The augmentation of insulin-like growth factor-I messenger ribonucleic acid in cultured rat hepatocytes: activation of protein kinase-A and -C is necessary, but not sufficient.
In previous studies it was shown that bovine GH (bGH) and glucagon, when individually added to primary rat hepatocyte cultures, modestly stimulated IGF-I mRNA levels 1.8- to 2.5-fold, but when combined, synergized to stimulate IGF-I mRNA levels by 10- to 12-fold. In the present study we have explored further the mechanism of this effect in primary rat hepatocyte cultures. Like glucagon, the addition of 3-isobutyl-1-methylxanthine (100 microM) or (Bu)2cAMP (150 microM) augmented IGF-I mRNA levels 1.8- to 2.0-fold, but when combined with bGH (50 ng/ml), they augmented levels up to 12-fold. The half-life of IGF-I mRNA, determined by incubating hepatocytes with actinomycin-D was 12 h. Although bGH did not affect the decay rate, glucagon (100 ng/ml) or (Bu)2cAMP (100 microM) reduced the rate of loss by about 70%. 4 beta-Phorbol 12 beta-myristate 13 alpha-acetate minimally stimulated IGF-I mRNA levels 1.2- to 1.4-fold, but displayed no synergism when added with bGH, glucagon, or (Bu)2cAMP. The stimulatory effect of bGH plus glucagon was inhibited 80% after preincubation with 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (10 microM) for 24 h. The addition of staurosporine, sphingosine, or H-7 [1-(5-isoquinolinyl sulfonyl)2-methyl piperazine] inhibited the stimulatory effect of bGH plus glucagon on hepatocyte IGF-I mRNA by 80%, 90%, and 85%, respectively. Preincubation with cycloheximide (10 micrograms/ml) blocked the synergistic effect of bGH plus either glucagon or (Bu)2cAMP by 65-80%. The effect of glucagon, mediated via the activation of adenylate cyclase, involves in part the posttranscriptional stabilization of IGF-I mRNA levels. The effect of GH, mediated in part by the activation of protein kinase-C, appears to be at the level of transcription. The synergistic augmentation of hepatocyte IGF-I mRNA levels by GH and glucagon involves the activation of PKA and PKC, but also appears to require the synthesis of one or more protein(s).
Kachra
Z
Z
Polypeptide Hormone Laboratory, Royal Victoria Hospital, Montreal, Quebec, Canada.
Yang
C R
CR
Posner
B I
BI
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D015152:Blotting, Northern; D003994:Bucladesine / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D004357:Drug Synergism; D004789:Enzyme Activation; D005934:Glucagon / Q000494:pharmacology; D013006:Growth Hormone / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis*; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013110:Sphingosine / Q000494:pharmacology; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000470:Alkaloids; D007546:Isoquinolines; D010879:Piperazines; D012333:RNA, Messenger; D003994:Bucladesine; D007334:Insulin-Like Growth Factor I; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D013006:Growth Hormone; D005934:Glucagon; D003513:Cycloheximide; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D019311:Staurosporine; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; D015056:1-Methyl-3-isobutylxanthine