Spatiotemporal pattern analysis of transcription factor 4 in the developing anorectum of the rat embryo with anorectal malformations
Spatiotemporal pattern analysis of transcription factor 4 in the developing anorectum of the rat embryo with anorectal malformations
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DOI：
10.1007/s00384-009-0705-3
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发表时间：
2009-09-01
影响因子：
2.8
通讯作者：
Wang, Wei Lin
中科院分区：
文献类型：
--
作者：
Zhang, Tao;Bai, Yu Zuo;Wang, Wei Lin
作为转录因子家族的一员，转录因子 4 (Tcf4) 已知可影响内胚层衍生组织（包括肺、肝、胰腺、胃和肠）的基因表达。然而，目前尚不清楚这种能力在正常和肛门直肠畸形（ARM）大鼠胚胎的肛门直肠发育过程中是否活跃。在本研究中，引入乙撑硫脲（ETU）诱导的ARM模型，通过免疫组化染色、逆转录酶聚合酶链反应（RT-PCR）和蛋白质印迹分析研究肛门直肠发育过程中Tcf4的表达模式。免疫染色显示，Tcf4表达在发育过程中表现出空间依赖性变化。肛门直肠：在正常胚胎中，Tcf4蛋白最初在胚胎第13天（E13）的尿直肠隔膜（URS）和后肠的背侧内胚层中表达。此外，单独的表达域在 E14 的背侧 CM 上集中发育。在 E15 上，在 URS 的融合组织中检测到阳性细胞，尤其是在肛膜中。然而，在 ARM 胚胎中，泄殖腔、URS 和肛门直肠上皮的 Tcf4 呈阴性或微弱。 Western blot和RT-PCR结果显示，肛肠发育过程中Tcf4蛋白和mRNA表达量随时间变化显着：E14、E14.5和E15表达量达到峰值； E16后，Tcf4表达逐渐下降。相反，在ARM胚胎中，从E13到E16的肛门直肠形态发生过程中，Tcf4的时空表达不平衡。这些数据表明，在泄殖腔分离为直肠和尿道时Tcf4的下调可能与ARM的发育有关。
As a member of the transcription factors family, transcription factor 4(Tcf4) is known to influence gene expression in endodermally derived tissues including lung, liver, pancreas, stomach, and intestine. However, it remained unknown if this capability is active during anorectal development in the normal and anorectal malformations (ARM) rat embryos.In this study, ethylenethiourea (ETU)-induced ARM model was introduced to investigate the expression pattern of Tcf4 during anorectal development using immunohistochemical staining, reverse transcriptase polymerase chain reaction (RT-PCR), and Western blot analysis.Immunostaining revealed that Tcf4 expression showed space-dependent changes in the developing anorectum: in normal embryos, Tcf4 protein is initially expressed in the dorsal endoderm of urorectal septum (URS) and hindgut on embryonic day 13 (E13). Additionally, separate expression domain develops intensively on the dorsal CM on E14. On E15, positive cells are then detected in the fused tissue of URS, and prominently in the anal membrane. In the ARM embryos, however, the epithelium of the cloaca, URS, and anorectum was negative or faint for Tcf4. In Western blot and RT-PCR, time-dependent changes of Tcf4 protein and mRNA expression were remarkable during the anorectal development: on E14, E14.5, and E15, the expression level reached the peak; after E16, Tcf4 expression gradually decreased. In contrast, in ARM embryos, spatiotemporal expression of Tcf4 was imbalanced during the anorectal morphogenesis from E13 to E16.These data implied that the downregulation of Tcf4 at the time of cloacal separation into rectum and urethra might be related to the development of ARM.