Mechanism Of PPARγ Agonists Improve Insulin Resistance Via Regulating Chemerin In Gestational Diabetes Mellitus

Part One Different Expressions of Chemerin and PPARγ in GDM Patients and Normal Pregnant WomenObjectiveTo examine the different expressions and significance of adipokine chemerin as well as peroxidase proliferator-activated receptor γ(PPARγ)in clinical specimens from normal pregnant women and patients with gestational diabetes mellitus(GDM).MethodsSpecimens of placental tissue,subcutaneous adipose tissue,umbilical cord blood and peripheral blood were collected from GDM patients and normal pregnant women that suffered cesarean surgery,and enzyme-linked immunosorbent assay was used to quantitatively analyze the concentrations of chemerin in cord blood and peripheral blood.The expressions of chemerin in placental tissue and subcutaneous adipose tissue were detected by immunohistochemistry.Western blot and RT-q PCR were used respectively to detect the expression of PPARγ in placenta.Results1.In GDM patients,the concentration of chemerin in plasma of umbilical cord blood was significantly higher than that of peripheral blood(p<0.05),while there were no significant differences between GDM patients and normal pregnant women with respect to the expressions of chemerin in plasma.2.Compared with normal pregnant women,the expressions of chemerin in placenta and subcutaneous adipose tissue of GDM patients increased(p<0.01).3.Compared with normal pregnant women,the protein and m RNA expression levels of PPARγ in the GDM placenta decreased,the differences were statistically significant(p<0.05).ConclusionsCompared with normal pregnant women,the level of chemerin is higher in the GDM placenta and subcutaneous adipose tissue,whereas the expression of PPARγ is down-regulated in the GDM placenta.Increased level of chemerin in the cord blood in GDM patients may be partly derived from the secretion of placental tissues.Part Two Mechanism of PPARγ Agonists Improve Insulin Resistance via Regulating Chemerin in Cell ModelObjective To investigate the effect of PPARγ on chemerin and its receptor,explore the influence of recombinant human chemerin on the key molecules of insulin signaling pathway,and clarify the role of chemerin receptors in the process of improving insulin resistance by PPARγ agonists.Methods High glucose cell model reported in multiple literatures was used in HTR-8/SVneo cells,a human chorionic trophoblast cell line,to formation a GDM placental trophoblast cell model.Immunocytofluorescense and western blot were used to detect the expressions of chemerin,chemokine-like receptor 1(CMKLR1)and G protein-coupled receptor 1(GPR1)in HTR-8/SVneo cells.RT-q PCR experiment was applied to detect the effect of 100ng/m L recombinant human chemerin as well as activation or inhibition of PPARγ on m RNA expression levels of phosphatidylinositol 3-kinase(PI3K),protein kinase B beta(AKT2)and mitogen-activated protein kinase 1(MAPK1).Western blot was used to detect the effect of PPARγ agonists(rosiglitazone and GW1929)on chemerin and its two receptors.Small interference RNA(si-RNA)method was used to silence the CMKLR1 and GPR1,respectively,and western blot was performed to explore the mechanism of action of chemerin receptors in the process of improving insulin resistance by the activation of PPARγ.Results 1.Chemerin,CMKLR1 and GPR1 were all expressed in high glucose cultured HTR-8/SVneo cells.2.Compared with the control group,100ng/m L recombinant human chemerin significantly increased the m RNA levels of PI3 K,AKT2 and MAPK1(p<0.05).3.Compared with the control group,the m RNA expressions of PI3 K and AKT2 increased in the rosiglitazone(PPARγ agonist)group and the m RNA levels of PI3 K and MAPK1 were higher in the GW1929(PPARγ agonist)group,while the m RNA levels of AKT2 and MAPK1 were lower in the GW9662(PPARγ inhibitor)group,the differences were all statistically significant(p<0.05).4.Compared with the control group,the protein expressions of PPARγ and GPR1 in the rosiglitazone(PPARγ agonist)group increased(p<0.05),and the protein expression levels of PPARγ,chemerin and CMKLR1 in the GW1929(PPARγ agonist)group raised,too(p<0.01).5.Compared with the si-CMKLR1 group,si-CMKLR1+PPARγ agonists(rosiglitazone or GW1929)increased the level of ERK2 phosphorylated protein(p<0.01),but they did not promote the expressions of PI3 K p110β protein and AKT2 phosphorylated protein.The results illustrate that silencing CMKLR1 may have an effect on the enhancement of the PI3K-AKT pathway by PPARγ agonists.6.Compared with the si-GPR1 group,si-GPR1+PPARγ agonists(rosiglitazone or GW1929)still improved the protein levels of PI3 K p110β and AKT2(p<0.05),however,they did not activate ERK2 phosphorylated protein.It is suggested that GPR1 may play an important role in the process of activation of MAPK(ERK1/2)pathway.Conclusions Chemerin may enhance the activation of insulin signaling pathway.PPARγ agonists may improve insulin resistance via promoting the expressions of chemerin and its receptors CMKLR1 and GPR1.