应用FRAP技术分析小鼠嵌合体和正常胚胎发育中细胞间隙连接介导通讯

应用激光扫描共聚焦显微镜的光漂白恢复(fluorescence redistribution after photobleaching,FRAP)技术分析小鼠嵌合体胚胎和正常胚胎的卵裂球之间细胞间隙连接介导通讯(gap junctional inter-cellular communica-tion,GJIC),结果发现:8-细胞期嵌合体胚胎的光漂白恢复率(24.3%)明显低于正常胚胎(64.2%),提示GJIC的降低可能是影响嵌合体胚胎发育率降低的因素之一;囊胚的光漂白恢复率也较低(22.7%),提示随着细胞分化,GJIC的水平有所降低。

Analysis of Gap Junctional Intercellular Communication in Mouse Preimpantation and Chimera Embryos by Fluorescence Redistribution after Photobleaching

In order to analyze the gap junctional inter-cellular communication(GJIC) between mouse blastomeres,we produced chimeric embryos by 2-cell blastomere exchange,and observed the fluorescence redistribution in normal and chimeric embryos after photobleaching(FRAP) and followed recovery by laser scanning confocal microscopy.The result showed that there were almost no recoveries after photobleaching at 2-cell and 4-cell stage.However,15 min after photobleaching in one blastmere of 8-cell embryo,the relative intensity was greatly recovered.Interestingly,the recovery rate of chimeric embryos at 8-cell stage was significantly lower than that of the normal embryos at the same stage(24.3 % vs 64.2 %),but similar to that in blastocyst.Our results indicated that decreased GJIC might be responsible for the lower developmental potential of chimera embryos.The low recovery rate in blastocyts may suggest that GJIC level is degraded gradually following the differentiation.