Testosterone inhibits adipogenic differentiation in 3T3-L1 cells: nuclear translocation of androgen receptor complex with beta-catenin and T-cell factor 4 may bypass canonical Wnt signaling to down-regulate adipogenic transcription factors.
Testosterone supplementation in men decreases fat mass; however, the mechanisms by which it inhibits fat mass are unknown. We hypothesized that testosterone inhibits adipogenic differentiation of preadipocytes by activation of androgen receptor (AR)/beta-catenin interaction and subsequent translocation of this complex to the nucleus thereby bypassing canonical Wnt signaling. We tested this hypothesis in 3T3-L1 cells that differentiate to form fat cells in adipogenic medium. We found that these cells express AR and that testosterone and dihydrotestosterone dose-dependently inhibited adipogenic differentiation as analyzed by Oil Red O staining and down-regulation of CCAAT/enhancer binding protein-alpha and -delta and peroxisome proliferator-activated receptor-gamma2 protein and mRNA. These inhibitory effects of androgens were partially blocked by flutamide or bicalutamide. Androgen treatment was associated with nuclear translocation of beta-catenin and AR. Immunoprecipitation studies demonstrated association of beta-catenin with AR and T-cell factor 4 (TCF4) in the presence of androgens. Transfection of TCF4 cDNA inhibited adipogenic differentiation, whereas a dominant negative TCF4 cDNA construct induced adipogenesis and blocked testosterone's inhibitory effects. Our gene array analysis indicates that testosterone treatment led to activation of some Wnt target genes. Expression of constitutively activated AR fused with VP-16 did not inhibit the expression of CCAAT/enhancer binding protein-alpha in the absence of androgens. Testosterone and dihydrotestosterone inhibit adipocyte differentiation in vitro through an AR-mediated nuclear translocation of beta-catenin and activation of downstream Wnt signaling. These data provide evidence for a regulatory role for androgens in inhibiting adipogenic differentiation and a mechanistic explanation consistent with the observed reduction in fat mass in men treated with androgens.
Singh
Rajan
R
Division of Endocrinology, Metabolism, and Molecular Medicine, Charles R. Drew School of Medicine, Los Angeles, California 90059, USA. rajansingh@mednet.ucla.edu
Artaza
Jorge N
JN
Taylor
Wayne E
WE
Braga
Melissa
M
Yuan
Xin
X
Gonzalez-Cadavid
Nestor F
NF
Bhasin
Shalender
S
D016475:3T3 Cells; D017667:Adipocytes / Q000166:cytology* / Q000187:drug effects; D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D051379:Mice; D021381:Protein Transport / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics; D011944:Receptors, Androgen / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology; D051820:TCF Transcription Factors / Q000378:metabolism*; D013739:Testosterone / Q000494:pharmacology*; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects; D051153:Wnt Proteins / Q000235:genetics / Q000502:physiology*; D051176:beta Catenin / Q000378:metabolism*
D012333:RNA, Messenger; D011944:Receptors, Androgen; D051820:TCF Transcription Factors; C544895:Tcf7l2 protein, mouse; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors; D051153:Wnt Proteins; D051176:beta Catenin; D013739:Testosterone