Enhancing insulin sensitivity in type 2 diabetes mellitus using apelin-loaded small extracellular vesicles from Wharton's jelly-derived mesenchymal stem cells: a novel therapeutic approach.

BACKGROUND: Type 2 diabetes mellitus (T2DM), characterized by ß-cell dysfunction and insulin resistance (IR), presents considerable treatment challenges. Apelin is an adipocyte-derived factor that shows promise in improving IR; however, it is limited by poor targeting and a short half-life. In the present study, engineered small extracellular vesicles (sEVs) derived from Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) loaded with apelin were used to address the limitations of the therapeutic application of apelin. METHODS: WJ-MSCs were transduced to obtain engineered sEVs loaded with overexpressed apelin (apelin-MSC-sEVs) and the control sEVs (MSC-sEVs). T2DM mice were injected with apelin-MSC-sEVs and MSC-sEVs, and blood glucose monitoring, glucose and insulin tolerance tests, confocal microscopy, and immunocytochemical analysis were performed. IR models of 3T3-L1 adipocytes were employed to detect GLUT4 expression in each group using western blotting; the affected pathways were determined by measuring the changes in Akt and AMPK signaling and phosphorylation. RESULTS: Upon successful engineering, WJ-MSCs demonstrated significant overexpression of apelin. The genetic modification did not adversely impact the characteristics of sEVs, ranging from surface protein markers, morphology, to particle size, but generated apelin-overexpressed sEVs. Apelin-MSC-sEVs treatment resulted in notable enhancement of Akt and AMPK pathway activities within 3T3-L1 adipocytes and adipose tissues of T2DM mice. Furthermore, the apelin-loaded sEVs significantly reduced plasma glucose levels, increased pancreatic ß-cell proliferation, improved insulin and glucose tolerance, and modulated pro-inflammatory cytokine profiles, compared to mice treated with the control sEVs. CONCLUSION: Our study developed novel genetically engineered apelin-loaded sEVs derived from WJ-MSCs, and demonstrated their potent role in augmenting insulin sensitivity and regulating inflammatory responses, highlighting their therapeutic promise in T2DM management. The findings open new avenues for the development of clinically viable treatments for T2DM in humans using the apelin-loaded sEVs.

Bone marrow mesenchymal stem cells transfer in patients with ST-segment elevation myocardial infarction: single-blind, multicenter, randomized controlled trial.

OBJECTIVE: Our aim was to evaluate the efficacy and safety of intracoronary autologous bone marrow mesenchymal stem cell (BM-MSC) transplantation in patients with ST-segment elevation myocardial infarction (STEMI). METHODS: In this randomized, single-blind, controlled trial, patients with STEMI (aged 39-76 years) were enrolled at 6 centers in Beijing (The People's Liberation Army Navy General Hospital, Beijing Armed Police General Hospital, Chinese People's Liberation Army General Hospital, Beijing Huaxin Hospital, Beijing Tongren Hospital, Beijing Chaoyang Hospital West Hospital). All patients underwent optimum medical treatment and percutaneous coronary intervention and were randomly assigned in a 1:1 ratio to BM-MSC group or control group. The primary endpoint was the change of myocardial viability at the 6th month's follow-up and left ventricular (LV) function at the 12th month's follow-up. The secondary endpoints were the incidence of cardiovascular event, total mortality, and adverse event during the 12 months' follow-up. The myocardial viability assessed by single-photon emission computed tomography (SPECT). The left ventricular ejection fraction (LVEF) was used to assess LV function. All patients underwent dynamic ECG and laboratory evaluations. This trial is registered with ClinicalTrails.gov, number NCT04421274. RESULTS: Between March 2008 and July 2010, 43 patients who had underwent optimum medical treatment and successful percutaneous coronary intervention were randomly assigned to BM-MSC group (n = 21) or control group (n = 22) and followed-up for 12 months. At the 6th month's follow-up, there was no significant improvement in myocardial activity in the BM-MSC group before and after transplantation. Meanwhile, there was no statistically significant difference between the two groups in the change of myocardial perfusion defect index (p = 0.37) and myocardial metabolic defect index (p = 0.90). The LVEF increased from baseline to 12 months in the BM-MSC group and control group (mean baseline-adjusted BM-MSC treatment differences in LVEF 4.8% (SD 9.0) and mean baseline-adjusted control group treatment differences in LVEF 5.8% (SD 6.04)). However, there was no statistically significant difference between the two groups in the change of the LVEF (p = 0.23). We noticed that during the 12 months' follow-up, except for one death and one coronary microvascular embolism in the BM-MSC group, no other events occurred and alanine transaminase (ALT) and C-reactive protein (CRP) in BM-MSC group were significantly lower than that in the control group. CONCLUSIONS: The present study may have many methodological limitations, and within those limitations, we did not identify that intracoronary transfer of autologous BM-MSCs could largely promote the recovery of LV function and myocardial viability after acute myocardial infarction.

Apelin: an endogenous peptide essential for cardiomyogenic differentiation of mesenchymal stem cells via activating extracellular signal-regulated kinase 1/2 and 5.

Growing evidence has shown that apelin/APJ system functions as a critical mediator of cardiac development as well as cardiovascular function. Here, we investigated the role of apelin in the cardiomyogenic differentiation of mesenchymal stem cells derived from Wharton's jelly of human umbilical cord in vitro. In this research, we used RNA interference methodology and gene transfection technique to regulate the expression of apelin in Wharton's jelly-derived mesenchymal stem cells and induced cells with a effective cardiac differentiation protocol including 5-azacytidine and bFGF. Four weeks after induction, induced cells assumed a stick-like morphology and myotube-like structures except apelin-silenced cells and the control group. The silencing expression of apelin in Wharton's jelly-derived mesenchymal stem cells decreased the expression of several critical cardiac progenitor transcription factors (Mesp1, Mef2c, NKX2.5) and cardiac phenotypes (cardiac α-actin, ß-MHC, cTnT, and connexin-43). Meanwhile, endogenous compensation of apelin contributed to differentiating into cells with characteristics of cardiomyocytes in vitro. Further experiment showed that exogenous apelin peptide rescued the cardiomyogenic differentiation of apelin-silenced mesenchymal stem cells in the early stage (1-4 days) of induction. Remarkably, our experiment indicated that apelin up-regulated cardiac specific genes in Wharton's jelly-derived mesenchymal stem cells via activating extracellular signal-regulated kinase (ERK) 1/2 and 5.

Therapeutic Potential of Umbilical Cord Mesenchymal Stem Cells for Inhibiting Myofibroblastic Differentiation of Irradiated Human Lung Fibroblasts.

Radiation-induced lung injury (RILI) limits the benefits of radiotherapy in patients with lung cancer. Radiation-induced differentiation of lung fibroblasts to myofibroblasts plays a key role in RILI. Recent studies have shown that mesenchymal stem cells (MSCs) can protect against lung fibrosis and that Wnt/ß-catenin signaling is involved in fibrotic processes. In the present study, we explored the therapeutic potential of human umbilical cord MSCs (HUMSCs) for preventing radiation-induced differentiation of human lung fibroblasts (HLFs) to myofibroblasts. There are two advantages in the use of HUMSCs; namely, they are easily obtained and have low immunogenicity. Irradiated HLFs were co-cultured with HUMSCs. Expression of α-smooth muscle actin (α-SMA), a myofibroblast marker, was measured by Western blot analysis and immunohistochemistry. Irradiation (X-rays, 5 Gy) induced the differentiation of HLFs into myofibroblasts, which was inhibited by co-culture with HUMSCs. Irradiation also caused activation of the canonical Wnt/ß-catenin signaling in HLFs, as judged by increased phosphorylation of glycogen synthase kinase 3ß, nuclear accumulation of ß-catenin, and elevated levels of Wnt-inducible signaling protein-1 (WISP-1) in the conditioned medium. However, co-culture with HUMSCs attenuated the radiation-induced activation of the Wnt/ß-catenin signaling. We also measured the expression of FRAT1 that can enhance the Wnt/ß-catenin signaling by stabilizing ß-catenin. Co-culture with HUMSCs decreased FRAT1 protein levels in irradiated nHLFs. Thus, co-culture with HUMSCs attenuated the radiation-induced activation of Wnt/ß-catenin signaling in HLFs, thereby inhibiting myofibroblastic differentiation of HLFs. Wnt/ß-catenin signaling is a potential therapeutic target for limiting RILI in patients receiving radiotherapy for lung cancer.

Human umbilical cord Wharton's jelly stem cells: immune property genes assay and effect of transplantation on the immune cells of heart failure patients.

Stem cells derived from umbilical cord Wharton's jelly (WJSCs) are not immunogenic and have immunosuppressive effects. To evaluate the related mechanisms and the effect of transplantation on body immune cells, we examined immune property genes expression in WJSCs and levels of T-lymphocytes subgroups and immunoglobulins (Ig) in heart failure (HF) patients with and without WJSCs transplantation. WJSCs express immune tolerance genes HLA-E, HLA-G and HLA-F and immunomodulation genes VEGF, TGFß1, HGF, HMOX1, IL1ß, IL-6, LIF, LGALS-1/3/8, COX1/2 and PTGE, while they do not express immune response-related genes HLA-DR, HLA-DQ, HLA-DP, CD80, CD86, CD40 and CD40L. No obvious changes of T-lymphocytes subgroups and plasma IgG/IgM were observed in HF patients with WJSCs transplantation. Our results suggest that the immune properties of WJSCs are due to the expression of immune avoidance and immunomodulation genes in the absence of immune response-related genes. WJSCs are secure in immunological aspects when used as seed cells for cardiac repair.

[Transfection of bone marrow mesenchymal stem cells by adeno-associated virus 2/1 vector].

This study was aimed to investigate the transfection efficacy of recombinant adeno-associated virus 2/1 (rAAV2/1) on bone marrow mesenchymal stem cells (BMMSCs) at different multiplicities of infection (MOI) and time, and effect of transfection on growth of rat BMMSCs. The rat BMMSCs cultured in vitro were transfected by using rAAV2/1 with enhanced green fluorescent protein (rAAV2/1-EGFP) at MOI of 1 x 10(4), 1 x 10(5) and 1 x 10(6); the EGFP expression was observed by fluorescent microscopy at 3, 7 and 14 days. The viability, proliferation multiple, differentiation ability of daughter cells were detected for evaluating the effect of rAAV2/1 on survival, proliferation and differentiation of BMMSCs and the fluorescence index (FI) were determined by flow cytometry. The results indicated that after transfection with rAAV2/1 for 24 hours the green fluorescence in BMMSCs were observed, but also the fluorescence gradually was enhanced along with prolonging of time, and reached to steady level after 7 days; the viability, proliferation multiple, differentiation ability of BMMSCs transfected by rAAV2/1-EGFP at different MOI showed no significant changes at 3,7 and 14 days (p > 0.05), meanwhile at same MOI the proliferation multiple obviously increased in comparison between 7 day vs 3 day and 14 days vs 7 days (p < 0.01). The flow cytometric detection showed that the transfection efficacy of rAAV2/1-EGFP on BMMSCs and FI increased significantly as the multiplicity of infection and culture time increased (p < 0.05). It is concluded that rAAV2/1-EGFP is able to transfect into BMMSCs effectively, but the transfection efficiency and fluorescence index increase significantly along with increase of multiplicity of infection and culture time. rAAV2/1-EGFP do not affect viability, proliferation multiple and differentiation ability of BMMSCs. rAAV2/1 is a kind of active vector for gene transfer to reform BMMSCs.

[Autologous mononuclear bone marrow cell transplantation by intracoronary route for patients with ischemic heart disease: observation of 2-years follow-up].

OBJECTIVE: To investigate the long-term effect and safety of intracoronary autologous bone marrow mononuclear cell (BMMC) transplantation in patients with ischemic heart disease (IHD). METHODS: Seventy-six patients with IHD, 26 patients with acute myocardial infarction (AMI) and 26 patients with chronic ischemic heart failure (CIHF), underwent routine treatment plus intracoronary autologous BMMC transplantation, and 24 patients, including 10 patients with AMI and 14 patients with CIHF underwent routine treatment as controls. Autologous BMMC transplantation was performed via a balloon catheter placed into the infarct-related artery during balloon dilatation by high pressure infusion to occlude the artery, which was performed 6 - 8 times for 2 minutes each with 2-minute interval or via a balloon catheter without occluding the infarct-related artery. Follow-up was conducted for 2 years. RESULTS: The surgery was safety without major periprocedural complications. There were no other new arrhythmias found by Holter recorder during the 2-years follow-up. In the AMI patients receiving BNNC transplantation, the left ventricular ejection fraction (LVEF) 1 and 2 years later increased by 5.79% (P < 0.05), 3.79% (P > 0.05) respectively; but there was no change in left ventricular end diastolic volume (LVEDV) and left ventricular end systolic volume (LVESV). The LVEF 1 and 2 years later of the control group increased by 8.8% and 9.2% respectively (both P < 0.01) and the LVESV 1 and 2 years later decreased by 20.4% and 27.8% respectively (both P < 0.05), the myocardium defect area 2 years later was not significantly different from that 3 months later. The heart function of the control group became markedly worse. CONCLUSION: Autologous BMMC intracoronary transplantation is safe and effective, especially in patients with CIHF.

[Transplantation of fetal cardiomyocyte regenerates infarcted myocardium in rats].

OBJECTIVE: To investigate the feasibility and efficiency of fetal cardiomyocyte transplantation into the rat model of myocardial infarction. METHODS: Cardiomyocytes were isolated from aborted human embryos aged 12 - 16 weeks and cultured for 5 days to confirm their viability. Rat model of extensive myocardial infarction (MI) was established in 18 male Wistar rats by ligating the descending anterior branch of left coronary artery and the 18 rats were randomly divided into 2 groups: transplantation group (n = 7, 2 x 10(6) fetal cardiomyocytes were transplanted into the myocardial scar) and culture medium injection group (n = 6, culture medium was injected into the myocardial scar) 5 days after extensive MI was caused. Another 6 rats undergoing sham operation were used as controls. Echocardiography was performed before and 60 +/- 3 days after the implantation to assess the left ventricular (LV) remodeling and cardiac function. Then the rats were killed and their heart were harvested to undergo HE staining, immunohistochemical examination with antibody against human alpha-actin smooth muscle (SMA) isoform, and light microscopy. RESULTS: Light microscopy revealed the presence of engrafted human fetal cardiomyocytes in the infarcted myocardium and the presence of nascent intercalated disks connecting the engrafted fetal cardiomyocytes and the host myocardium. The engrafted fetal cardiomyocytes were SMA positive. Serial echocardiography revealed that cell transplantation prevented scar thinning, LV further dilatation and dysfunction while the control animals developed scar thinning, significant LV dilatation accompanied by progressive deterioration in LV contractility. CONCLUSION: Fetal cardiomyocytes can be implanted and survive in the infarcted myocardial cells, thus preventing the scar thinning, and LV further dilatation and dysfunction.