MESENCHYMAL CHANGES ASSOCIATED WITH RETINOIC ACID-INDUCED CLEFT-PALATE IN CD-1 MICE

Retinoic acid (RA) is teratogenic in many species and is an effective inducer of cleft palate in mice. The pathogenesis of cleft formation v aries with the timing of exposure. It has been demonstrated, before fo rmation of the palatal shelves, that RA exposure results in insufficie nt mesenchymal tissue, and palatal shelves fail to make contact. Howev er, at the palatal shelf outgrowth stage, RA exposure affects shelf el evation and growth in rats, and possibly medial edge epithelium (MEE) differentiation in mice. The objective of this study was to examine th e morphologic and functional changes associated with cleft formation i n mice following exposure during shelf outgrowth. Particular emphasis was placed on evaluating the timing of palatal shelf elevation in RA e xposed embryos and on identifying differentiation events occurring con currently in the epithelium. On gestational day (GD) 12 (8:00 AM), gra vid CD-1 mice were gavaged with 70 mg/kg RA or vehicle. This protocol produced a 100% incidence of cleft palate at term, allowing us to corr elate the morphological and/or biochemical changes observed at pre-fus ion time points. Embryos were collected at 12 hr intervals through GD 15, beginning 4 hr after exposure. Serial sections of embryos were eit her stained with H&E, with a battery of lectins [Sambucus nigra (SNA), Arachis hypogaea (PNA), Ricinus communis (RCA-I), Glycine mas (SBA), Succinylated Wheat Germ (S-WGA)], or with a probe to hyaluronan. Throu ghout the period of normal palate development, the shelf mesenchyme sh owed increasing regional organization and progressive hydration and th ese changes were correlated with increase Hyaluronan (HA) deposition. RA treatment resulted in lose of regional organization and delayed mes enchyme hydration. In association with these changes there were reduct ions in HA deposition and extracellular matrix glycoconjugates recogni zed by PNA in the palate mesenchyme. Further there was a considerable delay in palatal shelf elevation and palate shelf did not make contact at the midline. Our data indicates, in embryos exposed on GD 12 to le vels of RA sufficient to induce a 100% incidence of clefting, that cle ft formation is a result of palatal shelves failing to make contact. A lterations in mesenchyme development and the subsequent delay in palat e shelve elevation are central to RA-induced cleft formation following exposure at the palate shelf out growth stage.