Mek and p38 MAPK-dependant pathways are involoved in the positive effect of interleukin-6 on human growth hormone gene expression in rat MtT/S somatotroph cells.
OBJECTIVE
To investigate the effect of interleukin-6 (IL-6) on the human growth hormone (hGH) gene expression in a rat somatotropic pituitary cell line MtT/S.


METHODS
The plasmids containing various lengths of hGH gene 5'-promoter fragments were constructed. Stably transfected MtT/S cells were created by cotransfecting the above plasmids and pcDNA3. 1(+) with DMRIE-C transfection reagent After the administration of these cells with IL-6 and/or various inhibitors of signaling transduction pathways, the luciferase activities in MtT/S cells lysis were assayed to demonstrate the effects of IL-6 on hGH gene promoter activity and possibly involved mechanism.


RESULTS
The 10(3) U/mL IL-6 stimulated GH secretion and synthesis, and promoted the 5'-promoter activity of GH gene in stably transfected MtT/SGL cells with the action of 1.69 times above the control. Among inhibitors of signaling transduction pathways, mitogen-activated protein kinase kinase (MAPKK/MEK) inhibitor PD98059 (40 micromol/L) and p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 (5 micromol/L) completely blocked the stimulatory effect of IL-6. Western blot analysis further confirmed the activation of phosphorylated MEK and p38 MAPK in MtT/SGL cells. Neither over-expression of Pit-1 nor inhibition of Pit-1 expression affected IL-6 induction of hGH promoter activity. A series of deletion constructs of hGH promoter were created to identify the DNA sequence that mediated the effect of IL-6. The results showed that the stimulatory effect of IL-6 was abolished following deletion of the -196 to - 132 bp fragment.


CONCLUSIONS
IL-6 promotes GH secretion and synthesis by rat MtT/S somatotroph cells. The stimulatory effect of IL-6 on hGH gene promoter appears to require the activation of MEK and p38 MAPK, and a fragment of promoter sequence that spans the - 196 to - 132 bp of the gene, but may be unlinked with Pit-1 protein.
Gong
Feng-Ying
FY
Department of Endocrinology, Key Laboratory of Endocrinology of Ministry of Health, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730. fygong5074@yahoo.com.cn
Deng
Jie-Ying
JY
Shi
Yi-Fan
YF
D000818:Animals; D002460:Cell Line; D005786:Gene Expression Regulation; D019382:Human Growth Hormone / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism*; D048031:JNK Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000502:physiology*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D051381:Rats; D052683:Somatotrophs / Q000166:cytology / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*
D015850:Interleukin-6; D019382:Human Growth Hormone; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020929:Mitogen-Activated Protein Kinase Kinases