当归补血汤对感音神经性聋大鼠耳蜗干细胞移植后细胞凋亡的影响

The Effect of Astragalus Membranaceus and Angelica Sinensis on Grafted Cochlea Stem Cells Apoptosis in Hearing Impaired Rats

目的探讨感音神经性聋大鼠耳蜗干细胞移植后干细胞凋亡的时程特点及当归补血汤的抗凋亡作用。方法分离胚胎大鼠Corti器的耳蜗干细胞进行细胞培养,在相差显微镜下观察细胞形态及生长状况,用Nestin免疫荧光法进行耳蜗干细胞鉴定。建立感音神经性聋大鼠模型,将其分为耳蜗干细胞移植组(45只)、当归补血汤干预组(45只)及对照组(35只),分别将耳蜗干细胞悬液、含有当归补血汤的耳蜗干细胞悬液和生理盐水移植入三组大鼠耳蜗鼓阶,于移植后1、3、5、10、15天采用原位末端标记法结合流式细胞仪及免疫荧光方法对三组动物干细胞凋亡情况进行定量分析。结果移植后1、3、5、10、15天耳蜗干细胞移植组的耳蜗干细胞凋亡率分别为27.84%、49.37%、44.78%、39.37%、34.82%,当归补血汤干预组分别为18.35%、26.92%、23.18%、21.38%、18.19%,细胞凋亡在移植后3～5天达到高峰;当归补血汤干预组各时间点凋亡率均明显低于耳蜗干细胞移植组(P<0.05)。结论感音神经性聋大鼠耳蜗干细胞移植后3～5天是干细胞凋亡的高峰期;当归补血汤具有抗移植耳蜗干细胞凋亡的作用。

Objective To investigate the effects of astragalus membranaceus and angelica sinensis on grafted cochlea stem cells apoptosis in hearing imparied rats. Methods The hearing impaired rats were divided into 3 groups： cochlea stem cells group（45 rats）, astragalus membranaceus and angelica sinensis group（45 rats）, and the control group（35 rats）. Cochlea stem cells only were grafted into the first group cochlea regions,mixture of stem cells and astragalus membranaceus and angelica sinensis were grafted into that of the second group. Saline was grafted into that of control rats. Observed was apoptosis at 1 day, 3 days, 5 days, 10 days, 15 days after the transplantation by CASPASE--3 and Bcl--2 immunofluorescence. Results After transplantationn 1 day, 3 days, 5 days, 10 days, and 15 days of five time, for the cochlea stem celt group, the rates of apoptotic cells were 27.84%, 49.37%, 44. 78%, 39.37%, and 34.82%, respectively. The rates of astragalus membranaceus and angelica sinensis group were 18.35%, 26.92%, 23.18%, 21.38%, and 18.19%, respectively. The percent of grafted cochlea stem cells apoptosis in astragalus membranaceus and angelica sinensis group was much lower than that in cochlea stem cells group and control group. The majority of apoptotic cells occurred within three to five days after transplantation. Conclusion The majority of apoptotic cells occur within three to five days after transplantation in the hearing impaired rats. The astragalus membranaceus and angelica sinensis could reduce the apoptotic rates of transplanted stem cells.