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1、树突状细胞通过NT-3/TrkC信号通路调控神经干细胞存活的体外研究王钟１，陈太邦１，王永飞１，郭乔楠2， 赵建华*１ （１第三军医大学大坪医院野战外科研究所脊柱外科，重庆，400042；2第三军医大学西南医院病理学研究所，重庆，400038）[摘要]目的:研究共培养状态下树突状细胞(dendriticcell,DC)对神经干细胞（neuralstemcell,NSC）存活的影响及其作用机制。方法：体外分离、纯化SD大鼠原代DC和NSC，将二者分隔培养，使用RT-PCR方法检测DC中神经营养素-3（neurotrophin-3,NT-3）的mRNA水平，流式细胞术
2、检测NSC凋亡比例，从而确定DC通过增加NT-3表达而促进NSC存活；同时，免疫荧光、westernblotting技术确定NSC表面NT-3特异性受体TrkC表达，以明确DC是通过NT-3/TrkC信号途径而产生促进NSC存活的作用。结果：和单纯DC组相比，共培养体系中，DC表达NT-3水平显著性增强（P<0.05)；同时共培养能显著增加NSC存活，而NT-3特异性中和性抗体能够选择性阻断该效应。共培养能够增加NSC中NT-3特异性受体TrkC表达，并上调TrkC磷酸化水平，而NT-3特异性抗体能够抑制TrkC表达和磷酸化上调。结论：DC和NSC共培养能够增加D
3、C中NT-3表达，转而通过NT-3/TrkC信号途径而增加NSC存活。[关键词]神经干细胞；树突状细胞；NT-3；TrkC；存活；信号通路。13DendriticCell-inducedNeuralStemCellSurvivalisdependentonNT-3/TrkCsignalingpathwayWangZhong1,ChenTaibang1,WangYongfei1,GuoQiaonan2,ZhaoJianhua*11DepartmentofSpinalSurgery,DapingHospital,InstituteofSurgeryResearch,T
4、hirdMilitaryMedicalUniversity,Chongqing,40042;2InstituteofPathology,SouthwestHospital,ThirdMilitaryMedicalUniversity,Chongqing,400038[Abstract]ObjectiveTostudytheeffectofdendriticcell(DC)onneuralstemcell(NSC)survivalwhencoculturedinvitro.MethodPrimaryDCandNSCfromSDratswerepurifiedands
5、eparatelycoculturedinvitro.TheexpressionofNT-3mRNAinDCwasdeterminedbyRT-PCRandtheapoptosisratioofNSCwasdetectedwithflowcytometrytodeterminetherelationshipbetweenNT-3expressionandNSCsurvival.Inaddition,theexpressionofNT-3specificreceptor,tropomyosinreceptorkinaseC(TrkC)inNSCwastestedwi
6、thimmunoflourescenceandwesternblottingtoconfirmthatNT-3/TrkCsignalingpathwayisinvolvedinNT-3inducedNSCsurvival.ResultsComparedwithsimpleDCgroup,coculturewithNSCsignificantlyincreasedtheexpressionofNT-3inDC(P<0.05);thesurvivalofNSCwasmarkedlyincreasedandthiseffectwasselectiveabrogatedb
7、yaNT-3specificneutralizingantibody.Inaddition,theexpressionandphosphorylationofNT-3specificreceptor,TrkCwasobviouslyenhancedinNSC,butinhibitedbyaNT-3specificneutralizingantibody.ConclusionCocultureofDCandNSCupregulatestheexpressionofNT-3inDCandpromotesthesurvivalofNSCreverselyinaNT-3/
8、TrkC-
