GH induced lipolysis stimulation in 3T3-L1 adipocytes stably expressing hGHR: analysis on signaling pathway and activity of 20K hGH

We have constructed a cell line of 3T3-L1 which can efficiently express hum an GHR (3T3-L1-hGHR) after differentiation to adipocytes. The expressed hGH R was detected as two bands with approximate molecular sizes of 120K by Wes tern analysis using hGHR specific monoclonal antibody. Maximum lipolytic ac tivity induced by hGH in the 3T3-L1-hGHR was enhanced l0-fold as compared t o that in 3T3-L1, suggesting that expressed hGHR is functionally active. Co mparative analysis using bGH and hGH revealed that 70% of lipolysis stimula tion by 1-10 ng/ml hGH could be attributed to hGHR-mediated response. Analy ses on inhibition and phosphorylation of signaling molecules suggested that GH-induced lipolysis stimulation is dependent on gene expression and not m ediated through PKA-, PKC-, PLA-, PLC-, nor MAPK-pathway but possibly throu gh JAK-STATs pathway. Duration of STAT5 activation by hGH continued up to 4 8 h. We also revealed that 22 K hGH isoform, 20K hGH which has been reporte d as a weaker agonist for GH-induced lipolysis stimulation, possesses equip otent activity and shows stronger action in the presence of hGHBP as compar ed to 22 K hGH. Taken together we conclude that the hGH-induced lipolysis w as not mediated through MAP-. PKA-, PKC-, nor PLA-pathway but might be medi ated through STAT pathway and that 20K hGH might show higher lipolytic acti vity than 22 K hGH in adipose tissue that produces a large amount of GHBP. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.