EFFECT OF POMC1-76, ITS C-TERMINAL FRAGMENT GAMMA-3-MSH AND ANTI-POMC1-76 ANTIBODIES ON DNA-REPLICATION IN LACTOTROPHS IN AGGREGATE CELL-CULTURES OF IMMATURE RAT PITUITARY

Treatment of aggregate cell cultures of 14-day-old rat pituitary for 4 0 h with purified human (h) POMC1-76 dose-dependently augmented the nu mber of DNA replicating lactotrophs as estimated by autoradiography of [H-3]-thymidine (H-3-T) incorporation in cells immunostained for prol actin (PRL). No such effect was seen on the total number of H-3-T labe lled cells (the majority of which did not contain any pituitary hormon e in a detectable amount) or on the total number of lactotrophs. The e ffect of hPOMC(1-76) on H-3-T incorporation in lactotrophs was blocked by concomitant treatment with anti-hPOMC(1-76) monoclonal and polyclo nal antibodies cross-reactive with rat POMC1-74. The latter anti-hPOMC (1-76) antibodies also decreased the number of H-3-T incorporating lac totrophs in the absence of hPOMC(1-76). gamma 3-MSH, which is the C-te rminal domain of hPOMC(1-76), mimicked the effect or hPOMC(1-76) on H- 3-T incorporation in lactotrophs but its potency was lower than that o f hPOMG(1-76). Other melanocortin (MC) peptides such as alpha- and bet a-MSH were also effective but were less potent than gamma 3-MSH. The d ifference in potency was not due to partial degradation of the peptide s. hPOMC(1-76) did not affect H-3-T incorporation in other pituitary c ell types. In contrast gamma(3)-MSH also augmented the number of H-3-T labelled somatotrophs and thyrotrophs. In the embryonic kidney 293 ce ll line stably transfected with the MC-3 receptor, gamma 3-MSH (10 nM) augmented cAMP formation up to 30 times. In contrast, hPOMC(1-76) (10 0 nM) was inactive in this test system,' indicating this peptide is no t an agonist at the MC-3 receptor. The present investigation further s upports the role of rat POMC1-74 as a paracrine growth factor in the d evelopment of lactotrophs. The active core of POMC1-76 does not seem t o be restricted to its C-terminal domain gamma 3-MSH as the latter pep tide displays a growth promoting effect that is different from that of POMC1-76 it is less potent, it is not specific for lactotrophs and wh ereas the effect of gamma 3-MSH may be mediated by the MC-3 receptor t hat of POMC1-76 is not.