The Role And Mechanism Of Bone Marrow Mesenchymal Stem Cells In Tumor Progression And Tumor Microenvironment

Objective:Mesenchymal stem cells (MSCs) were injected as cell mixture with tumor cells or were injected separately at different anatomic sites from the site of tumor injection can significantly enhance the growth of tumor xenograft in BALB/C nude mouse model, but the exact mechanism remains unclear. The aim of this study is to investigate the effect of MSCs on tumor microenvironment and the mechanism of MSCs on tumor initiation and progression. It will provide the new evidence that MSCs exert the tumor promoting function and provide a new target of tumor therapy based on MSC.Methods:We treated SGC-7901 gastric cancer line cells with hMSC-conditioned medium (hMSC-CM) or hMSC derived exosomes for 48 hours, followed by subcutaneously injection of the treated SGC-7901 cells (1x106) into BALB/c nude mice for the development of tumor xenograft. We also injected untreated SGC-7901 cells alone or co-injected with hMSC as control group. We determine in vitro proliferation index of SGC-7901 tumor cells before and after hMSC-CM treatment in culture using MTT assay. We set out to quantitative measure the presence of blood vessels by using immunostaining of vWF and Magnetic Resonance (MR) angiography was performed for analyzing tumor vascularity. We detected the effect hMSC secreted molecules on the expression of VEGF at both mRNA and protein levels in SGC-7901 cells by RT-PCR and ELISA assay, and to test neovascularization functionally in vivo by chick embryo chorioallantoic membrane (CAM) angiogenesis observation. The activation of ERK1/2 or RhoA-GTP were performed for gene expression profile analysis on hMSC-CM treated and untreated cells by gene chip and western bolt assay.We established nude mouse orthotopic transplantation models of human stomach cancer constructed using intact tumor tissue by "Fibrinogen-thrombin" paste technique. And gastric cancer-derived MSC-like cells (hGC-MSCs) were isolated from gastric cancer tissue samples. We treated hMSCs with SGC-7901 cells or gastric cancer tissue in vitro to observe whether hMSCs differentiate into tumor associated fibroblast (TAF).Results:Our results suggest that physical and continuous presence or interaction of MSCs with tumor cell is not required for enhanced tumor growth. Instead, one dose of pretreatment of tumor cells with hMSC conditioned-media is sufficient to recapitulate hMSC co-injection and the hMSCs secreted soluble factors are the major effort if not all tumor growth-promoting potential.The tumor tissue section exhibited a much more robust neovasculization in the xenograft derived from hMSC-CM pretreated tumor cells contrasting to that of untreated control tumor graft. Our data clearly demonstrated that signaling molecules secreted by hMSCs strongly activate VEGF expression of tumor cells at both mRNA and protein levels and the drastic ongoing activation pathways in these tumor cells upon the exposure to hMSC-CM leading to profound alteration in gene expression profiles of the tumor cells. After treatment with hMSCs-CM, the gene expression of SGC-7901 cell for N-cadherin,slug,snail was increased,but the expression of E-cadherin was decreased. The tumor tissue expressed stronger a-SMA in vivo. Our results showed that hMSCs-CM can induced epithelial-mesenchymal transition(EMT) for tumor cell. MSC-exosomes co-injected with tumor cell or pretreatment in vitro can promoted tumor growth, increased the expression of a-SMA,VEGF,CXCR4,PCNA and blood vessels density in tumor tissue. However, the MSC-exosomes did not induce any significant increase in tumor cell proliferation in vitro but a transient activation of ERK and enhanced expression of VEGF and CXCR4 genes were observed. The gastric cancer-derived MSC-like cells (hGC-MSCs) were isolated from 13 out of 20 cancer tissue samples. Their characteristics, including morphology, surface antigens, specific gene expression, and differentiation potential, were similar to those of MSCs derived from human bone marrow (hBM-MSCs). The nude mouse orthotopic transplantation model of human stomach cancer was constructed and 100% successful tumor-take rate was obtained in the model and we have succeed isolated MSCs from the nude mouse orthotopic transplantation model tissue.After treatment with tumor cell or tumor tissue in vitro, hBM-MSCs may be differentiate into TAF, also they can expression the special mark of TAFs, such as a-SMA,vimetin. Our findings suggest that MSCs are components of the tumor microenvironment, which can promote tumor growth.Conclusions:In this study, we show that through the action of secreted VEGF, MSCs potentiate tumor cells by activating angiogenic program as well as by altering the expression profile of a broad range of genes. These cells exhibit the ability to induced EMT or neovascularization for tumor. It also can promote tumor cell growth and the ability of migration.To our knowledge, it's the first report to present that MSC-exosomes had a significant acceleration effect on tumor growth in vivo, which offers a new mechanism responsible for the roles of MSCs in promoting tumor growth in vivo and the exosomes-meidated cell-cell interaction may be a new tumor therapeutic target. The existence of MSC-like cells in gastric cancer tissues suggests that MSC is the portion of tumor microenvironment, and the MSCs may be differentiate into TAF under tumor microenvironment in vitro. Taken together, these data suggest that hMSCs are a source of TAFs. This provides an experimental foundation for investigating the role of hMSCs in the initiation and progression of cancers and it may be a potential targets for cancer therapy.