冻融小鼠卵巢移植后细胞凋亡及血管内皮生长因子表达的变化及意义研究

Expression and clinical significance of VEGF and apoptosis in frozen-thawed mouse ovaries after transplantation

目的:探讨冻融小鼠卵巢同种异体移植后细胞凋亡及血管内皮生长因子表达的变化及意义。方法:收集C57BL/6j雌鼠和BALB/c雄鼠杂交后F1代4周龄小鼠卵巢,慢冻速融后移植至杂交后F1代8～12周雄鼠的肾被膜下,分别于移植后1d(24h)、2d(48h)和7d回收移植物,将冻融以及移植后不同时间段的卵巢组织进行HE染色、全卵巢卵泡计数、电镜观察、免疫组织化学分析细胞凋亡及RT-PCR检测VEGF基因表达。结果:冻融小鼠卵巢移植后随着时间的推移、各级卵泡数和卵泡存活率逐渐下降;移植后48h内细胞凋亡指数最高;电镜观察发现小鼠卵巢组织移植后损伤主要发生在移植后48h内;移植后VEGF的表达有上升的趋势,至第7d仍维持较高水平;移植后48hVEGF120mRNA和VEGF188mRNA水平明显升高(P<0.05),至7d下降恢复至移植前水平,而VEGF164mRNA水平移植后无明显变化(P>0.05)。结论:小鼠卵巢组织移植后48h内细胞凋亡最为严重,移植后引起大量卵泡的丢失;在移植后血管化的过程中VEGFmRNA表达量增加,VEGF120mRNA和VEGF188mRNA可能参与卵巢移植后早期血管化过程。

Objective：To investigate expression and clinical significance of VEGF and apoptosis in frozen-thawed mouse ovaries after transplantation.Methods：Ovaries from B6C2F1（C57BL/6j×BALB/c） 4 week old mice were cryopreservation and the thawed ovaries were xenografted into kidney capsules of 8～12 week old adult male mice.The grafted were recovered 1 d,2 d and 7 d after transplantation respectively,the grafts and frozen-thawed were removed for follicle counting and immunohistochemically,ultrastructure,and detection of the mRNA expression of vascular endothelial growth factor（VEGF）.Results：The follicle numbers were decreased gradually after transplantation,the cell apoptosis increased especially in 48 h after transplantation.Transmission electron microscopy（TEM） showed the tissue damaged was severest 48 h after transplantation;the expression of VEGF increased after transplantation,peaked on day 7,the mRNA expression of VEGF120 and VEGF188 was more on 48 h after transplantation,decreased on day 7 （P0.05）.Conclusion：The number of follicles was decreased after transplantation,the apoptosis index was increased especially in 48 h after transplantation;the expression of VEGF increased after transplantation,an increase in the VEGF188 and VEGF164 isoform might suggest the positive effect in the early stages of angiogenesis.