Reports of adverse human pregnancy outcomes including cleft palate have increased as the clinical use of isotretinoin (13-cis-retinoic acid) and other retinoic acid (RA) derivatives have increased, but the mechanisms by which their effects are exerted are not understood. Research in craniofacial development is generally performed in rodents, and mouse palatal shelves exposed in organ cultures to retinoids and epidermal growth factor (EGF) display altered medial epithelial cell morphology blocking normal union of apposing shelves. In the present study, precontacting human palatal shelves were maintained in organ culture for 2, 3, or 6 days and exposed to labeled thymidine (3H-TdR) during the last 16 hr. Retinoids and EGF were included in the media so that each shelf was exposed to one of the following: control, EGF, trans-RA at 10(-5)M, cis-RA at 10(-7) or 10(-9) M, or RA + EGF. After exposure of cultured human embryonic palatal shelves to 13-cis-RA and trans-RA with or without EGF, medial epithelial cells do not degenerate, cell surface morphology shifts toward a nasal type, glycogen deposits decrease, smooth endoplasmic reticulum (SER) increases, and basal lamina appear altered. In shelves exposed to EGF and trans-RA early in their development, DNA synthesis appears to terminate prematurely as compared to shelves cultured in control media, and this effect is accompanied by excessive mesenchymal extracellular space expansion. Exposure of shelves to EGF alone is sufficient to block degeneration and induce hyperplasia of the medial epithelial cells but does not induce other ultrastructural changes seen with both EGF and RA. The observed alterations in medial cell morphology could interfere with adhesion of the palatal shelves and may play a role in retinoid-induced cleft palate in the human embryo.