重庆医学
重慶醫學
중경의학
CHONGQING MEDICAL JOURNAL
2013年
33期
4048-4050
,共3页
海马%Hes1%神经分化%神经干细胞%逆转录-聚合酶链反应
海馬%Hes1%神經分化%神經榦細胞%逆轉錄-聚閤酶鏈反應
해마%Hes1%신경분화%신경간세포%역전록-취합매련반응
hippocampus%Hes1%neuronal differentiation%neural stem cells%RT-PCR
目的：探讨Hes1 mRNA在神经干细胞（NSC）向神经细胞分化过程中的表达变化。方法构建乳鼠海马NSC的细胞培养模型，对NSC增殖分化进行形态学观察。细胞分化前后，采用免疫化学染色法分别检测巢蛋白（Nestin）和神经元特异性烯醇化酶（NSE）的表达，以鉴定细胞类型。流式细胞仪检测细胞周期以确定细胞增殖活性。用逆转录-PCR（RT-PCR）检测NSC中Hes1 mRNA的表达变化情况。结果从海马分离培养的细胞生长状态良好，有克隆增殖能力，呈Nestin表达阳性，细胞分化后呈NSE表达阳性。流式细胞仪检测结果显示，诱导前NSC增殖活跃，其S期的细胞比例明显高于诱导分化各阶段的细胞比例（P＜0．01）。与诱导前NSC相比，分化细胞各阶段G0G1期细胞比例则显著增多（P＜0．01），细胞停滞于G0G1期。RT-PCR结果表明，Hes1 mRNA在NSC诱导分化前后均有表达，与诱导前相比，分化后的各阶段 Hes1 mRNA表达量均明显减少（P＜0．05），而分化各阶段 Hes1 mRNA表达量差异无统计学意义（P＞0．05）。结论 Hes1 mRNA 的高表达可能与 NSC的增殖有关，而Hes1 mRNA表达的减弱则有利于神经细胞的大量分化。
目的：探討Hes1 mRNA在神經榦細胞（NSC）嚮神經細胞分化過程中的錶達變化。方法構建乳鼠海馬NSC的細胞培養模型，對NSC增殖分化進行形態學觀察。細胞分化前後，採用免疫化學染色法分彆檢測巢蛋白（Nestin）和神經元特異性烯醇化酶（NSE）的錶達，以鑒定細胞類型。流式細胞儀檢測細胞週期以確定細胞增殖活性。用逆轉錄-PCR（RT-PCR）檢測NSC中Hes1 mRNA的錶達變化情況。結果從海馬分離培養的細胞生長狀態良好，有剋隆增殖能力，呈Nestin錶達暘性，細胞分化後呈NSE錶達暘性。流式細胞儀檢測結果顯示，誘導前NSC增殖活躍，其S期的細胞比例明顯高于誘導分化各階段的細胞比例（P＜0．01）。與誘導前NSC相比，分化細胞各階段G0G1期細胞比例則顯著增多（P＜0．01），細胞停滯于G0G1期。RT-PCR結果錶明，Hes1 mRNA在NSC誘導分化前後均有錶達，與誘導前相比，分化後的各階段 Hes1 mRNA錶達量均明顯減少（P＜0．05），而分化各階段 Hes1 mRNA錶達量差異無統計學意義（P＞0．05）。結論 Hes1 mRNA 的高錶達可能與 NSC的增殖有關，而Hes1 mRNA錶達的減弱則有利于神經細胞的大量分化。
목적：탐토Hes1 mRNA재신경간세포（NSC）향신경세포분화과정중적표체변화。방법구건유서해마NSC적세포배양모형，대NSC증식분화진행형태학관찰。세포분화전후，채용면역화학염색법분별검측소단백（Nestin）화신경원특이성희순화매（NSE）적표체，이감정세포류형。류식세포의검측세포주기이학정세포증식활성。용역전록-PCR（RT-PCR）검측NSC중Hes1 mRNA적표체변화정황。결과종해마분리배양적세포생장상태량호，유극륭증식능력，정Nestin표체양성，세포분화후정NSE표체양성。류식세포의검측결과현시，유도전NSC증식활약，기S기적세포비례명현고우유도분화각계단적세포비례（P＜0．01）。여유도전NSC상비，분화세포각계단G0G1기세포비례칙현저증다（P＜0．01），세포정체우G0G1기。RT-PCR결과표명，Hes1 mRNA재NSC유도분화전후균유표체，여유도전상비，분화후적각계단 Hes1 mRNA표체량균명현감소（P＜0．05），이분화각계단 Hes1 mRNA표체량차이무통계학의의（P＞0．05）。결론 Hes1 mRNA 적고표체가능여 NSC적증식유관，이Hes1 mRNA표체적감약칙유리우신경세포적대량분화。
Objective To explore the expression of Hes1 mRNA during neural stem cells(NSC) differentiation toward neurons . Methods To establish the model of cultivation NSC in the hippocampal of newborn (24 h) SD rats ,and then to observe the mor-phology of NSC in the course of proliferation and differentiation .Before and after cellular induction ,the expression of Nestin and NSE were respectively measured to detect cell types by immunochemistry method .And flow cytometry was used to determine cell cycle phases ,so as to detect proliferative activity of these cells .Meanwhile ,the expression of Hes1 mRNA in NSC was determined by reverse transcription-PCR(RT-PCR) .Results The results demonstrated that NSC isolated from hippocampal showed vigorously clonal proliferation in vitro ,and positive Nestin expression .In addition ,the differentiated cells demonstrated positive NSE expres-sion .Flow cytometry analysis showed that the percentage of NSC in S phase was obviously higher than that of induced differentia-tion of all time(P<0 .01) ,which indicated that NSC were actively dividing induction before .Compared to NSC ,the percentages of cells in G0 G1 phases were increased significantly after neuronal differentiation (P<0 .01) ,which indicated that differented cells have arrested in G0 G1 phases .Meanwhile ,the results from RT-PCR showed that :Hes1 mRNA was expressed in NSC both before and af-ter induced differention .Compared to induction before ,the level of Hes1 mRNA expression in NSC during different stages of differ-entiation after induction were significantly decreased (P<0 .05) ,and Hes1 mRNA did not show any obvious changes among these stages of differentiation(P>0 .05) .Conclusion The high level of Hes1 mRNA was probably involved proliferation of NSC .How-ever ,low level of Hes1 mRNA might contribute to neuronal differentiation .
