferation but did increase their migration by activating the PI3K/Akt pathway. Our results demonstrated that CRP can upregulate vascular endothelial growth factor-A (VEGF-A) expression by activating hypoxia inducible factor-1α (HIF-1α) in ADSCs, which significantly increased tube formation on Matrigel and functional vessels in the Matrigel plug angiogenesis assay. The inhibition of CRP-activated phosphorylation of ERK and Akt can suppress CRP-stimulated HIF-1α activation and VEGF-A expression. CRP can also stimulate proteolytic activity of matrix metalloproteinase-2 in ADSCs. Furthermore, CRP binds activating CD64 on ADSCs, rather than CD16/32.ConclusionOur findings implicate that CRP might play a role in vasa vasorum growth by activating the proangiogenic activity of ADSCs.

MiR-142-3p Attenuates the Migration of CD4+ T Cells through Regulating Actin Cytoskeleton via RAC1 and ROCK2 in Arteriosclerosis Obliterans

The migration of CD4+ T cells plays an important role in arteriosclerosis obliterans (ASO). However, the molecular mechanisms involved in CD4+ T cell migration are still unclear. The current study is aimed to determine the expression change of miR-142-3p in CD4+ T cells from patients with ASO and investigate its role in CD4+ T cell migration as well the potential mechanisms involved. We identified by qRT-PCR and in situ hybridization that the expression of miR-142-3p in CD4+ T cells was significantly down-regulated in patients with ASO. Chemokine (C-X-C motif) ligand 12 (CXCL12), a common inflammatory chemokine under the ASO condition, was able to down-regulate the expression of miR-142-3p in cultured CD4+ T cells. Up-regulation of miR-142-3p by lentivirus-mediated gene transfer had a strong inhibitory effect on CD4+ T cell migration both in cultured human cells in vitro and in mouse aortas and spleens in vivo. RAC1 and ROCK2 were identified to be the direct target genes in human CD4+ T cells, which are further confirmed by dual luciferase assay. MiR-142-3p had strong regulatory effects on actin cytoskeleton as shown by the actin staining in CD4+ T cells. The results suggest that the expression of miR-142-3p is down-regulated in CD4+ T cells from patients with ASO. The down-regulation of miR-142-3p could increase the migration of CD4+ T cells to the vascular walls by regulation of actin cytoskeleton via its target genes, RAC1 and ROCK2.

High density lipoprotein from patients with valvular heart disease uncouples endothelial nitric oxide synthase

Normal high density lipoprotein (HDL) protects vascular function; however these protective effects of HDL may absent in valvular heart disease (VHD). Because vascular function plays an important role in maintaining the circulation post-cardiac surgery and some patients are difficult to stabilize, we hypothesized that a deleterious vascular effect of HDL may contribute to vascular dysfunction in VHD patients following surgery. HDL was isolated from age-match 28 healthy subjects and 84 patients with VHD and during cardiac surgery. HDL pro-inflammation index was measured and the effects of HDL on vasodilation, protein interaction, generation of nitric oxide (NO) and superoxide were determined. Patients with VHD received either simvastatin (20mg/d) or routine medications, and endothelial effects of HDL were characterized. HDL inflammation index significantly increased in VHD patients and post-cardiac surgery. HDL from VHD patients and post-cardiac surgery significantly impaired endothelium-dependent vasodilation, inhibited both Akt and endothelial nitric oxide synthase (eNOS) phosphorylation at S1177, eNOS associated with heat shock protein 90 (HSP90), NO production and increased eNOS phosphorylation at T495 and superoxide generation. Simvastatin therapy partially reduced HDL inflammation index, improved the capacity of HDL to stimulate eNOS and Akt phosphorylation at S1177, eNOS associated with HSP90, NO production, reduced eNOS phosphorylation at T495 and superoxide generation, and improved endothelium-dependent vasodilation. Our data demonstrated that HDL from VHD patients and cardiac surgery contributed to endothelial dysfunction through uncoupling of eNOS. This deleterious effect can be reversed by simvastatin, which improves the vasoprotective effects of HDL. Targeting HDL may be a therapeutic strategy for maintaining vascular function and improving the outcomes post-cardiac surgery.

Retinol Binding Protein-Dependent Cholesterol Uptake Regulates Macrophage Foam Cell Formation and Promotes Atherosclerosis.

Background: Retinol-binding protein 4 (RBP4) is an adipokine that plays decisive roles in glucose metabolism and insulin sensitivity. Elevated circulating RBP4 levels were reported to be associated with increased risk for cardiovascular disease, but the precise role of RBP4 in atherosclerotic diseases and its mechanisms of action remain elusive. Methods: Serum RBP4 levels of 1683 participants from South China were evaluated and the occurrence of major adverse cardiovascular events was followed up for 5 years. Apolipoprotein E-deficient mice infected with RBP4-overexpressing/silencing adenovirus, J774A.1 macrophages, and primary peritoneal macrophages from RBP4 transgenic mice were used for investigating the function of RBP4 in foam cell formation. Results: Prospective cohort studies revealed that baseline serum RBP4 level was an independent predictor for incidence of adverse cardiovascular events after adjustment for traditional risk factors. Increased RBP4 expression was observed in atherosclerotic lesions of aortic specimens from both humans and apolipoprotein E-deficient mice, and RBP4 was localized to areas rich in macrophage foam cells. RBP4 inhibition attenuated whereas overexpression accelerated atherosclerosis progression in apolipoprotein E-deficient mice. Both treatment with exogenous recombinant RBP4 and overexpression of RBP4 gene promoted macrophage-derived foam cell formation through the activation of scavenger-receptor CD36-mediated cholesterol uptake, and RBP4 transcriptionally upregulated CD36 expression in a manner dependent on jun N-terminal kinase and signal transducer and activator of transcription 1. The tyrosine kinase c-Src was identified as the upstream regulator of jun N-terminal kinase-signal transducer and activator of transcription 1-mediated CD36-dependent cholesterol uptake, and RBP4 challenge was found to alter the membrane distribution of c-Src and cause c-Src to partition into lipid-raft membrane subdomains, where the kinase was activated. Lastly, Toll-like receptor 4, but not retinol or stimulated by retinoic acid 6, mediated the inductive effects of RBP4 in macrophages. Conclusions: Inclusion of RBP4 levels in traditional models enhances the predictive ability for the incidence of atherosclerotic events. RBP4 promotes atherogenesis by inducing macrophage-derived foam cell formation.

The oxidized phospholipid POVPC impairs endothelial function and vasodilation via uncoupling endothelial nitric oxide synthase.

Endothelial dysfunction is an early stage of atherosclerosis. We recently have shown that 25-hydroxycholesterol found in atherosclerotic lesions could impair endothelial function and vasodilation by uncoupling and inhibiting endothelial nitric oxide synthase (eNOS). 1-Palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (POVPC), the oxidation product of oxidized low-density lipoprotein, is another proinflammatory lipid and has also been found in atherosclerotic lesions. However, whether POVPC promotes atherosclerosis like 25-hydroxycholesterol remains unclear. The purpose of this study was to explore the effects of POVPC on endothelial function and vasodilation. Human umbilical vein endothelial cells (HUVECs) were incubated with POVPC. Endothelial cell proliferation, migration and tube formation were measured. Nitric oxide (NO) production and superoxide anion generation (O2-) were determined. The expression and phosphorylation of endothelial nitric oxide synthase (eNOS), AKT, PKC-βII and P70S6K as well as the association of eNOS and heat shock protein 90 (HSP90) were detected by immunoblotting and immunoprecipitation. Endothelial cell apoptosis was monitored by TUNEL staining. The expression of Bcl-2, Bax, and Cleaved Caspase 3 were detected by immunoblotting. Finally, aortic ring from C57BL6 mice were isolated and treated with POVPC and the endothelium-dependent vasodilation was evaluated. POVPC significantly inhibited HUVECs proliferation, migration, tube formation, decreased NO production but increased O2- generation. POVPC inhibited the phosphorylation of Akt and eNOS at Ser1177, increased activation of PKC-βII, P70S6K and the phosphorylation of eNOS at Thr495, reduced the association of HSP90 with eNOS. Meanwhile, POVPC induced endothelial cell apoptosis by inhibiting Bcl-2 expression, increasing Bax and cleaved caspase-3 expressions as well as caspase-3 activity and impaired endothelium-dependent vasodilation. These data demonstrated that POVPC impaired endothelial function by uncoupling and inhibiting eNOS as well as by inducing endothelial cell apoptosis. Therefore, POVPC may play an important role in the development of atherosclerosis and may be considered as a potential therapeutic target for atherosclerosis.

The Cardioprotective Effect of Vitamin E (Alpha-Tocopherol) Is Strongly Related to Age and Gender in Mice.

Vitamin E (VitE) only prevented cardiovascular diseases in some patients and the mechanisms remain unknown. VitE levels can be affected by aging and gender. We hypothesize that age and gender can influence VitE’s cardioprotective effect. Mice were divided into 4 groups according to age and gender, and each group of mice were divided into a control group and a VitE group. The mice were administered water or VitE for 21 days; Afterward, the cardiac function and myocardial infarct size and cardiomyocyte apoptosis were measured after myocardial ischemia reperfusion(MI/R). VitE may significantly improved cardiac function in young male mice and aged female mice by enhancing ERK1/2 activity and reducing JNK activity. Enhanced expression of HSP90 and Bcl-2 were also seen in young male mice. No changes in cardiac function and cardiac proteins were detected in aged male mice and VitE was even liked to exert a reverse effect in cardiac function in young mice by enhancing JNK activity and reducing Bcl-2 expression. Those effects were in accordance with the changes of myocardial infarction size and cardiomyocyte apoptosis in each group of mice. VitE may reduce MI/R injury by inhibiting cardiomyocyte apoptosis in young male mice and aged female mice but not in aged male mice. VitE was possibly harmful for young female mice, shown as increased cardiomyocyte apoptosis after MI/R. Thus, we speculated that the efficacy of VitE in cardiac protection was associated with age and gender.

Disruption of calcium homeostasis by cardiac-specific over-expression of PPAR-γ in mice: A role in ventricular arrhythmia.

AIMS Adverse cardiovascular effects induced by peroxisome proliferator activator receptor-γ (PPAR-γ) activation were observed in clinical setting. But the underlying mechanism is unclear. Now, transgenic mice with cardiac specific peroxisome proliferator activator receptor-γ overexpression (TG-PPAR-γ) were used to explore the possible mechanisms. MATERIALS AND METHODS Cardiac tissues from TG-PPAR-γ mice, a PPAR-γ over-expressing human cardiomyocyte line AC16 cell, and PPAR-γ agonist-treated primary cardiomyocytes were used to evaluate the expression of cardiac calcium regulatory proteins as sarcoplasmic reticulum Ca2+ ATPase, Na+/Ca2+ exchanger 1, ryanodine receptor 2 and phospholamban. Intracellular Ca2+ levels were also examined by flow cytometry and confocal microscopy with Fluo-4/AM in these cells. KEY FINDINGS In this study, frequent ventricular premature contraction and polymorphic ventricular tachycardia were observed in TG-PPAR-γ but not in wild-type mice. Besides, we found the calcium regulatory proteins expression were higher in the TG-PPAR-γ mice, PPAR-γ overexpressing human cardiomyocyte line AC16 cell and PPAR-γ agonist-treated primary cardiomyocytes than the control group respectively. In addition, an increase of intracellular calcium levels and CaMKII δ expression in PPAR-γ overexpression and PPAR-γ activation group. Moreover, Inhibition of CaMKII δ could improve the intracellular calcium levels and reduce the occurrence of ventricular arrhythmia. SIGNIFICANCE PPAR-γ over-expression perturbs the intracellular calcium homeostasis in cardiomyocytes which contribute to the ventricular arrhythmias and cardiac sudden death in TG-PPAR-γ mice.