Effects of tyrosine kinase inhibitors on tyrosine phosphorylations and the insulin-like effects in response to human growth hormone in isolated rat adipocytes.
Recent data suggest involvement of the Janus tyrosine kinase-2 (JAK2) in human GH-induced tyrosine phosphorylation of the GH receptor and the insulin receptor substrates 1 and 2 (IRS-1 and IRS-2), leading to activation of the phosphatidylinositol 3-kinase and the acute insulin-like effects in primary rat adipocytes. To investigate the functional role of this kinase, we screened a number of tyrosine kinase inhibitors for their ability to inhibit three rapid effects of GH on primary adipocytes: increased lipogenesis, inhibition of noradrenaline-induced lipolysis, and promotion of JAK2 tyrosine phosphorylation. Only staurosporine was found to inhibit all three effects. The inhibition of lipogenesis and antilipolysis exhibited the same staurosporine dose dependency (IC50, approximately 40 nM) as inhibition of JAK2 and IRS-1 tyrosine phosphorylation as well as binding of the p85 subunit of phosphatidylinositol 3-kinase to IRS-1 and IRS-2. The unidentified cytosolic tyrosine-phosphorylated protein pp95, in contrast, was not affected, suggesting that it is not phosphorylated primarily by JAK2. Protein kinase C does not seem to be directly involved in the insulin-like effects, because the selective protein kinase C inhibitor calphostin C had no effect at levels up to 100 nM above which unspecific cellular effects occurred. Methyl-2,5-dihydroxy cinnamate inhibited GH-induced lipogenesis from [3-3H]glucose and nonstimulated lipogenesis from [2-14C]-pyruvate and [3H]acetate, but was without effect on GH-induced 2-deoxy-D-[1-3H]glucose uptake, JAK2 phosphorylation and antilipolysis, suggesting unspecific effects on mitochondrial metabolism rather than a direct effect on the GH-mediated signal. Tyrphostin 25 and herbimycin A had no effect on any of the parameters studied, except for a slight increase in JAK2 phosphorylation in response to tyrphostin 25. In summary, these data support the role for JAK2 in mediating the insulin-like effects of GH in adipocytes.
Department of Pediatrics, University of Lund, Sweden.
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Ridderstråle
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Tornqvist
D000085:Acetates / Q000378:metabolism; D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D016227:Benzoquinones; D002478:Cells, Cultured; D002934:Cinnamates / Q000494:pharmacology; D003847:Deoxyglucose / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004822:Epididymis; D019382:Human Growth Hormone / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D047029:Lactams, Macrocyclic; D008055:Lipids / Q000096:biosynthesis; D008066:Lipolysis / Q000187:drug effects; D008297:Male; D009281:Naphthalenes / Q000494:pharmacology; D009570:Nitriles / Q000494:pharmacology; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019289:Pyruvic Acid / Q000378:metabolism; D011809:Quinones / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D019311:Staurosporine / Q000494:pharmacology; D020032:Tyrphostins
D000085:Acetates; D016227:Benzoquinones; D002934:Cinnamates; D004791:Enzyme Inhibitors; D007328:Insulin; D047029:Lactams, Macrocyclic; D008055:Lipids; D009281:Naphthalenes; D009570:Nitriles; D011809:Quinones; D011994:Recombinant Proteins; D020032:Tyrphostins; C079836:tyrphostin 25; D019382:Human Growth Hormone; C029010:cinnamic acid; D017828:Rifabutin; D019000:Phosphotyrosine; C020754:herbimycin; D019289:Pyruvic Acid; D003847:Deoxyglucose; D011505:Protein-Tyrosine Kinases; D019311:Staurosporine; C058819:calphostin C