N-TERMINAL 10 AND 12-KDA POMC FRAGMENTS STIMULATE DIFFERENTIATION OF LACTOTROPHS

Medium conditioned by a highly enriched population of gonadotrophs, cu ltured as reaggregates in the presence of 0.01 nM GnRH, was concentrat ed, separated on a reversed-phase HPLC column, and tested for activity on lactotroph development in pituitary reaggregate cell cultures of 1 4-day-old rats. The number of cells expressing prolactin (PRL) mRNA wa s estimated by image analysis after in situ hybridization of paraffin- embedded sections. The number of these cells entering the mitotic cycl e was estimated by autoradiography of [H-3]thymidine ([H-3]T) incorpor ation. One HPLC column fraction expanded the section area occupied by PRL. mRNA cells without displaying an effect on [H-3]T labeling of the se cells, indicating that this fraction induces differentiation in the lactotroph Lineage. The latter fraction was further purified on a sec ond reversed-phase HPLC column, a gel filtration column, and a final r eversed-phase HPLC column. From the last column, four substances were isolated that all selectively induced differentiation of lactotrophs. Each of them had an N-terminal amino acid sequence identical to the N- terminal domain of rat proopiomelanocortin (POMC). As determined by ma ss spectrometric analysis, the M(r)s were 10,091, 10,289, 12,238, and 12,247 Da, respectively. The C-terminal extension of these compounds i s possibly up to Gln(74) for the former two compounds and up to Gly(95 ) for the latter two. Authentic purified human POMC(1-76) mimicked the effects of the purified 10- and 12-kDa rat POMC fragments. The presen t data suggest that certain isoforms of rat POMC(1-74) and human POMC( 1-76) can stimulate lactotroph growth through a differentiation-induci ng action on progenitor cells. Copyright (C) 1996 Elsevier Science Inc .