CAJ | 학술논문

　　目的研究同型半胱氨酸（Hcy）对体外大鼠神经干细胞(NSCs)增殖及Notch信号通路相关基因Notch1和Hes1 mRNA表达的影响。方法采用无血清培养原代培养新生大鼠NSCs，细胞分为对照组(Hcy-C)、Hcy低剂量组（Hcy-L）、Hcy中剂量组（Hcy-M）、Hcy高剂量组（Hcy-H）。采用免疫组化法检测Nestin、β-tubulinⅢ及GFAP的表达对NSCs进行鉴定，MTT法检测细胞增殖情况，Real-time PCR检测Notch1、Hes1基因mRNA表达。结果无血清培养条件下，所培养细胞形成大量呈Nestin抗原阳性细胞组成的神经球；诱导分化6 d后，形成β-tubulinⅢ表达呈阳性的神经元和GFAP表达呈阳性的星形胶质细胞；Hcy干预组细胞活力低于对照组，且有剂量依赖性(P<0.05)。Hcy干预组Hes1 mRNA表达低于对照组(P<0.05)。Hcy-H组Notch1 mRNA表达低于其他3组(P<0.05)；Hcy-M组Notch1 mRNA表达低于对照组和低剂量组(P<0.05)。结论 Hcy能抑制大鼠新生鼠NSCs的增殖，可能与下调Notch信号通路中Notch1和Hes1 mRNA表达有关。
　　목적연구동형반광안산（Hcy）대체외대서신경간세포(NSCs)증식급Notch신호통로상관기인Notch1화Hes1 mRNA표체적영향。방법채용무혈청배양원대배양신생대서NSCs，세포분위대조조(Hcy-C)、Hcy저제량조（Hcy-L）、Hcy중제량조（Hcy-M）、Hcy고제량조（Hcy-H）。채용면역조화법검측Nestin、β-tubulinⅢ급GFAP적표체대NSCs진행감정，MTT법검측세포증식정황，Real-time PCR검측Notch1、Hes1기인mRNA표체。결과무혈청배양조건하，소배양세포형성대량정Nestin항원양성세포조성적신경구；유도분화6 d후，형성β-tubulinⅢ표체정양성적신경원화GFAP표체정양성적성형효질세포；Hcy간예조세포활력저우대조조，차유제량의뢰성(P<0.05)。Hcy간예조Hes1 mRNA표체저우대조조(P<0.05)。Hcy-H조Notch1 mRNA표체저우기타3조(P<0.05)；Hcy-M조Notch1 mRNA표체저우대조조화저제량조(P<0.05)。결론 Hcy능억제대서신생서NSCs적증식，가능여하조Notch신호통로중Notch1화Hes1 mRNA표체유관。
Objective To explore the effects of homocysteine (Hcy) on neural stem cell (NSC) proliferation and the mRNA expression level of Notch1 and Hes1 related Notch signaling pathway from neonatal rats in vitro. Methods NSCs from neonatal rats were cultured by serum-free culture method in vitro. Cells were divided into four groups: control group (Hcy-C), low dose Hcy (Hcy-L) group, middle dose Hcy (Hcy-M) group and high dose Hcy (Hcy-H) group. NSCs were iden- tified by immunofluorescent staining using the antibodies against Nestin, β-tubulin Ⅲ and GFAP. The proliferation ability of NSCs was detected by MTT. The mRNA expressions of Notch1 and Hes1 were detected by Real-time PCR. Results In the serum free suspension medium, neurospheres that consisted of a great number of nestin-positive cells were found. β-tu- bulin Ⅲ positive neurons and GFAP positive astrocytes were detected by immunofluorescence staining on the 6 th day of cell induction. MTT assay showed that the cell viability was significantly lower in three Hcy treatment groups than those of con- trol group (P < 0.05). And the effect of concentration-dependent was observed. The results of RT-PCR showed that mRNA expression of Hes1 was significantly lower in three Hcy treatment groups than that in control group (P < 0.05). The mRNA ex- pression of Notch1 was significantly lower in Hcy-H group than that of other three groups (P < 0.05). The mRNA expression of Notch1 was significantly lower in Hcy-M group than that of Hcy-L group and control group (P < 0.05). Conclusion Hcy could inhibit the proliferation of NSCs by down-regulating mRNA expression levels of Notch1 and Hes1 genes related to Notch signal pathway.