wnt信号通路介导的基因调控毛囊干细胞定向分化的分析-analysis of gene regulation and differentiation of hair follicle stem cells mediated by wnt signaling pathway.docxVIP

ml(约5mmol/L)时明显抑制细胞生长。实验研究发现在HFSCs分化过程中，LiCl可激活Wnt/β-catenin信号通路，当浓度为5mM/l时促使β-catenin及其拮抗物GSK3β、核内转录因子Tcf3、下游基因c-myc、cyclinD1的转录上调，浓度为10mM/l时除cyclinD1转录仍上调外，其余基因转录下调。3.扫描电镜观察细胞在PLGA-ADM材料上生长良好，分泌的胞外基质丰富，并随时间进展，细胞逐渐融合，细胞外基质分泌逐渐增多。HE染色观察发现HFSCs生长状态良好，单层排列分布于ADM表皮面上，LiCl干预组表皮面有毛囊样小凹形成，成纤维细胞与ADM真皮面贴附紧密，能分泌胞外基质形成连续细胞层包绕ADM。结论1.我们改进的培养方法能简便快捷的获取人HFSCs，且得到的人HFSCs在体外有较强的增殖及传代能力，1×104/ml接种密度是促进HFSCs增殖的最佳密度。2.LiCl促HFSCs分化明显，对HFSCs的干预有临界毒性浓度，当浓度大于200μg/ml时，细胞生长增殖受到明显抑制，同时这一浓度也是促HFSCs分化的最佳浓度。LiCl促HFSCs分化作用可能与激活Wnt/β-catenin信号通路、通路下游各因子转录上调相关，Tcf3在维持细胞未分化状态中发挥了一定的作用，cyclinD1的转录与氯化锂浓度作用呈正相关，在HFSCs的分化中起到重要作用。3.复乳-溶剂挥发法能快速高效的制备出载有LiCl的纳米微囊，其PLGA材料具有良好的细胞相容性，人HFSCs能在其上较好的粘附和生长。构建载有人HFSCs的组织工程皮肤，在LiCl诱导下能够促使种子细胞发生分子学改变形成毛囊样小凹，使构建带有皮肤附属器的组织工程替代物成为可能。【关键词】毛囊干细胞氯化锂Wnt信号通路纳米微囊定向分化ABSTRACTBackgroundandObjectiveIthasbecomeahotspotinmedicalresearchesthathairfolliclestemcells(HFSCs)areselectedtobeseedcellstoconstructtissueengineeredskintorepairskinmissingcau-sedbyburns,traumasandchroniculcers.Thehairfollicleisanimportantsubsidiaryorganoftheskin,andisconsistofhairshaft,internalrootsheathandexternalrootsheath.T-hehairfolliclebulgeisintheexternalrootsheathneartheepidermalside,atthearrectormuscleattachingbelowsebaceousglands.Thecellsatthisplaceculturedinvitroturno-uttobecharacteristicsofstemcells,whichareabilitiesofself-duplicating,slowcyclicityandmulti-directionaldifferentiation.ThesecellsareprovedtobeHFSCs.HFSCshavetheabilityofmulti-directionaldifferentiation,whichparticipateepidermalrenewal,sebaceousglandsmaintainingandhairfollicleformation.ThedifferentiationofHFSCsisco-instructedbymulti-factors,includingelementsandgenesincellsandmicroenvironment,evensignalingpathways.Withtheeffortsofmanyresearchers,WntsignalingpathwayisprovedtohavecriticaleffectsonproliferationanddifferentiationinHFSCs.Wnt/β-cateninsignalingpathwayexistsinnearlyallcells.Wntproteinisreleasedaftercellstimulation,activatesaseriesofmolecularsignalingsubstancesindownstreamandf-inallychangesgeneexpression,whichisamolecularcascade