Conditional deletion of platelet derived growth factor receptor alpha (Pdgfr[alpha]) in urorectal mesenchyme causes defective urorectal development in mice

Abstract

In mammals, the urorectal development starts at an early embryonic stage, and is divided into three phases: the genital tubercle (GT) outgrowth, the cloaca septation and the urethra tubularization. The defective urorectal development results in anorectal malformations (ARMs), which is the most common congenital urorectal defects in newborns. Although multiple genes and signaling pathways have been implicated in the pathogenesis, the underlying etiology and embryology of ARMs remain largely unknown. Platelet derived growth factor receptor alpha (Pdgfrα) was identified by Affymetrix array analysis for genes/pathways implicated in urorectal development at our laboratory, and down-regulation of Pdgfrα expression was associated with urorectal defects in ETU-induced ARMs model. Pdgfrα is a cell surface receptor tyrosine kinase, upon binding to its ligands (Pdgf-a, -b and -c), controls cellular proliferation, survival, apoptosis, migration and differentiation both in embryos and in adults. Firstly, I showed that the expression of Pdgfrα is tightly regulated in the developing urorectal mesenchyme, and is correlated closely with the GT outgrowth, the cloaca septation of mouse embryos. I hypothesize that (1) Tempo-spatial expression of Pdgfrα is crucial for the GT outgrowth and the cloaca septation of the external genitalia development, and (2) Perturbation of Pdgfrα results in the defective urorectal development. Deletion of Pdgfrα causes early embryo lethality which precludes the investigation of regulatory functions of Pdgfrα in the urorectal development. To demonstrate the roles of Pdgfrα in the urorectal development, I employed a conditional deletion of Pdgfrα gene approach by the tamoxifen inducible Crerecombination, and investigated the urorectal developmental defects of mice in details. I conditionally deleted Pdgfrα in urorectal mesenchyme in mice at different embryonic stages, and investigated the urorectal defects of these mutants. Both female and male mutant embryos exhibited ARMs-like phenotypes including incomplete cloaca septation, anteriorly displaced anus, defective development of the urogenital folds, and the midline unfused urethra. I also detected an elevated apoptosis in the urorectal mesenchyme of mutant embryos, associated with p53 phosphorylation and Caspase3 activation. In conclusion, my study shows that (1) the correct expression and timing of Pdgfrα are essential for the urorectal development; (2) dysregulation of Pdgfrα expression results in defective cloaca septation and multiple urorectal defects; and (3) Pdgfrα may function as a survival factor and modulate mesenchymal cell death in the urorectal development. In summary, my study demonstrates for the first time that Pdgfrα plays an important role in the urorectal development and conditional knockout of Pdgfrα in Pdgfrα-expressing tissues of mouse embryos resulted in urorectal defects which resemble the congenital developmental anomalies of human ARMs. The remarkable resemblance of conditional Pdgfrα-CKO embryos to human ARMs, suggesting that dysregulation in the time and level of Pdgfrα expression could cause these anomalies in human.