Roles of transforming growth factor-alpha and epidermal growth factor in chick limb development.

We have examined the distribution of transforming growth factor-alpha (TGF-alpha), epidermal growth factor (EGF), and the chicken EGF receptor (c-erbB), in embryonic chick limbs. Prior to limb budding, TGF-alpha is present in prospective limb-forming mesoderm and in prospective apical ectodermal ridge (AER)-forming ectoderm, but is not detected in non-limb-forming flank mesoderm or ectoderm, nor in presumptive non-AER-forming limb ectoderm, suggesting possible roles in initial limb formation and AER induction. Consistent with this possibility, TGF-alpha is present in the mesoderm of the wing buds of the amelic chick mutants limbless and wingless, which form and bud normally, but is absent from limbless and wingless ectoderm, which fails to form an AER. TGF-alpha and EGF are present in the AER of the developing limb, and TGF-alpha, EGF, and c-erbB are present in the underlying subridge mesoderm, suggesting possible roles in reciprocal AER/subridge mesoderm interactions required for limb outgrowth. We found that exogenous TGF-alpha and EGF can promote the outgrowth of limb mesoderm in the absence of the AER in vitro and can also promote the outgrowth of limbless and wingless wing bud explants. EGF is present in ventral but not dorsal limb ectoderm, suggesting a role for EGF in specification of ventral ectoderm. TGF-alpha and EGF are not detected in the differentiating cartilaginous elements or muscle primordia of the limb, suggesting that cessation of TGF-alpha and EGF expression may be required for cartilage and muscle formation. We have found that exogenous TGF-alpha and EGF inhibit chondrogenesis and myogenesis of limb mesenchyme in vitro. Together these results indicate that signaling through the EGF receptor via endogenous TGF-alpha and EGF may be important for initial limb formation, AER induction, outgrowth of limb mesoderm, and regulation of limb chondrogenic and myogenic differentiation.