Study On The Regulation And Mechanism Of TNF-? On Visfatin Expression In Trophoblast Cells

Visfatin,an adipocytokine that is mainly expressed and secreted by placenta and visceral adipose tissue,can be involved in the regulation of inflammatory response,and has insulin mimetic effects,and influences the metabolism of glucose and lipid.In recent years,it has been found that visfatin is associated with insulin resistance(IR)and plays an important role in the occurrence and development of gestational diabetes mellitus(GDM).The nuclear receptor peroxisome proliferator-activated receptor ?(PPAR-?)is predominantly produced in adipose tissue.It can be activated by antidiabetic thiazolidinediones,regulating glucose and lipid metabolism,inhibiting the expression of inflammatory factors and exerting anti-inflammatory effects.c-Jun n-terminal kinase(JNK),as a serine/threonine protein kinase,can be activated by a variety of cytokines and stress stimulation,and is considered to be an important signal transduction molecule linking inflammation and insulin resistance.Objective: The aim of this study is to investigate the effect of TNF-? on the expression of visfatin in cultured BeWo cells and the regulatory mechanisms involved.Methods: BeWo cells were incubated with TNF-? at various concentrations(0,1,10 and 100 ng/mL)for 48 hours.The transcription levels of visfatin and PPAR-? mRNA were detected by FQ RT-PCR,and the optimum concentration was selected.BeWo cells were treated for 12,24,48 or72 h with the effective concentration of TNF-? to determine the transcription levels of visfatin and PPAR-? mRNA by FQ RT-PCR and select an effective time.After intervening BeWo cells with the effective concentration and time,the expression levels of visfatin,PPAR-?,JNK and p-JNK proteins were detected by Western blot.Then,after adding the JNK inhibitor SP600125,theexpression levels of visfatin,JNK and p-JNK protein were detected by Western-blot.Extracellular visfatin secretion level was detected by ELISA.Results:1.The different concentrations of TNF-?(1,10,100ng/mL)all reduced the expression of visfatin and PPAR-? mRNA in a concentration-dependent manner.Correlation analysis indicated that visfatin and PPAR-? mRNA expression were positively correlated(r=0.794,P<0.01).2.Treated with TNF-?(100ng/mL)for 12,24,48 or 72 h,BeWo cells PPAR-? mRNA expression level was only decreased at 48 h,while visfatin mRNA expression levels were not obviously altered after 12 or 24 h,but its expression levels decreased after 48 and 72h(P<0.01)and reached to the minimum level at 48 h.There was positive correlation between visfatin and PPAR-? mRNA expression(r=0.749,P<0.01).3.BeWo cells were incubated with TNF-?(100ng/mL)for 48 h,PPAR-?and JNK proteins expression had no significant change,while visfatin protein expression was decreased(P<0.01),and p-JNK protein expression level increaced(P<0.05),which was negatively correlated with visfatin protein level(r=-0.816,P=0.048).After adding JNK inhibitor SP600125,compared with no inhibitor group,p-JNK protein expression decreased,while visfatin expression increased(P<0.01).4.In the control group,TNF-? group and TNF-?+SP600125 group,after48 h treatment,the secretion level of visfatin in TNF-? group was slightly lower than that in control group(0.110±0.066 vs.0.169±0.092 ng/ml,P=0.38).The visfatin secretion in TNF-?+SP600125 group was slightly higher than that in TNF-? group(0.235±0.067 vs.0.110±0.066 ng/ml,P=0.09).Conclusions:1.TNF-? could downregulate the expression of visfatin in BeWo cells by activating the JNK signaling pathway.2.TNF-? inhibited the mRNA expression of PPAR-? in BeWo cells,which was positively correlated with visfatin mRNA expression.The PPAR-?may be involved in the regulation of visfatin expression in Be Wo cells by TNF-? mainly at the transcriptional level.