连接蛋白40、45在连接蛋白43基因敲除小鼠胚胎心脏的时空表达

Spatio-temporal expression of connexion （Cx） 40 and Cx45 in Cx43 knockout embryonic mouse hearts

目的检测连接蛋白（Cx）40、45在Cx43基因敲除小鼠胚胎心脏的时空表达。方法选取Cx43基因敲除胎鼠，通过PCR方法鉴定其基因型，以Cx43基因敲除胎鼠纯合子作为研究对象，野生型为对照组，获取胎龄10．5、11．5、12．5、13．5、14．5、15．5d纯合子及野生型各2只胎鼠，免疫组化检测Cx40、Cx45在心脏各部位的表达，SCIM显微图像分析系统对染色强度进行定量分析。结果（1）野生型小鼠心室原始小梁网在胎龄10．5d就已经有Cx40表达（A值为8．6），随后在心房、心室、小梁网、房室间隔和主、肺动脉壁表达逐日增加，胎龄14．5d在心室部位达到高峰（A值为94．8），然后逐渐减少；Cx43基因敲除小鼠胚胎心脏发育中Cx40在各部位的时空表达趋势与野生型相似，胎龄10．5d在肌小梁A值为7．9，胎龄14．5d心室部位为75．8。纯合基因敲除型胎鼠Cx40的表达量明显弱于野生型。（2）野生型小鼠肌小梁部在胎龄10．5d出现Cx45的表达（A值为20．0），随后在房室各部位相继表达，胎龄12．5d在心房部位达到高峰（A值为49．6），然后表达逐渐减少；Cx43基因敲除小鼠胚胎心脏发育中Cx45在各部位的时空表达趋势与野生型相似，胎龄10．5d在肌小梁A值为17．8，胎龄12．5d心房部位为31．5。纯合基因敲除型胎鼠Cx45的表达量明显弱于野生型。结论Cx40和Cx45在Cx43基因敲除小鼠纯合子胎龄10．5～15．5d这一心脏分隔、瓣膜发育的关键时期表达异常，可能与Cx43基因敲除小鼠心脏的异常发育有关。

Objective To investigate the spatio-temporal expression of connexin （Cx） 40 and Cx45 genes in Cx43 knockout embryonic mouse hearts. Methods Cx43 knockout heterozygous mice were raised. PCR was performed to identify genotypes of their offsprings. The homozygute （ Cx43 - / - ） was used for study and the wild type （ Cx43 ＋ / ＋ ） was used as control. Immuno-histochemistry was used to detect the Cx40 and Cx45 expression in different parts of the fetal hearts at the embryonic days （EDs） 10.5, 11.5, 12.5,＊ 13. 5, 14. 5, and 15. 5, respectively. SCIM microscopic image analytic system was used for quantitative analysis of staining intensity. Results （ 1 ） Cx40 expression was detected in ventricles of Cx43 ＋/＋ fetal heart as early as ED10.5 with the intensity represented by A value of 8.6. Subsequently it was distributed in the atria and ventricles with the peak expression observed at ED14.5 （ A value = 94.8 ） , and faded afterwards. Less Cx40 expression was observed in the Cx43 -/- fetal hearts as compared with Cx43 ＋/＋ although its expression pattern was similar in both groups. （2） Cx45 expression was detected in ventricles at ED 10.5 （A value = 20.0）. It was subsequently distributed in the atria and ventricles with the peak expression observed at ED12.5 （A value = 49.6） , and then faded. Less Cx45 expression was observed in the Cx43 -/- fetal hearts as compared with Cx43 ＋/＋ although its expression pattern was similar as well in both groups. Conclusion Down-regulated expression of the genes Cx40 and Cx45 may be associated with the abnormal heart development in Cx43 knockout animals.