Effect Of GDF11 On Obesity-related Insulin Resistance And Inflammation And Its Mechanism

PART I EFFECT OF GDF11 ON OBESITY-RELATED INSULIN RESISTANCEObjective: GDF11 is a member of the TGF-? superfamily.Our previous study found that GDF11 levels in serum and liver of obese mice with insulin resistance(IR)were significantly reduced compared to normal mice.The liver plays a crucial role in metabolism,and obesity often causes liver lipid accumulation and IR.This study was to investigate the effect of GDF11 on obesity-related IR from the perspective of hepatic metabolism by using a model of insulin resistance induced by palmitate(PA)-induced Hep G2.Methods:(1)A high-fat diet(HFD)-induced obesity mice model was established in vivo.IPGTT and ITT tests were used to detect glucose tolerance and insulin sensitivity in mice.The mice serum levels of GDF11 were detected by ELISA.The protein and m RNA expression levels of GDF11 in liver were determined by Western blot and real-time PCR.(2)In vitro,a model of insulin resistance was induced by palmitate(PA)-induced Hep G2 hepatocytes,and the protein and m RNA expression levels of GDF11 were detected.(3)PA-induced Hep G2 hepatocytes were treated with GDF11.Levels of triglyceride and cholesterol were detected.Western blot were used to detect the the protein expression levels of P-AKT/AKT,and RT-PCR were used to detect the the m RNA expression level of PEPCK and G-6-Pase.Results:(1)In vivo: compared with low fat diet(LFD)group,the HFD group had higher body and liver weights,decreased insulin sensitivity.The HFD group had decreased serum and liver expression levels of GDF11(P < 0.05).(2)In vitro: compared with control,PA-induced Hep G2 had lower protein and m RNA expression levels of GDF11(P < 0.05),higher levels of triglyceride and cholesterol,lower protein levels of P-AKT/AKT,higer m RNA levels of PEPCK and G-6-Pase(P < 0.05).(3)Compared with PA group,the levels of triglyceride and cholesterol,protein levels of P-AKT/AKT,m RNA levels of PEPCK and G-6-Pase did not change significantly in GDF11 combined PA treatment group(P > 0.05).Conclusion: We found that the expression levels of GDF11 were significantly reduced in serum and liver tissues in obese-induced mice.The expression levels of GDF11 also decreased in PA-induced Hep G2.GDF11 treatment did not improve lipid accumulation and insulin resistance in PA-induced Hep G2.PART II EFFECT OF GDF11 ON PA-INDUCED MACROPHAGES AND ITS MECHANISMObjective: Obesity is closely related to chronic inflammation.Our previous study found that the level of inflammation in obese mice increased and the level of GDF11 decreased.GDF11 is a member of the TGF-? superfamily,and TGF-? can inhibit the inflammatory response of macrophages.Studies have shown that GDF11 can reduce endothelial inflammation and reduce atherosclerosis in mice.Therefore,we speculate that GDF11 has anti-inflammatory effect.This study was to investigate the effect of GDF11 on macrophage inflammation and explore possible mechanisms by treating RAW264.7 macrophages with PA.Methods:(1)The mice serum levels of IL-6 and TNF-? in obese mice were detected by ELISA.(2)Raw264.7 macrophages were treated with PA to induce inflammation.Western blot and real-time PCR were used to detect the the protein and m RNA expression level of GDF11.(3)PA-induced Raw264.7 were treated with GDF11,the supernatant levels of IL-6 and TNF-? were detected by ELISA,the protein expression levels of P-smad2/smad2,P-smad3/smad3 and P-JNK/JNK were determined by Western blot.(4)After pretreated with smad2/3 inhibitor SB431542,PA-induced Raw264.7 were treated with GDF11.The supernatant levels of IL-6 and TNF-? were detected by ELISA,the protein expression levels of P-JNK/JNK were determined by Western blot.Results:(1)In vivo: compared with low fat diet(LFD)group,the HFD group had higher serum levels of IL-6 and TNF-?,lower serum levels of GDF11.(2)Compared with control,PA-induced RAW264.7 had lower protein and m RNA expression levels of GDF11(P < 0.05).(3)Compared with PA group,GDF11 combined PA treatment group had lower levels of IL-6 and TNF-?,lower protein expression levels of P-JNK/JNK and higher protein expression levels of P-smad2/smad2 and P-smad3/smad3(P < 0.05).(4)Compared with GDF11 group,SB431542 pretreatment group had higher levels of IL-6 and TNF-?,higher protein expression levels of P-JNK/JNK(P < 0.05).Conclusion: We found that the expression levels of GDF11 were significantly reduced in PA-induced RAW264.7.GDF11 alleviate the inflammation of PA-induced RAW264.7 by inhibiting JNK via smad2/smad3 pathways.