Poor long-term patency of vein grafts remains an obstacle in coronary artery bypass grafting (CABG) surgery using an autologous saphenous vein graft. Recent studies have revealed that miR-126-3p promotes vascular integrity and angiogenesis. We aimed to identify the role of miR-126-3p in the setting of vein graft disease and investigate the value of miR-126-3p agomir as a future gene therapy in vein graft failure. Expression analysis of circulating miR-126-3p in plasma from CABG patients established the basic clues that miR-126-3p participates in CABG. The in vitro results indicated that elevated miR-126-3p expression significantly improved proliferation and migration in human saphenous vein endothelial cells (HSVECs) by targeting sprouty-related protein-1 (SPRED-1) and phosphatidylinositol-3-kinase regulatory subunit 2 (PIK3R2), but not in human saphenous vein smooth muscle cells (HSVSMCs). Moreover, the therapeutic potential of miR-126-3p agomir was demonstrated in cultured human saphenous vein (HSV) ex vivo. Finally, local delivery of miR-126-3p agomir was confirmed to enhance reendothelialization and attenuate neointimal formation in a rat vein arterialization model. In conclusion, we provide evidence that upregulation of miR-126-3p by agomir possesses potential clinical value in the prevention and treatment of autologous vein graft restenosis in CABG.

Background In our previous research, we found that overexpression of miR-126-3p in human umbilical cord MSCs (hucMSCs) promoted human umbilical vein endothelial cells angiogenic activities through exosome-mediated mechanisms. The present study aimed to investigate the role of miR-126-3p-modified hucMSCs derived exosomes (miR-126-3p-hucMSCs-exosomes) on the treatment of premature ovarian failure (POF). Methods Primary hucMSCs were isolated from human umbilical cords and identified by differentiation experiments and flow cytometry. miR-126-3p-hucMSCs were obtained by miR-126-3p lentivirus infection. miR-126-3p-hucMSCs-exosomes were purified by ultracentrifugation method and characterized by transmission electron microscopy and western blot analysis. Primary rat ovarian granulosa cells (OGCs) were collected from ovarian tissues and identified by cell immunohistochemistry. The effects of miR-126-3p-hucMSCs-exosomes and miR-126-3p on OGCs function were determined by cell proliferation and apoptosis assays in a cisplatin induced POF cell model. The levels of suitable target genes were analyzed by PCR and Western blot analysis and subsequent Dual-Luciferase reporter assay. The signal pathway was also analyzed by western blot analysis. A cisplatin-induced POF rat model was used to validate the therapeutic effects of miR-126-3p-hucMSCs-exosomes to treat POF. Ovarian function was evaluated by physical, enzyme-linked immunosorbent assay, and histological examinations in chemotherapy-treated rats. The angiogenesis and apoptosis of ovarian tissues were assessed by immunohistochemical staining and Western blots. Results Primary hucMSCs and miR-126-3p-hucMSCs-exosomes and primary rat OGCs were successfully isolated and identified. The cellular uptake experiments indicated that miR-126-3p-hucMSC-exosomes can be internalized into OGCs in vitro. Annexin V-FITC/PI staining and EDU assays revealed that both miR-126-3p-hucMSCs-exosomes and miR-126-3p promoted proliferation and inhibited apoptosis of OGCs damaged by cisplatin. PCR and western blot analysis and subsequent dual-luciferase reporter assay verified that miR-126-3p targets the sequence in the 3' untranslated region of PIK3R2 in OGCs. Further analysis showed that PI3K/AKT/mTOR signaling pathway took part in miR-126-3p/PIK3R2 mediated proliferation and apoptosis in OGCs. In rat POF model, administration of miR-126-3p-hucMSCs-exosomes increased E2 and AMH levels, increased body and reproductive organ weights and follicle counts, and reduced FSH levels. But more importantly, immunohistochemistry results indicated miR-126-3p-hucMSCs-exosomes significantly promoted ovarian angiogenesis and inhabited apoptosis in POF rats. Additionally, the analysis of angiogenic-related factors and apoptosis-related factors showed miR-126-3p-hucMSCs-exosomes had pro-angiogenesis and anti-apoptosis effect in rat ovaries. Conclusions Our findings revealed that hucMSCs-derived exosomes carrying miR-126-3p promote angiogenesis and attenuate OGCs apoptosis in POF, which highlighted the potential of exosomes containing miR-126-3p as an effective therapeutic strategy for POF treatment.

Bone marrow‐derived mesenchymal stem cells (BMSCs) have great therapeutic potential for many diseases. However, the homing of BMSCs to injury sites remains a difficult problem. Recent evidence indicates that simvastatin stimulates AKT phosphorylation, and p‐AKT affects the expression of chemokine (CXC motif) receptor‐4 (CXCR4). Therefore, simvastatin may improve the expression of CXCR4 in BMSCs, and microRNAs (miRs) may participate in this process. In this study, we demonstrated that simvastatin increased both the total and the surface expression of CXCR4 in BMSCs. Stromal cell‐derived factor‐1α (SDF‑1α)‐induced migration of BMSCs was also enhanced by simvastatin, and this action was inhibited by AMD 3100(a chemokine receptor antagonist for CXCR4). The PI3K/AKT pathway was activated by simvastatin in this process, and LY294002 reversed the overexpression of CXCR4 caused by simvastatin. MiR‐9 directly targeted CXCR4 in rat BMSCs, and simvastatin decreased miR‐9 expression. P‐AKT affected the expression of miR‐9; as the phosphorylation of AKT increased, miR‐9 expression decreased. In addition, LY294002 increased miR‐9 expression. Taken together, our results indicated that simvastatin improved the migration of BMSCs via the PI3K/AKT pathway. MiR‐9 also participated in this process, and the phosphorylation of AKT affected miR‐9 expression, suggesting that simvastatin might have beneficial effects in stem cell therapy.

The oxidative stress caused by endothelial injury is involved in intimal hyperplasia (IH) in vein grafts. Mesenchymal stem cells (MSCs) can home to injured intima and promote endothelial repair. However, MSC apoptosis is increased accompanied by decreased functional activity under oxidative stress. Thus, we investigate whether tumour necrosis factor‐α (TNF‐α) can promote the survival and activity of MSCs under oxidative stress to reduce IH more effectively, and establish what role the NF‐κB pathway plays in this. In this study, we preconditioned MSCs with TNF‐α (TNF ‐α‐PCMSCs) for 24 hrs and measured the activation of the IKK/NF‐κB pathway. EdU and transwell assays were performed to assess proliferation and migration of TNF ‐α‐PCMSCs. Apoptosis and migration of TNF ‐α‐ PCMSCs were evaluated in conditions of oxidative stress by analysis of the expression of Bcl‐2 and CXCR4 proteins. TNF ‐α‐ PCMSCs were transplanted into a vein graft model, so that cell homing could be tracked, and endothelial apoptosis and IH of vein grafts were measured. The results demonstrated that TNF‐α promotes proliferation and migration of MSCs. Furthermore, survival and migration of TNF ‐α‐ PCMSCs under oxidative stress were both enhanced. A greater number of MSCs migrated to the intima of vein grafts after preconditioning with TNF‐α, and the formation of neointima was significantly reduced. These effects could be partially abolished by IKK XII (NF‐κB inhibitor). All these results indicate that preconditioning with TNF‐α can promote survival and migration of MSCs under oxidative stress via the NF‐κB pathway and thus attenuate IH of vein grafts.

Background The aim of this study was to determine whether the combination of MSC implantation with miRNA-126-3p overexpression would further improve the surgical results after vein grafting. Methods human umbilical cord MSCs (hucMSCs) and human umbilical vein endothelial cells (HUVECs) were isolated from human umbilical cords and characterized by a series of experiments. Lentivirus vector encoding miRNA-126-3p was transfected into hucMSCs and verified by PCR. We analyzed the miRNA-126-3p-hucMSC function in vascular endothelial cells by using a series of co-culture experiments. miRNA-126-3p-hucMSCs-exosomes were separated from cell culture supernatants and identified by WB and TEM. We validated the role of miRNA-126-3p-hucMSCs-exosomes on HUVECs proliferative and migratory and angiogenic activities by using a series of function experiments. We further performed co-culture experiments to detect downstream target genes and signaling pathways of miRNA-126-3p-hucMSCs in HUVECs. We established a rat vein grafting model, CM-Dil-labeled hucMSCs were injected intravenously into rats, and the transplanted cells homing to the vein grafts were detected by fluorescent microscopy. We performed historical and immunohistochemical experiments to exam miRNA-126-3p-hucMSC transplantation on vein graft neointimal formation and reendothelialization in vitro. Results We successfully isolated and identified primary hucMSCs and HUVECs. Primary hucMSCs were transfected with lentiviral vectors carrying miRNA-126-3p at a MOI 75. Co-culture studies indicated that overexpression of miRNA-126-3p in hucMSCs enhanced HUVECs proliferation, migration, and tube formation in vivo. We successfully separated hucMSCs-exosomes and found that miRNA-126-3p-hucMSCs-exosomes can strengthen the proliferative, migratory, and tube formation capacities of HUVECs. Further PCR and WB analysis indicated that, SPRED-1/PIK3R2/AKT/ERK1/2 pathways are involved in this process. In the rat vein arterialization model, reendothelialization analysis showed that transplantation with hucMSCs modified with miRNA-126-3p had a higher reendothelialization of the vein grafts. The subsequent historical and immunohistochemical examination revealed that delivery with miRNA-126-3p overexpressed hucMSCs significantly reduced vein graft intimal hyperplasia in rats. Conclusion These results suggest hucMSC-based miRNA-126-3p gene therapy may be a novel option for the treatment of vein graft disease after CABG.