PDGF通路在内皮细胞对小鼠AoM来源的MSC-第三军医大学学报.DOC

内皮细胞条件培养基对小鼠AoM来源的MSC迁移的实验研究 阎新龙1，兰雨2，王晓燕1，贺文艳3，熊加祥4，要晖宇1，李韧5，刘兵3*，毛宁1*（军事医学科学院：1基础医学研究所细胞生物学研究室，北京 100850，2生物工程研究所疾病与遗传发育研究室，北京100072，3附属307医院肿瘤分子研究室，北京100071；4第三军医大学生理教研室，重庆 400038；5武警总医院肿瘤中心，北京 100039） 摘 要：目的 在小鼠AoM区分离MSC,探讨内皮细胞对MSC的趋化作用。方法 取小鼠E11.5 AoM区，胶原酶消化后，贴壁培养获得MSC，流式细胞仪及免疫荧光表型鉴定，诱导分化功能鉴定MSC，收集内皮条件培养基，Transwell方法测定AoM迁移能力。结果 P2-P3代的AoM-MSC不表达造血和内皮的标记，表达周细胞的标记（CD140b,CD140a, a-SMA，NG2），具有体外分化为脂肪，成骨，软骨的能力。比较了11种生长因子对AoM-MSC迁移的影响，其中bFGF、PDGF-BB、TGF-β1对MSC显著促进迁移。内皮条件培养基能够趋化AoM-MSC迁移，该作用可被PDGF受体抑制剂抑制，而TGF-β受体抑制剂和bFGF受体抑制剂则无抑制效应；可被JNK和AKT通路抑制剂抑制，而ERK和P38通路抑制剂不起作用。结论 小鼠AoM存在三系分化的MSC，PDGF在内皮条件培养基对AoM-MSC的迁移中起重要作用，且JNK和AKT通路抑制剂抑制内皮对AoM-MSC的迁移，而ERK和P38通路则无抑制作用。 关键词：AGM区；间充质干细胞；分化；内皮条件培养基；迁移；JNK通路 Endothelial cell conditional medium induced migration of AoM derived mesenchymal stem cells Xin-Long Yan1, Yu Lan2, Xiao-Yan Wang1, Wen-Yan He 3, Jia-Xiang Xiong4, Hui-Yu Yao1, Bing Liu3, Ning Mao1（1Department of Cell Biology, Institute of Basic Medical Sciences, Beijing 1008502Genetic Laboratory of Development and Diseases, Institute of Biotechnology, Beijing 100071; 3Laboratory of Oncology, Affiliated Hospital of Academy of Military Medical Sciences, Beijing 100071; 4Department of Physiology, Third Military Medical University, Chongqing 400038, 5Cancer Center,General hospital of Chinese people’s armed police forces,beijing,100039,China） Abstract：Objecve To explore the possibility of isolating mesenchymal stem cells from mouse aorta and mesenchyme (AoM) and study the chemotactic effect of endothelial cell conditional medium(ECM) on the AoM-MSCs. Methods E11.5 mouse AoM was longitudinally dissected from the urogenital ridges and digested with collagenase , MSC adhered to plastic and amplified remarkably. The phenotypes of MSC were identified by flow cytometry analysis and immunofluorescence staining. The transwell was applied in chemotactic study. Results The P2-P3 AoM-MSC did not express hematopoietic and endothelial markers and expressed pericyte markers (CD140b, CD140a,