Department of Pediatric Surgery, Shengjing Hospital of China Medical University, Shenyang 110004, P.R. China.
Exp Biol Med (Maywood). 2018 Feb;243(4):327-333. doi: 10.1177/1535370217751073. Epub 2018 Jan 7.
Congenital anorectal malformation is the most common digestive tract malformation in newborns. It has been reported that FOXD3/FOXD4, a forkhead transcription factor, regulates the generation, migration, and differentiation of neural crest cells. However, whether FOXD3/FOXD4 takes part in anorectal malformation remains unclear. In the present study, we used ethylene thiourea to induce the animal models of anorectal malformation in rat embryos and to interrogate the role of FOXD3/FOXD4 in anorectal malformation pathogenesis. Hindgut samples of the animal models were collected at E15, E17, E19, and E21 days of age. The expression of FOXD3/FOXD4 was detected by immunohistochemistry, western blot, and quantitative real-time fluorescence PCR. By immunohistochemical staining, FOXD3/FOXD4 was observed in epithelial cells of the rectum and the anus both in normal and rat embryos with anorectal malformation. Expression level analysis by western blot indicated that FOXD3/FOXD4 expression increased in ethylene thiourea-induced anorectal malformation groups. mRNA expression as determined by quantitative real-time fluorescence PCR analysis was consistent with the western blot results. Tentative conclusions were drawn that FOXD3/FOXD4 is expressed in the hindgut in rat embryos and is upregulated in anorectal malformation. FOXD3/FOXD4 is required for the development of the hindgut, and its aberrant expression may be an important factor leading to the incidence of anorectal malformation. Impact statement Congenital anorectal malformation (ARM) is the most common digestive tract malformation in newborns. The pathophysiological ground remains unclear. In this study, we used animal models of ARM for the first time to interrogate the role of FOXD3/FOXD4 in ARM pathogenesis. The animal models of ARM were successfully induced by ethylene thiourea (ETU) in rat embryos providing a strong basis for pathogenesis study of this disease. Expression analysis of FOXD3/FOXD4 was carried out in these models, and the results shape a deeper understanding of FOXD3/FOXD4 being required for the normal development of the hindgut. The aberrant expression of FOXD3/FOXD4 may be an important factor leading to ARM incidence.
先天性肛门直肠畸形是新生儿最常见的消化道畸形。有报道称，叉头框转录因子 FOXD3/FOXD4 调节神经嵴细胞的生成、迁移和分化。然而，FOXD3/FOXD4 是否参与肛门直肠畸形尚不清楚。本研究采用乙硫尿（ETU）诱导大鼠胚胎肛门直肠畸形动物模型，探讨 FOXD3/FOXD4 在肛门直肠畸形发病机制中的作用。收集 E15、E17、E19 和 E21 天龄动物模型的后肠样本，通过免疫组织化学、Western blot 和实时荧光定量 PCR 检测 FOXD3/FOXD4 的表达。免疫组织化学染色显示，FOXD3/FOXD4 在正常和 ETU 诱导的肛门直肠畸形大鼠胚胎的直肠和肛门上皮细胞中均有表达。Western blot 分析表明，ETU 诱导的肛门直肠畸形组 FOXD3/FOXD4 表达增加。实时荧光定量 PCR 分析的 mRNA 表达与 Western blot 结果一致。结论认为 FOXD3/FOXD4 在大鼠胚胎的后肠中表达，并在肛门直肠畸形中上调。FOXD3/FOXD4 是后肠发育所必需的，其异常表达可能是导致肛门直肠畸形发生的重要因素。
