A Tumor Necrosis Factor-α and Hypoxia-Induced Secretome Therapy for Myocardial Repair.

BACKGROUND: Poor viability and retention of transplanted bone marrow mesenchymal stem cells (BM-MSC) remains an obstacle in promoting healing after myocardial infarction (MI). This study aimed to understand the migratory, angiogenic, and cardioprotective effects induced by tumor necrosis factor (TNF)-α and hypoxia through rat BM-MSC (rBM-MSC) paracrine secretions, collectively referred to as secretome, after MI. METHODS: Secretome from rBM-MSC cultures treated with various combinations of H9c2 cardiomyoblast-conditioned medium, TNF-α, and hypoxia was initially collected. Immunocytochemistry, Western blot analyses, and transwell cell migration assays were conducted. In vivo, echocardiography was performed on rats with induced MI after their treatment with TNF-α and hypoxia-induced secretome. RESULTS: Immunocytochemistry confirmed the presence of TNF receptors 1 and 2 on rBM-MSCs. Western blot analyses of rBM-MSCs treated with TNF-α and hypoxia showed an overall increasing trend in the expression of antiinflammatory proteins and angiogenic and migratory cytokines (transforming growth factor-ß, fibroblast growth factor-2, angiopoietin-2, vascular endothelial growth factor-1). In addition, the TNF-α and hypoxia-induced secretome significantly increased the in vitro rBM-MSCs migration. In the rat MI model, the rats treated with the TNF-α and hypoxia-induced secretome had a significantly higher left ventricular fractional shortening than the control group. CONCLUSIONS: Our data suggest that after MI, rBM-MSCs secrete paracrine factors in response to TNF-α and hypoxia that work together to manipulate the microenvironment and decrease inflammation. In addition, these signaling factors trigger angiogenic and migratory effects at the site of the infarct to promote myocardial healing and improve the cardiac function.

Hypoxia modulates cell migration and proliferation in placenta-derived mesenchymal stem cells.

OBJECTIVES: For more than a decade, stem cells isolated from different tissues have been evaluated in cell therapy. Among them, the human bone marrow-derived mesenchymal stem cells (hBM-MSCs) were investigated extensively in the treatment of myocardial infarction. Recently, the human placenta-derived mesenchymal stem cells (hPD-MSCs), which are readily available from a biological waste, appear to be a viable alternative to hBM-MSCs. METHODS: C-X-C chemokine receptor type 4 (CXCR4) gene expression and localization were detected and validated in hPD-MSCs and hBM-MSCs via polymerase chain reaction and immunofluorescence. Subsequently, cell culture conditions for CXCR4 expression were optimized in stromal-derived factor-1 alpha (SDF1-α), glucose, and cobalt chloride (CoCl2) by the use of cell viability, proliferation, and migration assays. To elucidate the cell signaling pathway, protein expression of CXCR4, hypoxia-inducible factor-1α, interleukin-6, Akt, and extracellular signal-regulated kinase were analyzed by Western blot. CXCR4-positive cells were sorted and analyzed by florescence-activated cell sorting. RESULTS: CXCR4 was expressed on both hPD-MSCs and hBM-MSCs at the basal level. HPD-MSCs were shown to have a greater sensitivity to SDF-1α-dependent cell migration compared with hBM-MSCs. In addition, CXCR4 expression was significantly greater in both hPD-MSCs and hBM-MSCs with SDF-1α or CoCl2-induced hypoxia treatment. However, CXCR4+ hPD-MSCs population increased by 10-fold in CoCl2-induced hypoxia. In contrast, only a 2-fold increase was observed in the CXCR4+ hBM-MSCs population in similar conditions. After CoCl2-induced hypoxia, the CXCR4/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase signaling pathway was activated prominently in hPD-MSCs, whereas in hBM-MSCs, the CXCR4/phosphatidylinositol 3-kinase/Akt pathway was triggered. CONCLUSIONS: Our current results suggest that hPD-MSCs could represent a viable and effective alternative to hBM-MSCs for translational studies in cardiocellular repair.

Conditioned medium of H9c2 triggers VEGF dependent angiogenesis by activation of p38/pSTAT3 pathways in placenta derived stem cells for cardiac repair.

AIMS: Cardiomyocytes are understood to possess a limited regenerative capacity. Any myocardial insult leads to an irreversible injury. Mesenchymal stem cell differentiation into cardiomyocyte-like cells stands as one of the leading experimental therapies. However, a candidate cell source has yet to be defined. Here, we examined the in vitro and in vivo cardiac differentiation potential of human placenta derived stem cells (hPDSCs); a unique, abundant, and non-immunogenic cell source. MAIN METHODS: H9c2 cell culture medium was applied to hPDSCs at different ratios for a period of 4weeks. In parallel, hPDSCs, human bone marrow stem cells, or cell free culture medium was injected in peri-infarcted regions induced in rat hearts. KEY FINDINGS: In vitro, hPDSCs pre-conditioned with H9c2 cell culture medium proportionally over-expressed alpha sarcoplasmic actinin and displaced connexin 43 from the cytoplasm to the cell membrane. Additionally, pre-conditioning promoted hPDSCs survival and triggered vascular endothelial growth factor (VEGF) dependent angiogenesis by activating the pAkt and p38MAPK/pSTAT3 pathways. In vivo, echocardiography analysis showed a significant improvement in cardiac parameters in the rats injected with hPDSCs, similar to the human bone marrow stem cells injected group. Moreover, hPDSCs detected within rat cardiac tissues expressed troponin I and myosin heavy chain. In accordance with the pre-conditioning findings, VEGF positive neovessels were observed in hearts injected with hPDSCs. SIGNIFICANCE: hPDSCs have the potential to differentiate into cardiac-like cells and induce angiogenesis via paracrine effects. With the advantages of easy availability and young age, these cells could be more suitable for clinical translation.