Induction of circadian gene expression in human subcutaneous adipose-derived stem cells.
OBJECTIVE
Genes encoding the circadian transcriptional apparatus exhibit robust oscillatory expression in murine adipose tissues. This study tests the hypothesis that human subcutaneous adipose-derived stem cells (ASCs) provide an in vitro model in which to monitor the activity of the core circadian transcriptional apparatus.


METHODS
Primary cultures of undifferentiated or adipocyte-differentiated ASCs were treated with dexamethasone, rosiglitazone, or 30% fetal bovine serum. The response of undifferentiated ASCs to dexamethasone was further evaluated in the presence of lithium chloride. Lithium inhibits glycogen synthase kinase 3, a key component of the circadian apparatus. Total RNA was harvested at 4-hour intervals over 48 hours and examined by real-time reverse transcription polymerase chain reaction (RT-PCR).


RESULTS
Adipocyte-differentiated cells responded more rapidly to treatments than their donor-matched undifferentiated controls; however, the period of the circadian gene oscillation was longer in the adipocyte-differentiated cells. Dexamethasone generated circadian gene expression patterns with mean periods of 25.4 and 26.7 hours in undifferentiated and adipocyte-differentiated ASCs, respectively. Both rosiglitazone and serum shock generated a significantly longer period in adipocyte-differentiated ASCs relative to undifferentiated ASCs. The Bmal1 profile was phase-shifted by approximately 8 to 12 hours relative to Per1, Per3, and Cry2, consistent with their expression in vivo. Lithium chloride inhibited adipogenesis and significantly lengthened the period of Per3 and Rev-erbalpha gene expression profiles by >5 hours in dexamethasone-activated undifferentiated ASCs.


CONCLUSIONS
These results support the initial hypothesis and validate ASCs as an in vitro model for the analysis of circadian biology in human adipose tissue.
Wu
Xiying
X
Stem Cell Laboratory, Pennington Biomedical Research Center, Baton Rouge, Louisiana 70808, USA.
Zvonic
Sanjin
S
Floyd
Z Elizabeth
ZE
Kilroy
Gail
G
Goh
Brian C
BC
Hernandez
Teri L
TL
Eckel
Robert H
RH
Mynatt
Randall L
RL
Gimble
Jeffrey M
JM
D056930:ARNTL Transcription Factors; D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D050156:Adipogenesis / Q000235:genetics; D000328:Adult; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D002940:Circadian Rhythm / Q000235:genetics*; D056931:Cryptochromes; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D005260:Female; D005420:Flavoproteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors; D006801:Humans; D018021:Lithium Chloride / Q000494:pharmacology; D008875:Middle Aged; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D056953:Nuclear Receptor Subfamily 1, Group D, Member 1; D056950:Period Circadian Proteins; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D000077154:Rosiglitazone; D013234:Stem Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D045162:Thiazolidinediones / Q000494:pharmacology; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
D056930:ARNTL Transcription Factors; D051792:Basic Helix-Loop-Helix Proteins; D018797:Cell Cycle Proteins; D056931:Cryptochromes; D004268:DNA-Binding Proteins; D003907:Dexamethasone; D005420:Flavoproteins; D038362:Glycogen Synthase Kinase 3; D018021:Lithium Chloride; D009687:Nuclear Proteins; D056953:Nuclear Receptor Subfamily 1, Group D, Member 1; D056950:Period Circadian Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D012097:Repressor Proteins; D000077154:Rosiglitazone; D045162:Thiazolidinediones; D014157:Transcription Factors; C538960:BMAL1 protein, human; C538966:CRY2 protein, human; C064353:NR1D1 protein, human; C090977:NR1D2 protein, human; C539022:PER1 protein, human; C512644:PER3 protein, human