[Effects of ghrelin on the proliferation and differentiation of 3T3-L1 preadipocytes and its possible mechanisms].
OBJECTIVE
To investigate the effects of ghrelin on the proliferation and differentiation of 3T3-L1 preadipocyte, and study the possible mechanisms.


METHODS
3T3-L1 preadipocytes were cultured in vitro. The proliferation potentials of 3T3-L1 preadipocytes that were treated with different concentrations of ghrelin were evaluated by MTT methods. The levels of c-myc and thymidine kinase mRNA were detected using RT-PCR. 3T3-L1 preadipocytes were differentiated into the matured adipocytes with insulin (INS) or ghrelin. The morphological changes of 3T3-L1 adipocytes were observed and the differentiation rate was assayed by oil-red O staining. Total RNA was extracted from adipocytes at various times, and the levels of peroxisome proliferation activated receptor gamma (PPARgamma) and CAAT/enhancer binding protein(C/EBPalpha) mRNA expressions were detected using RT-PCR.


RESULTS
Ghrelin at concentrations of 10(-7) to 10(-15) mol/L significantly stimulated preadipocyte proliferation (p<0.05). The levels of c-myc and thymidine kinase mRNA significantly increased in 3T3-L1 preadipocytes with 10(-9) mol/L and 10(-11) mol/L ghrelin treatment (p<0.01). The 3T3-L1 preadipocytes treated with 10(-11) mol/L ghrelin had lots of lipid droplets in the cytoplasma, but the differentiation rate was lower than those treated with INS. Ghrelin of 10(-11) mol/L significantly increased the mRNA expression of PPARgamma and C/EBPalpha in the course of 3T3-L1 preadipocyte differentiation, compared with the normal control group (p<0.05). The PPARgamma and C/EBPalpha mRNA expression increased with the prolonged differentiation of preadipocytes induced by ghrelin or INS. There were significant differences in the levels of PPARgamma and C/EBPalpha mRNA expression between the 2nd and 8th days of differentiation(p<0.01).


CONCLUSIONS
Ghrelin promotes the proliferation and differentiation of 3T3-L1 preadipocytes. The proliferation of 3T3-L1 preadipocytes induced by ghrelin may be associated with increased c-myc levels. Ghrelin may promote differentiation of 3T3-L1 preadipocytes by increasing mRNA expression of PPARgamma and C/EBPalpha, thus enhances the sensitivity of adipocytes to INS.
Liu
Jing
J
Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Lin
Han-Hua
HH
Cheng
Pei-Xuan
PX
Hu
Xiu-Fen
XF
Lu
Hui-Ling
HL
D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology / Q000187:drug effects*; D000818:Animals; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000235:genetics; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D016259:Genes, myc; D054439:Ghrelin / Q000494:pharmacology*; D051379:Mice; D047495:PPAR gamma / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects*; D013937:Thymidine Kinase / Q000235:genetics
D022763:CCAAT-Enhancer-Binding Protein-alpha; D054439:Ghrelin; D047495:PPAR gamma; D012333:RNA, Messenger; D013937:Thymidine Kinase