CAJ | 학술논문

目的初步探讨c-kit在肠神经节细胞发育中的作用.方法体外培养大鼠肠神经节细胞及骨髓间充质干细胞(BMSCs),并在GDNF与胎肠培养基(FGCM)培养的条件下诱导BMSCs分化为肠神经节细胞.免疫组织化学方法鉴定原代培养的肠神经节细胞神经丝蛋白(neurofilament,NF)的表达,用血管活性肽(vasoactive intestinal peptide,VIP)及NF鉴定BMSCs诱导分化的肠神经节细胞.RT-PCR分别检测原代肠神经节细胞、BMSCs以及诱导之肠神经节细胞的c-kit的表达.结果免疫组化结果显示纯化后的肠神经节细胞表达NF,流式细胞学检测第四代后的BMSCs高表达CD90,而不表达CD45,诱导后的肠神经节细胞可表达VIP及NF,而其余两组则不表达.RT-PCR结果示原代肠神经节细胞及BMSCs不表达c-kit,诱导后的肠神经节细胞可表达c-kit.结论在体外能成功培养大鼠肠神经节细胞及BMSCs并纯化,BMSCs可被诱导为肠神经节细胞并表达肠神经递质.在肠神经系统的发育过程中,c-kit可能在某一阶段起到一定作用,在肠神经细胞逐渐发育成熟时接受到某些信号而关闭.
목적 초보탐토c-kit재장신경절세포발육중적작용.방법 체외배양대서장신경절세포급골수간충질간세포(BMSCs),병재GDNF여태장배양기(FGCM)배양적조건하유도BMSCs분화위장신경절세포.면역조직화학방법 감정원대배양적장신경절세포신경사단백(neurofilament,NF)적표체,용혈관활성태(vasoactive intestinal peptide,VIP)급NF감정BMSCs유도분화적장신경절세포.RT-PCR분별검측원대장신경절세포、BMSCs이급유도지장신경절세포적c-kit적표체.결과면역조화결과현시순화후적장신경절세포표체NF,류식세포학검측제사대후적BMSCs고표체CD90,이불표체CD45,유도후적장신경절세포가표체VIP급NF,이기여량조칙불표체.RT-PCR결과시원대장신경절세포급BMSCs불표체c-kit,유도후적장신경절세포가표체c-kit.결론 재체외능성공배양대서장신경절세포급BMSCs병순화,BMSCs가피유도위장신경절세포병표체장신경체질.재장신경계통적발육과정중,c-kit가능재모일계단기도일정작용,재장신경세포축점발육성숙시접수도모사신호이관폐.
Objective To investigate the possible role of c-kit on the development of enteric neurons. Methods Enteric neurons and bone marrow stromal cells (BMSCs) were harvested and cultured from rats. At passage 4,BMSCs were induced into enteric neurons by GDNF of 10ng/ml in fetal gut condition medium (FGCM) for 7 days. Primary enteric neurons were identified by neurofilament (NF) and induced enteric neurons were identified by va～active intestinal peptide (VIP) and NF with immunohistochemical assay. The expression of c-kit was detected by RT-PCR. Results After purification, primary enteric neurons showed expression of NF. BMSCs at passage 4 were CD90 positive and CD45 negative detected by flow cytometry. After 7 days of induction, induced enteric neurons expressed VIP,while BMSCs were VLP negative in the control group. Primary enteric neurons and BMSCs didn't express c-kit mRNA. The expression of c-kit mRNA of induced enteric neurons increased highly after induction. Conclusions Enteric neurons and BMSCs can be harvested and cultured in vitro,and BMSCs can be induced to differentiate into enteric neurons. C-kit may play a role at a certain stage of development of enteric nervous system and be blocked after receiving a signal of mature enteric neurons.