肠和脑来源神经干细胞的nNOS表达研究

Study of Expression of nNOS in the NSCs of Enteric and Cerebral Origins

目的对同一胎鼠的肠神经干细胞和脑神经干细胞进行体外培养,比较两种不同来源神经干细胞的神经型一氧化氮合酶(nNOS)的表达情况。方法从孕20天的SD胚鼠中分别提取肠神经干细胞和脑神经干细胞,采用改良的神经干细胞培养液反复传代。取传代培养6～10天后得到的两种不同来源的神经干细胞球,通过琼脂糖预包埋免疫细胞化学检测法及流式细胞仪检测法测定神经干细胞nNOS的表达。结果通过本实验室改良的神经干细胞培养液及分离方法能培养出两种不同来源的神经干细胞。琼脂糖预包埋免疫细胞化学检测显示,两种不同来源神经干细胞的nNOS表达,分别与相应对照组比较,均有统计学显著性差异(P<0.05),两种不同来源神经干细胞间的nNOS表达比较,也均有统计学显著性差异(P<0.05)。流式细胞仪检测显示,nNOS阳性细胞在肠和脑来源神经干细胞分别占14.13%和5.80%,有统计学显著性差异(P<0.05)。结论两种不同来源的神经干细胞中均有表达nNOS,但消化道来源的肠神经干细胞nNOS表达较中枢神经系统来源的神经干细胞高。

Objective To investigate and compare the expression of nNOS in the NSCs of two origins by culture in vitro of the EN- SCs and CNS - NSCs from the same fetal rat. Methods The ENSCs and CNS - NSCs were extracted from the same 20 - day SD rat em- bryos by repeated subculture adopting the improved neural stem cell medium. Two different kinds of nerve bulbs originating from 6 - 10 days＇ subculturing were respectively divided into two groups,the control group and nNOS group. The expressions of nNOS in two kinds of NSCs were detected by immunohistochemistry assay with agarose pre - embedding and FCM. Results Our improved culture medium and the isolation methods can bring up two kinds of NSCs of different sources. After analyzing the data obtained by imunohistochemistry assay with agarose pre- embedding, we feund that the differences of relative parameter were statistically significant （P 〈 0.01 ） between the nNOS groups and corresponding control groupsand between two nNOS groups. FCM indicated nNOS - positive cells accounted for 14.13% and 5.80% of the enteric and cerebral original NSCs respectively. Conclusion nNOS is expressed both in two kinds of NSCs, but the quantity of nNOS expressed in the NSCs of digestive tract origin is higher than those of cerebral origin.