Gao Fang, Yi Qi-Jian, Tian Yun-Jiao, Wang Fang-Jie, Liu Guan-Xin
Heart Center, Children's Hospital, Chongqing Medical University, Chongqing 400014, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2012 Mar;43(2):161-5.
To investigate the role of nesprin-1 in mouse embryonic stem cells differentiation into cardiomyocyte.
Hanging drop-suspension-adherence method was applied for the differentiation of mouse embryonic stem cells into cardiomyocytes under the inducing of salvia miltorrhiza and 5-azacytidine. Changes in nesprin-1 gene expression were detected by using Western blotting and immunofluorescent assay. RNA interference was used to reduce nesprin-1 protein levels to further investigate the importance of nesprin-1 in mouse embryonic stem cells differentiation, group I (target sequence AAAGCCAAGCACGCAACTA), group II (target sequence GGGAACCAACAGTGAGATT), group III (target sequence ACCAGGACATTGCGTACTA), and group IV (control group).
The nesprin-1 isoform profile was altered in mouse embryonic stem cells differentiation. The rates of differentiation of the four groups were (17.78 +/- 1.92)%, (36.67 +/- 3.34)%, (44.42 +/- 5.08)%, (77.78 +/- 1.92)%; The rate of differentiation of group IV was higher than RNAi groups and the difference was significant (P < 0.05). In addition, compared with the control group, myosin in RNAi groups were dramatically reduced.
Nesprin-1 played important roles in mouse embryonic stem cells differentiation into cardiomyocyte. Nesprin-1 isoforms might perform different functions in the process of mouse embryonic stem cells differentiation.
探讨核膜伸展蛋白-1（nesprin-1）在小鼠胚胎干细胞向心肌细胞分化中的作用。
采用悬滴-悬浮-贴壁法，在丹参和5-氮杂胞苷诱导下将小鼠胚胎干细胞分化为心肌细胞。运用蛋白质免疫印迹法和免疫荧光分析法检测nesprin-1基因表达的变化。采用RNA干扰降低nesprin-1蛋白水平，进一步研究nesprin-1在小鼠胚胎干细胞分化中的重要性，分为Ⅰ组（靶序列AAAGCCAAGCACGCAACTA）、Ⅱ组（靶序列GGGAACCAACAGTGAGATT）、Ⅲ组（靶序列ACCAGGACATTGCGTACTA）和Ⅳ组（对照组）。
在小鼠胚胎干细胞分化过程中，nesprin-1异构体谱发生改变。四组的分化率分别为（17.78±1.92）%、（36.67±3.34）%、（44.42±5.08）%、（77.78±1.92）%；Ⅳ组的分化率高于RNA干扰组，差异有统计学意义（P<0.05）。此外，与对照组相比，RNA干扰组的肌球蛋白显著减少。
nesprin-1在小鼠胚胎干细胞向心肌细胞分化中起重要作用。nesprin-1异构体在小鼠胚胎干细胞分化过程中可能发挥不同功能。
