胃癌MSC源性miRNAs再编程骨髓MSC作用及机制研究

Bone marrow mesenchymal stem cell (BM-MSC) is an important source of tumor stroma cells. Our research group firstly isolated and got a kind of important stroma cell from gastric cancer tissue, which was named as gastric cancer tissue-derived MSC (GC-MSC), and also got the corresponding gastric non-cancerous tissue-derived MSC(GCN-MSC). When BM-MSC was induced by gastric cancer cells, its phenotype was similar to that of GC-MSC. However,how BM-MSC transdifferentiate into GC-MSC and its molecular mechanism remain unclear. On the basis of the miRNAs differently expressed between GC-MSC and GCN-MSC by microarray screening, we further validate and determine the differential miRNAs for GC-MSC. We set gastric cancer cell-induced BM-MSC and GC-MSC as cell models, take the differential expression of miRNAs as the breakthrough point and combine BM-MSC transdifferentiation with GC-MSC dedifferentiation as the bidirectional model to determine the key miRNAs that are participated in reprogramming BM-MSC to GC-MSC. Then we perform screening, validation of the target genes for miRNAs and the target gene expression interferencing to elucidate the molecular mechanism of miRNAs reprogramming BM-MSC. This study is aiming at explaining the molecular mechanism of reprogramming BM-MSC to GC-MSC, providing a new proof for the source and transdifferentiation mechanism of gastric cancer stroma cell and supply new therapeutic targets for gastric cancer, which shows a great significance and potential application in the future.

骨髓MSC是肿瘤微环境细胞的重要来源。本课题组率先从胃癌组织分离培养获得一种重要的微环境细胞即胃癌MSC以及相应癌旁MSC，发现胃癌细胞诱导后的骨髓MSC表型与胃癌MSC相似。但骨髓MSC如何向胃癌MSC转分化及其分子机制尚不清楚。本研究在前期筛选出胃癌MSC与癌旁MSC差异表达miRNAs的工作基础之上，进一步验证并确定胃癌MSC差异表达的miRNAs。以胃癌细胞诱导后的骨髓MSC和胃癌MSC为研究的细胞模型，以差异表达的miRNAs为切入点，结合骨髓MSC转分化与胃癌MSC逆分化双向研究模式，确定参与再编程骨髓MSC向胃癌MSC转分化的关键miRNAs；通过靶基因的筛选、验证及干预阐明miRNAs再编程的分子机制。本研究旨在解析再编程骨髓MSC向胃癌MSC转分化的分子调控机制，为胃癌微环境细胞来源与转分化机制提供新的实验依据，为胃癌提供新的治疗靶点，具有重要的意义和潜在的应用价值。

骨髓间充质干细胞MSC是肿瘤微环境细胞的重要来源，胃癌MSC是本课题组率先从胃癌组织中分离培养获得的一种重要的微环境细胞，其较骨髓MSC具有更强的促癌表型和功能，但骨髓MSC如何向胃癌MSC转分化及其分子机制尚不清楚。前期芯片筛选及定量PCR检测验证发现胃癌MSC与癌旁MSC存在着差异表达的miRNAs，结合体外HGC-27和SGC-7901胃癌细胞上清诱导骨髓MSC向胃癌MSC转分化细胞模型检测，确定诱导后骨髓MSC与胃癌MSC差异变化一致的miRNAs包括miR-155-5p、miR-99a-5p和miR-4669。模拟胃癌MSC中miRNAs表达模式，分别抑制骨髓MSC中miR-155-5p、miR-99a-5p水平或过表达miR-4669可促进其获得胃癌MSC样表型和功能。反之，分别过表达miR-155-5p、miR-99a-5p或低表达miR-4669可逆转胃癌MSC促癌表型和功能。靶基因预测、筛选、验证及功能试验确定miR-155-5p通过靶向作用IKBKE和NF-κB p65调控骨髓MSC转分化；miR-99a-5p靶向调控FGFR3参与骨髓MSC再编程； miR-4669 靶向抑制TIMP3和NKIRAS2发挥促骨髓MSC转分化作用。靶向干预胃癌MSC中miRNAs调控靶基因可消除其促癌表型和功能。本项目严格按照研究计划顺利完成，已发表基金标注SCI论文4篇，核心期刊论文3篇，已培养博士研究生1名，硕士研究生3名。本项目以胃癌MSC差异表达的miRNAs为研究切入点，确定胃癌MSC差异表达的miR-155-5p、miR-99a-5p和miR-4669是直接再编程骨髓MSC转分化的关键miRNAs，阐明了其调控骨髓MSC向胃癌MSC样细胞转分化分子机制，为胃癌MSC来源与如何转分化形成提供了新的实验依据，有望为胃癌治疗提供新的有效分子靶标，具有重要的研究意义和临床应用前景。