Xinqun Hu

Mesenchymal stem cells modified with miR-126 release angiogenic factors and activate Notch ligand Delta-like-4, enhancing ischemic angiogenesis and cell survival

The endothelial cell-specific microRNA (miRNA), miR-126, is considered a master regulator of physiological angiogenesis. Transplanted mesenchymal stem cells (MSCs) release soluble factors contributing to neoangiogenesis and cardiac repair. Therefore, we hypothesized that the over-expression of miR-126 may prolong MSC survival and enhance the cell secretome, thereby improving post-infarction angiogenesis and cardiac function. In this study, MSCs harvested from male C57BL/6 mouse bone marrow were infected in vitro with miR-126 (MSC(miR-126)) by using recombinant lentiviral vectors; the control cells were either non-transfected or transduced with mock vectors (MSC(null)). The results showed an increased secretion of angiogenic factors and a higher resistance against hypoxia in MSC(miR-126) compared with the control cells. The expression of the Notch ligand Delta-like (Dll)-4 in the MSC(miR-126) group was also increased. For in vivo experiments, MSC(miR-126) cultures were intramyocardially injected into the infarct region of the hearts of female C57BL/6 mice (an acute myocardial infarction model) who had undergone ligation of the left anterior descending coronary artery. The survival of MSC(miR-126) cultures, determined by Sry expression, was increased at 7 days after transplantation. MSC(miR-126)-treated animals showed significantly improved cardiac function as assessed by echocardiography 2 weeks later. The expression levels of angiogenic factors and Dll-4 in the infarcted myocardium were further increased by MSC(miR-126) compared with MSCs or MSC(null) cultures. Furthermore, fluorescent microsphere and histological studies revealed that myocardial blood flow and microvessel density were significantly increased in the MSC(miR-126)-transplanted animals. In addition, we found increased immature vessel proliferation following the transplantation of MSC(miR-126) cultures in which the expression of Dll-4 had been knocked down. However, blood vessels with lumen were barely detected, which indicated that Dll-4 plays a key role in tubulogenesis. We conclude that the transplantation of MSCs overexpressing miR-126 can further enhance functional angiogenesis in the ischemic myocardium possibly by the secretion of angiogenic factors and the activation of Dll-4, thus increasing MSC survival. Therefore, MSCs modified with miR-126 may represent a novel and efficient therapeutic approach for ischemic angiogenesis and the improvement of cardiac function.

Overexpression of MicroRNA-1 Improves the Efficacy of Mesenchymal Stem Cell Transplantation after Myocardial Infarction

Background: The aim of this research was to study whether transplantation of mesenchymal stem cells (MSCs) overexpressing microRNA-1 into mouse infarcted myocardium can enhance cardiac myocyte differentiation and improve cardiac function efficiently. Methods: Eight-week-old female C57BL/6 mice underwent ligation of the left coronary artery to produce models of myocardial infarction. The ligated animals were randomly divided into 4 groups (20 in each). One week later, they were intramyocardially injected at the heart infarcted zone with microRNA-1-transduced MSCs (MSCmiR-1 group), mock-vector-transduced MSCs (MSCnull group), MSCs (MSC group) or medium (PBS group). At 4 weeks post-transplantation, transthoracic echocardiographic assessment, histological evaluation and Western blot were performed. Results: The transplanted MSCs were able to differentiate into cardiomyocytes in the infarcted zone. Cardiac function in the MSC, MSCnull and MSCmiR-1 groups was significantly improved compared to the PBS group (p < 0.01 or p < 0.001). However, treatment of MSCs expressing microRNA-1 was more effective for cardiac repair and improved cardiac function more efficiently by enhancing cell survival and cardiac myocyte differentiation compared to the MSC group or the MSCnull groups (p < 0.05 or p < 0.01, respectively). Conclusions: Transplantation of microRNA-1-transfected MSCs was more conducive to repair of infarct injury and improved heart function by enhancing transplanted cells survival and cardiomyogenic differentiation.

Overexpression of miR-126 promotes the differentiation of mesenchymal stem cells toward endothelial cells via activation of PI3K/Akt and MAPK/ERK pathways and release of paracrine factors.

Abstract The endothelial cell (EC)-specific miRNA, miR-126, is known to promote angiogenesis in response to angiogenic factors by repressing negative regulators of signal transduction pathways; however, whether miR-126 might regulate the differentiation of stem cells toward endothelial lineage remains unknown. To answer this question, in this study mesenchymal stem cells (MSCs) harvested from C57BL/6 mouse bone marrow were transfected with miR-126 (MSCmiR-126) using recombinant lentiviral vectors. Results showed the para-secretion and the expression levels of phosphorylated PI3K p85, Akt, p38, ERK1 protein in the MSCmiR-126 group were dramatically increased when compared with the control group. With half culture medium refreshed every 3 days, a small number of 6-day-cultured MSCmiR-126 differentiated into endothelial-like cells and most of 9-day-cultured MSCmiR-126 formed a cobblestone-like structure. These differentiated cells evidently expressed EC-specific makers and possessed mature ECs function, while inhibition of paracrine factors suppressed the MSC-EC differentiation. Strikingly, the increased secretion of MSCmiR-126 and their endothelial-differentiated potential were deprived by using a PI3K or MEK chemical inhibitor. Our results suggest that overexpression of miR-126 agumenting the endothelial differentiation of MSCs might in part be attributable to the activation of PI3K/Akt and MAPK/ERK pathways and an increased release of paracrine factors.

miR-1-Mediated Induction of Cardiogenesis in Mesenchymal Stem Cells via Downregulation of Hes-1

MicroRNAs (miRNAs, miRs) have the potential to control stem cells fate decisions. The cardiac- and skeletal-muscle-specific miRNA, miR-1, can regulate embryonic stem cells differentiation to cardiac lineage by suppressing gene expression of alternative lineages. Accordingly, we hypothesized that overexpression of miR-1 may also promote cardiac gene expression in mesenchymal stem cells. Since Notch signaling could inhibit muscle differentiation, a process in contrast with the effect of miR-1, miR-1-mediated repression of Notch signaling may contribute to the observed effects of miR-1 in mesenchymal stem cells. Thus, mesenchymal stem cells were infected by lentiviral vectors carrying miR-1, and cells expressing miR-1 were selected. Alterations in Notch signaling and cardiomyocyte markers, Nkx2.5, GATA-4, cTnT, and CX43, were identified by Western blot in the infected cells on days 1, 7, and 14. Our study showed that the downstream target molecule of Notch pathway, Hes-1, was obviously decreased in mesenchymal stem cells modified with miR-1, and overexpression of miR-1 promotes the specific cardiac gene expression in the infected cells. Knockdown of Hes-1 leads to the same effects on cell lineage decisions. Our results indicated that miR-1 promotes the differentiation of MSCs into cardiac lineage in part due to negative regulation of Hes-1.

Effect of Delayed vs Immediate Stent Implantation on Myocardial Perfusion and Cardiac Function in Patients With ST-Segment Elevation Myocardial Infarction Undergoing Primary Percutaneous Intervention With Thrombus Aspiration

BACKGROUND Optimizing microcirculation in STEMI patients with thrombus-containing lesion undergoing percutaneous coronary intervention (PCI) remains challenging. Our objective was to compare the effects on myocardial perfusion and cardiac function of delayed vs immediate stent implantation after thrombus aspiration in STEMI patients undergoing PCI. METHODS Eighty-seven STEMI patients with thrombus-containing lesion undergoing PCI were enrolled. After thrombus aspiration was performed, subjects were divided into 2 groups according to residual thrombus score (TS): immediate stent implantation (ISI) group (n = 47, residual TS < 2; stenting was performed immediately), and delayed stent implantation (DSI) group (n = 40, residual TS ≥ 2; stenting was performed 7 days later). Corrected thrombolysis in myocardial infarction frame count and myocardial blush grade were analyzed immediately after PCI. The wall motion score index was assessed on admission and at 6-month follow-up. RESULTS At the end of the PCI procedure, the corrected thrombolysis in myocardial infarction frame count was significantly shorter and the myocardial blush grade 3 was more frequent in the DSI group than in the ISI group. Compared with the ISI group, the DSI group had a lower incidence of thrombus-related angiographic events, including distal embolization and no reflow. A significantly greater improvement in wall motion score index from baseline to 6-month follow-up was observed in the DSI group compared with the ISI group. CONCLUSIONS In STEMI patients presenting with thrombus containing lesion undergoing PCI, delayed stent implantation after thrombus aspiration leads to better myocardial perfusion and cardiac functional recovery in comparison with immediate stent implantation.

Complications associated with transcatheter closure of perimembranous ventricular septal defects

Objectives: To identify the complications associated with transcatheter closure of perimembranous ventricular septal defects (PmVSD) using the Amplazter PmVSD occluder (AGA Medical, USA). Methods: Between October 2002 and November 2006, transcatheter closure PmVSD was attempted in 210 patients and performed in 206 patients. Those patients were followed‐up for 6–24 months (mean, 10.6 ± 3.9 months) to identify the complications. Results: Device implantation was successfully accomplished in 206 of the 210 patients (98%). Serious complications such as high degree atrioventricular block (AV block), infective endocarditis, and device embolization occurred in eight cases (3.8%). Other complications including mild aortic or tricuspid regurgitation, femoral pseudoaneurysm, and femoral arteriovenous fistula occurred in four cases. Conclusions: Transcathter closure of PmVSD can be performed safely and successfully. But further studies should continue to evaluate the potential complications associated with this procedure.

The roles of autophagy in vascular smooth muscle cells

Autophagy, which is an evolutionarily conserved mechanism and links to several cellular pathways, impacts vascular smooth muscle cells (VSMCs) survival and function. Activation of autophagy by intercellular and/or extracellular stimuli has protective effects on VSMCs against cell death, while on the contrary, overloading autophagy has been recognized as a deleterious process by excessive self-digestion. Alterations in autophagy has been documented in VSMC in response to various stimuli, resulting in modulation of VSMC functions, including proliferation, migration, matrix secretion, contraction/relaxation, and differentiation. Each of these changes in VSMC functions plays a critical role in the development of vascular diseases. Importantly, emerging evidence demonstrates that autophagy deficiency in VSMCs would contribute to atherosclerosis and restenosis, shedding novel light on therapeutic target of the vascular disorders. Herein, this review summarizes the recent progress associated with the roles of autophagy in VSMC and offers the perspectives to several challenges and future directions for further studies.

Atrioventricular block: a serious complication in and after transcatheter closure of perimembranous ventricular septal defects.

Transcatheter closure is an effective approach for perimembranous ventricular septal defects (PMVSD). However, atrioventricular blocks (AVB) emerged possibly due to the close proximity of the PMVSD to the conduction system, but concern for the complication was not adequately emphasized. In this study, we report the incidence of AVBs, in and after transcatheter closure of a PMVSD, and the outcome of the complication in our center.

Apelin/APJ signaling promotes hypoxia‑induced proliferation of endothelial progenitor cells via phosphoinositide‑3 kinase/Akt signaling

Endothelial progenitor cells (EPCs) can adhere to the endothelium at sites of hypoxia/ischemia and participate in the formation of novel vessels through differentiating into endothelial cells (ECs). Apelin is an endogenous ligand for the G protein‑coupled receptor APJ, and apelin/APJ signaling has a role in cardiovascular function. The present study aimed to investigate the role of apelin/APJ signaling in the regulation of EPC proliferation under hypoxia. The results showed that hypoxia was able to induce EPC proliferation, accompanied with an upregulation of hypoxia‑inducible factor (HIF)‑1α as well as apelin/APJ signaling. Further investigation indicated that siRNA‑mediated knockdown of apelin or APJ expression attenuated the hypoxia‑induced proliferation of EPCs, suggesting that apelin/APJ signaling has an important role in hypoxia‑induced EPC proliferation. Moreover, the phosphoinositide‑3 kinase (PI3K)/Akt signaling pathway was found to be involved in the apelin/APJ‑mediated EPC proliferation under hypoxia. Based on these findings, the present study suggested that hypoxia‑induced upregulation of HIF‑1α promotes the expression of apelin and APJ, which further activate the downstream PI3K/Akt signaling pathway, a key promoter of EPC proliferation. In conclusion, the present study highlighted the role of apelin/APJ in the regulation of EPC proliferation, and apelin/APJ may therefore serve as a potential target for the prevention of hypoxic ischemic injury.

Hypoxia induces the proliferation of endothelial progenitor cells via upregulation of Apelin/APLNR/MAPK signaling

Endothelial progenitor cells (EPCs) can form new vessels through differentiation into endothelial cells (ECs), thus being important in the prevention of hypoxia/ischemia. Apelin can activate different signaling pathways through its receptor, APLNR, which regulate diverse biological functions, including cardiovascular function. However, the molecular mechanism by which Apelin mediates hypoxia-induced EPCs proliferation remain to be fully elucidated. The present study aimed to determine the role of Apelin/APLNR signaling in hypoxia-induced proliferation of EPCs. MTT assay was used to determine cell proliferation. Reverse transcription-quantitative polymerase chain reaction and western blotting analysis were conducted to examine mRNA and protein expression. It was revealed that hypoxia promoted the proliferation of the EPCs. Further investigation demonstrated that hypoxia promoted the expression levels of hypoxia-inducible factor (HIF)-1α, Apelin and APLNR in the EPCs. In addition, upregulation of Apelin or APLNR promoted the hypoxia-induced proliferation of the EPCs, while knockdown of Apelin or APLNR by small interfering RNA suppressed the hypoxia-induced proliferation of the EPCs, suggesting that the Apelin/APLNR axis is involved in hypoxia-induced proliferation of EPCs. Furthermore, pretreatment of the EPCs with SB-239063 or PD98059, two inhibitors of mitogen-activated protein kinase (MAPK), eliminated the Apelin upregulation-induced EPC proliferation, suggesting that MAPK signaling is a downstream effecter of Apelin/APLNR in EPCs. Therefore, the findings of the present study indicated that the production of HIF-1α, induced by hypoxia, activated the Apelin/APLNR and the downstream MAPK signaling pathways, leading to upregulated proliferation of the EPCs. These findings suggested that Apelin/APLNR signaling may be used as a potential therapeutic target for hypoxic/ischemic injury.