Conditional heterozygous loss of Kit receptor tyrosine kinase in neural crest cell lineage is associated with midline cleft lip and bifid nose deformity.

Abstract

Objectives: The receptor tyrosine kinase Kit is expressed in cells derived from the trunk neural crest (NC), such as melanocytes; however, its role in cranial NC cell development is not fully understood.

Methods: We investigated the effects of the heterozygous loss of Kit in NC cells during embryonic development by mating Kit^2lox/+ mice with Wnt1-Cre mice to produce Wnt1-Cre; Kit^2lox/+ embryos. In addition, Wnt1-Cre mice were mated with Rosa26R-yellow fluorescent protein (YFP) mice to visualize the tissue regions expressing Cre recombinase. Histological studies of the craniofacial regions of these mice were performed using samples from embryonic day (E) 12.5 and postnatal day (P) 1. Cellular apoptosis and proliferation were both analyzed through the immunostaining of tissue sections collected on E13.5 and E14.5 using anti-cleaved caspase 3 (CC3) to detect apoptosis and anti-Ki67 to detect proliferation. Cells from YFP-positive tissue regions of the facial areas of Wnt1-Cre; Kit^+/+; Rosa26R-YFP embryos and Wnt1-Cre; Kit^2lox/+; Rosa26R-YFP embryos collected on E12.5 and E15.5 were cultured and evaluated for cell proliferation.

Results: Compared with control littermates, Wnt1-Cre; Kit^2lox/+ embryos exhibited midline cleft lip and bifid nose deformities. Substantial early (P1) postnatal lethality was observed in Wnt1-Cre; Kit^2lox/+ mice, with none surviving to 3 weeks of age. YFP-positive cells from the maxillary regions of Wnt1-Cre; Kit^2lox/+; Rosa26R-YFP embryos exhibited defective cell growth and self-renewal in vitro.

Conclusion: Conditional heterozygous loss of Kit in Wnt1-Cre; Kit^2lox/+ embryos is associated with craniofacial dysplasia and exhibit defective NC development in vitro and in vivo.