生理学报
生理學報
생이학보
ACTA PHYSIOLOGICA SINICA
2006年
6期
547-555
,共9页
分化%增殖%胆碱能神经元%胚胎神经干细胞%溶血磷脂酸
分化%增殖%膽堿能神經元%胚胎神經榦細胞%溶血燐脂痠
분화%증식%담감능신경원%배태신경간세포%용혈린지산
differentiation%proliferation%cholinergic neuron%embryonic neural stem cells%lysophosphatidic acid
溶血磷脂酸(lysophosphatidic acid,LPA)是一种细胞外磷脂信号.本研究用[3H]-胸腺嘧啶掺入法、免疫细胞化学和Western blot等技术,观察了LPA对体外培养的大鼠胚胎神经干细胞(neural stem cells,NSCs)的增殖以及向MAF2标记的一般神经元和ChAT标记的胆碱能神经元的分化的影响.结果显示:(1)在特殊的无血清培养基中加入低浓度的LPA(0.01～1.0 gmol/L)后,NSCs对[3H]-胸腺嘧啶的摄入呈剂量依赖性增加,表明LPA对NSCs有显著的促增殖作用;(2)在培养基中加入胎牛血清以诱导NSCs的分化,发现低浓度的LPA增加MAP2阳性和ChAT阳性神经元的比例,0.1 gmol/L LPA引起的增加达到峰值;(3)Westem blot分析显示LPA促进了MAP2和ChAT的表达;(4)在诱导NSCs出现分化早期,用倒置显微镜观察到低浓度的LPA明显促进细胞突起的生长和细胞的迁移.以上结果表明,低浓度LPA在一定范围内可以促进NSCs的增殖、并分化为一般的MAP2阳性神经元和特殊的胆碱能神经元,而且LPA可以促进在分化早期出现的神经元或神经胶质细胞前体细胞的迁移和突起生长.
溶血燐脂痠(lysophosphatidic acid,LPA)是一種細胞外燐脂信號.本研究用[3H]-胸腺嘧啶摻入法、免疫細胞化學和Western blot等技術,觀察瞭LPA對體外培養的大鼠胚胎神經榦細胞(neural stem cells,NSCs)的增殖以及嚮MAF2標記的一般神經元和ChAT標記的膽堿能神經元的分化的影響.結果顯示:(1)在特殊的無血清培養基中加入低濃度的LPA(0.01～1.0 gmol/L)後,NSCs對[3H]-胸腺嘧啶的攝入呈劑量依賴性增加,錶明LPA對NSCs有顯著的促增殖作用;(2)在培養基中加入胎牛血清以誘導NSCs的分化,髮現低濃度的LPA增加MAP2暘性和ChAT暘性神經元的比例,0.1 gmol/L LPA引起的增加達到峰值;(3)Westem blot分析顯示LPA促進瞭MAP2和ChAT的錶達;(4)在誘導NSCs齣現分化早期,用倒置顯微鏡觀察到低濃度的LPA明顯促進細胞突起的生長和細胞的遷移.以上結果錶明,低濃度LPA在一定範圍內可以促進NSCs的增殖、併分化為一般的MAP2暘性神經元和特殊的膽堿能神經元,而且LPA可以促進在分化早期齣現的神經元或神經膠質細胞前體細胞的遷移和突起生長.
용혈린지산(lysophosphatidic acid,LPA)시일충세포외린지신호.본연구용[3H]-흉선밀정참입법、면역세포화학화Western blot등기술,관찰료LPA대체외배양적대서배태신경간세포(neural stem cells,NSCs)적증식이급향MAF2표기적일반신경원화ChAT표기적담감능신경원적분화적영향.결과현시:(1)재특수적무혈청배양기중가입저농도적LPA(0.01～1.0 gmol/L)후,NSCs대[3H]-흉선밀정적섭입정제량의뢰성증가,표명LPA대NSCs유현저적촉증식작용;(2)재배양기중가입태우혈청이유도NSCs적분화,발현저농도적LPA증가MAP2양성화ChAT양성신경원적비례,0.1 gmol/L LPA인기적증가체도봉치;(3)Westem blot분석현시LPA촉진료MAP2화ChAT적표체;(4)재유도NSCs출현분화조기,용도치현미경관찰도저농도적LPA명현촉진세포돌기적생장화세포적천이.이상결과표명,저농도LPA재일정범위내가이촉진NSCs적증식、병분화위일반적MAP2양성신경원화특수적담감능신경원,이차LPA가이촉진재분화조기출현적신경원혹신경효질세포전체세포적천이화돌기생장.
Effects of lysophosphatidic acid(LPA),an extracellular phospholipid signal,on the proliferation of rat embryonic neural stem cells fNSCs)and their differentiation into microtubule-associated protein 2(MAP2)-positive and choline acetyltransferase (CHAT)-positive,i.e.cholinergic-committed neurons,were observed in vitro by[3H]-thymidine incorporation,immunocytochemistry,Western blot and other techniques.The results showed that:(1)Lower concentrations of LPA(0.01～1.0μmol/L)dose-dependently enhanced the uptake of[3H]-thymidine by NSCs cultured in specific serum-free medium,indicating a significant promotlve action of LPA on the Droliferation of NSCs.(2)After fetal bovine serum which induces and commences the differentiation of NSCs,was used in the medium.the lower concentrations of LPA increased the percentages of both MAP2-and ChAT-immunoreactive neurons,with a Deak at 0.1 μmol/L LPA in two cases.(3)The promotive effects of LPA on the differentiation of MAP2-and ChAT-positive neurons were also supported by the up-regulation of the expressions of both MAP2 and ChAT proteins detected by Western blot.(4)At the earlv phase of differentiation of NSCs,the cell migration and neurite extension were enhanced significantly by lower dosages of LPA under phase-contrast microscope.These results suggest that LPA within certain lower range of concentrations promotes the proliteration of NSCs and their differentiation into unspecific MAP2-positive and specific cholinergic-committed neurons,and also strengthens the migration and neurite extension of the newly-generated neuronal (and also glial as reported elsewhere)progenitors.
