ACTH and alpha-MSH inhibit leptin expression and secretion in 3T3-L1 adipocytes: model for a central-peripheral melanocortin-leptin pathway.
Leptin is the 167 amino-acid protein product of the Lep (obese) gene that is released predominantly from adipose tissue and circulates at levels related to the amount of fat. Leptin expression is hormonally regulated: insulin and glucocorticoids are stimulators, while inhibitors include beta-adrenergic agonists and testosterone. Recently, adenylate cyclase-coupled melanocortin receptors have been identified in murine adipose tissue, the 3T3-L1 adipocyte cell line, and in human fat tissue. These studies prompted us to evaluate the effects of pro-opiomelanocortin (POMC)-derived peptides on leptin production and expression in 3T3-L1 adipocytes in culture. 3T3-L1 pre-adipocytes differentiated by the insulin/indomethacin (I/I) method produced leptin at levels that were two times higher than those obtained in cells differentiated by the more traditional insulin/dexamethasone/isobutylmethylxanthine (I/D/M) method. By RT-PCR studies, 3T3-L1 cells expressed both the melanocortin 2 receptors (MC2-R) and melanocortin 5 receptors (MC5-R) isoforms of the melanocortin receptor at an early stage of differentiation. When I/I differentiated 3T3-L1 adipocytes were incubated with different concentrations of dibutyryl cAMP (db-cAMP) or POMC-derived peptides (ACTH and alpha-MSH), ACTH and alpha-MSH stimulated cAMP production after 30 min (2-fold increase) associated with a dose-dependent inhibition of leptin secretion (ACTHz.Gt;alpha-MSH; IC(50)=3.2+/-0.4 SE and 36+/-5 nM, respectively), maximal after 3 h of incubation (30% inhibition). In addition, 100 nM ACTH and alpha-MSH induced a 60% reduction in leptin expression by RT-PCR. Incubation of cells with 0.5 mM db-cAMP led to a more prominent inhibition of leptin expression and secretion (up to 80% at 1 and 24 h, respectively). The ACTH and alpha-MSH inhibitory effects on leptin secretion were mediated by activation of the MC2-R and MC5-R and were reversed by the MC-R antagonists ACTH(11-24) and ACTH(7-38). In summary, we have shown that POMC-peptides are potent inhibitors of leptin expression and production in 3T3-L1 adipocytes. The finding of ACTH/alpha-MSH receptor-induced inhibition of leptin production and expression in adipocytes support the possibility that there is a control mechanism for modulation of adipose tissue function via a melanocortin-leptin axis.
Norman
Dennis
D
Department of Endocrinology, St. Bartholomew's Hospital, London EC1A 7BE, UK.
Isidori
Andrea M
AM
Frajese
Vanni
V
Caprio
Massimiliano
M
Chew
Shern L
SL
Grossman
Ashley B
AB
Clark
Adrian J
AJ
Michael Besser
G
G
Fabbri
Andrea
A
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000378:metabolism*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D019070:Cell Lineage; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D050536:Glycerol-3-Phosphate Dehydrogenase (NAD+); D005993:Glycerolphosphate Dehydrogenase / Q000378:metabolism; D006801:Humans; D020738:Leptin / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D010446:Peptide Fragments / Q000378:metabolism; D011333:Pro-Opiomelanocortin / Q000378:metabolism; D044102:Receptor, Melanocortin, Type 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D054411:Receptors, Leptin; D000521:alpha-MSH / Q000494:pharmacology*
C515448:LEPR protein, human; D020738:Leptin; D010446:Peptide Fragments; D044102:Receptor, Melanocortin, Type 1; D054411:Receptors, Leptin; C515447:leptin receptor, mouse; D000521:alpha-MSH; D011333:Pro-Opiomelanocortin; D000324:Adrenocorticotropic Hormone; D000242:Cyclic AMP; D005993:Glycerolphosphate Dehydrogenase; D050536:Glycerol-3-Phosphate Dehydrogenase (NAD+)