TSH signaling and cell survival in 3T3-L1 preadipocytes.
Thyroid-stimulating hormone (TSH) action in adipose tissue remains largely unknown. Our previous work indicates that human preadipocytes express functional TSH receptor (TSHR) protein, demonstrated by TSH activation of p70 S6 kinase (p70 S6K). We have now studied murine 3T3-L1 preadipocytes to further characterize TSH signaling and cellular action. Western blot analysis of 3T3-L1 preadipocyte lysate revealed the 100-kDa mature processed form of TSHR. TSH activated p70 S6K and protein kinase B (PKB/Akt), as measured by immunoblot analysis. Preincubation with wortmannin or LY-294002 completely blocked TSH activation of p70 S6K and PKB/Akt, implicating phosphoinositide 3-kinase (PI3K) in their regulation. TSH increased phosphotyrosine protein(s) in the 125-kDa region and augmented the associated PI3K activity fourfold. TSH had no effect on cAMP levels in 3T3-L1 preadipocytes, suggesting that adenylyl cyclase is not involved in TSH activation of the PI3K-PKB/Akt-p70 S6K pathway. 3T3-L1 preadipocyte cell death was reduced by 29-76% in serum-deprived (6 h) preadipocytes treated with 1-20 microM TSH. In the presence of 20 microM TSH, an 88% reduction in terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL)-positive cells was observed in serum-starved (3 h) 3T3-L1 preadipocytes as well as a 93% reduction in the level of cleaved activated caspase 3. In summary, TSH acts as a survival factor in 3T3-L1 preadipocytes. TSH does not stimulate cAMP accumulation in these cells but instead activates a PI3K-PKB/Akt-p70 S6K pathway.
Bell
Andrea
A
Department of Biochemistry, Microbiology, and Immunology, Ottawa Health Research Institute, University of Ottawa, Ottawa, Ontario, K1Y 4E9, Canada.
Gagnon
Annemarie
A
Dods
Patti
P
Papineau
Denise
D
Tiberi
Mario
M
Sorisky
Alexander
A
D016475:3T3 Cells; D017667:Adipocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002458:Cell Fractionation; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D000242:Cyclic AMP / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020287:In Situ Nick-End Labeling; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger / Q000096:biosynthesis; D011989:Receptors, Thyrotropin / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D019893:Ribosomal Protein S6 Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D013972:Thyrotropin / Q000494:pharmacology*
D000242:Cyclic AMP; D004791:Enzyme Inhibitors; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D012333:RNA, Messenger; D011989:Receptors, Thyrotropin; D019893:Ribosomal Protein S6 Kinases; D013972:Thyrotropin