Cx43敲除小鼠胚胎心脏流出道内第二生心区和心脏神经嵴来源间充质细胞减少

A downtrend of mesenchymal cells derived from the second heart field and cardiac neural crest in the outflow tract of Cx43 knockout embryonic mouse heart

背景:Cx43被认为在人类先天性心脏病中起着重要的作用,目前对于Cx43基因敲除后形成心脏畸形的相关机制尚不清楚。目的:探讨敲除Cx43后心脏发育缺陷以及第二生心区和心脏神经嵴前体细胞的迁移分化是否异常。方法:取胚龄10-13 d的Cx43基因敲除纯合(Cx43^-/-)小鼠和Cx43野生型(Cx43^+/+)小鼠胚胎石蜡包埋连续切片,进行免疫组织化学和免疫荧光显色以及心脏的三维重建。结果与结论:①Cx43基因敲除后的小鼠在胚龄10-11 d,Isl1阳性第二生心区细胞仍由前肠腹侧经弓动脉延续至心包腔背侧壁,以及经鳃弓核心间充质、心包腔背侧壁延续至流出道远端;至胚龄13 d,升主动脉及肺动脉干形成后,其管壁以及心室左、右流出道壁均可见Isl1阳性细胞的分布,但Isl1阳性细胞数量明显减少(P<0.01);②Cx43基因敲除后小鼠在胚龄10-11 d,Ap2α阳性神经嵴细胞仍可见于弓动脉壁以及动脉囊背侧与腹侧壁,但神经嵴细胞数量少于野生型小鼠胚胎(P<0.01);③结果说明,Cx43基因敲除后第二生心区Isl1阳性细胞和Ap2α阳性神经嵴细胞的迁移路径和分布模式未发生异常,只是表现为这两种细胞迁移数量的减少,由此提示除心脏神经嵴来源细胞外,第二生心区前体细胞数量的减少可能也与Cx43基因缺失后流出道的畸形发生相关。

BACKGROUND:Cx43 plays an important role in human congenital heart disease.However,there is still no consistent conclusion about the formation mechanism of cardiac malformation in Cx43 knockout mouse embryos.OBJECTIVE:To investigate the cardiac development defects and the migration and differentiation of progenitor cells of the second heart field and cardiac neural crest in Cx43 knockout mouse embryo.METHODS:Serial sections of Cx43 gene knockout homozygous(Cx43^-/-)mouse and Cx43 wild-type(Cx43^+/+)mouse embryos from embryonic day(ED)10 to ED13 were made for immunohistochemical and immunofluorescent staining,and three-dimensional reconstruction of the heart.RESULTS AND CONCLUSION:(1)In Cx43 gene knockout mouse embryos at ED10-ED11,Isl1 positive second heart field cells in the ventral mesenchyme of the foregut extended through the area between the bilateral arch arteries to the dorsal wall of pericardial cavity.Meanwhile,Isl1 positive cells in the core mesenchyme of the branchial arches were continuous with those in the dorsal wall of pericardial cavity and the distal wall of the outflow tract.At ED13,the distribution of Isl1 positive cells was observed in the wall of the ascending aorta and pulmonary trunk as well as the wall of the left and right outflow tracts of the septated ventricles.However,compared with wild-type mouse embryos,fewer Isl1 positive second heart field cells were found in Cx43 gene knockout mouse embryos(P<0.01).(2)During ED10 to ED11,Ap2αpositive neural crest cells were still found in the wall of the arch artery and the dorsal and ventral walls of the aortic sac in Cx43 gene knockout mouse embryos,but the number of neural crest cells was less than that of wild-type mouse embryos(P<0.01).(3)These results indicate that the migration path and distribution pattern of Isl1 positive second heart field cells and Ap2αpositive cardiac neural crest cells are similar between the Cx43 gene knockout and wild-type mouse embryos,but the number of two kinds of migrating cells is reduced after Cx43 gene knockout.This suggests that in addition to cardiac neural crest derived cells,the decrease of second heart field progenitor cells might be involved in the formation of outflow tract malformations in Cx43 knockout mouse embryos.