胎盘来源的滋养层细胞在血管形成中的应用研究

To construct the optimal culture condition and microenvironment for vessle formation in vitro, which is the main research contents and the bottleneck problems in blood vessel tissue engineering. The human placenta is a highly vascular organ. Substances secreted by placenta trophoblast cells participante in the regulation of placenta blood vessel formation. So trophoblast cells are important control factors in local microenvironment for placenta vessel formation. In previous research, we have obtained mesenchymal stem cells and cytotroblast from placenta of different pregnacy duration, and induced placenta-derived mesenchymal stem cells(hPDMSCs) to differentiate into endothelial cells in vitro, then performed micro tube-like formation with endothelial cells. We will detect the contents of various angiogenic substances secreted by cytotrophoblast, and observe the effect of cytotroblast and/or matirals secreted by them on mesenchymal stem cells proliferation, endothelial cells proliferation and differentiation, micro tube-like formation and in mouse corneal micropocket assay via co-culture in vitro and co-transplantation in vivo with placenta cytotroblast and mesenchymal stem cells, then explore the effects and mechanism of cytotrophoblast in placenta vessle formation. The results will help to explain the role of cytotrophoblast in blood vessel formation, and they will also provide valuable evidence for hPDMSCs' application as seed cells and construction the effective culture condition in vitro with placenta cytotroblast and/or matirals secreted by cytotroblast in vascular tissue engineering.

构建有效的体外培养条件和微环境，是血管组织工程的主要研究内容和关键技术难题。人类胎盘是一个强大的毛细血管网。胎盘滋养层细胞合成的各种物质可以参与胎盘血管生成的调控，因此这些细胞是重要的局部微环境调控因素。我们前期已提取出胎盘间充质干细胞和滋养层细胞，并体外诱导间充质干细胞分化为内皮细胞，形成微管腔。本研究选择不同妊娠时期的胎盘，提取胎盘来源的间充质干细胞和滋养层细胞。检测滋养层细胞分泌的各种促血管生成物质的含量，采用体外共培养及体内共移植等研究方法，观察滋养层细胞及其分泌物质对间充质干细胞、内皮细胞分化增殖、微管腔形成及小鼠角膜血管生成的影响，探讨滋养层细胞对血管形成的作用及其机制。研究结果有助于阐述滋养层细胞对血管形成的作用；为血管组织工程中选用胎盘来源的间充质干细胞作为种子细胞，利用滋养层细胞和（或）其细胞分泌物质构建有效的组织工程血管体外培养环境，提供有价值的理论和实验依据。

胎盘来源的间质干细胞（Human placenta-derived mesenchymal stem cell,hPDMSCs）和滋养层细胞（cytotrophoblast,CT）是胎盘的两大主要细胞成分，这两类细胞及其合成和分泌的物质是胎盘血管生成过程中重要的调控因素。本研究选择不同妊娠时期的胎盘，提取胎盘来源的间充质干细胞。利用胎盘间充质干细胞（hPDMSC）和滋养层细胞系（HTR-8）作为种子细胞，收集不同时期胎盘组织块培养液和hPDMSC、HTR-8细胞培养上清液，采用MTS、Transwell等方法，通过胎盘来源的两种主要细胞培养液以及胎盘组织块培养液与DMEM对比，研究其对HuVEC和hPDMSC细胞的增殖、粘附、迁移、侵袭以及管腔形成的影响，发现均可促进细胞增殖、粘附、迁移和管腔形成，但可抑制细胞的侵袭能力，总体hPDMSC效果优于HTR,24Hours和48Hours培养液均优于6Hours。通过抗体芯片初步筛查发现GRO、IL-8、IL-6、TIMP-1、TIMP-2、MCP-1、ENA-78等因子含量有所不同。为进一步进行机制方面的深入研究打下坚实的基础。