Impact of retinoic acid exposure on midfacial shape variation and manifestation of holoprosencephaly in Twisted gastrulation mutant mice

Abstract

Abstract Holoprosencephaly (HPE) is a developmental anomaly characterized by inadequate or absent midline division of the embryonic forebrain and midline facial defects. It is believed that gene-environment interactions play a role in the widely variable penetrance and expressivity of HPE, although a direct investigation of such effects has been limited. The goal of this study was to examine if mice carrying a mutation in a gene encoding a BMP antagonist Twisted gastrulation (Twsg1) associated with a low penetrance of HPE are sensitized to retinoic acid (RA) teratogenesis. Pregnant Twsg1+/- dams were treated by gavage with a low dose of all-trans RA (3.75 mg/kg). Embryos were analyzed between E9.5 and E11.5 by microscopy and geometric morphometric analysis by microCT. P19 embryonal carcinoma cells were used to examine potential mechanisms mediating combined effects of increased BMP and retinoid signaling. While only 7% of wild type embryos exposed to RA showed overt HPE or neural tube defects (NTD), 100% of Twsg1 null mutants exposed to RA manifested severe HPE compared to 17% without RA. Remarkably, up to 30% of Twsg1+/- mutants also showed HPE (23%) or NTD (7%). The majority of shape variation among Twsg1+/- mutants was associated with narrowing of the midface. In P19 cells, RA induced the expression of Bmp2, acted in concert with BMP to increase p53 expression, caspase activation, and oxidative stress. This study provides direct evidence for modifying effects of the environment in a genetic mouse model carrying a predisposing mutation for HPE in the Twsg1 gene. Further study of the mechanisms underlying these gene-environment interactions in vivo will contribute to better understanding of the pathogenesis of birth defects and present an opportunity to explore potential preventive interventions.