Effects And Mechanisms Of Green EGCG On Oxidative Stress And Inflammation Of 3T3-L1 Adipocytes

Obesity is a multi-factor-induced chronic disease.In recent years,with the improvement of people's living standard and nutritional conditions,obesity has become a global public concern,especially in industrialized countries.Obesity is defined as a state of excessive accumulation of adipose tissue due to hyperplasia andhypertrophy of adipocytes..Adipose tissue is not only an organ that stores fat,butalso an important place to regulate endocrine metabolism.It can secrete Leptin,Monocyte Chemotactic Protein-1(MCP-1)and interleukin-6(IL-6)and other fatty factors,These factors are associated with many physiological and pathological processes.Obese people with chronic inflammatory response and oxidative stress response,which in turn induced cell damage,dysfunction and decreased insulin sensitivity,leading to a series of pathological reactions.Therefore,the study of adipose tissue oxidative stress and inflammation and its molecular mechanism provides a new strategy for the prevention and treatment of obesity and relateddiseases.Epigallocatechin-3-gallate(EGCG)is the most abundant and active ingredient in green tea.The role of EGCG in the prevention and treatment of obesity has been confirmed by a large number of studies.At the same time,more and more studies have shown that EGCG has a strong anti-inflammatory and anti-oxidation effect.However,whether EGCG can directly effect on adipocytes and have the potential to inhibit the inflammation and oxidative stress of adipocytes and the related mechanism there are few studies reported.In this study,3T3-L1 adipocytes were used to observe the effect of EGCG on oxidative stress and inflammation of 3T3-L1 adipocytes,and the effect of EGCG on Nrf2 / HO-1 of 3T3-L1 adipocytes was preliminary explored.This study provided a theoretical basis for the treatment of obesity and related chronic diseases by antioxidative and/or anti-inflammatory intervention.Purpose:1.To investigate the effect of EGCG on basal state and Lipopolysaccharide(LPS)induced 3T3-L1 adipocytesoxidative stress and inflammation;2.To explore the molecular mechanism of EGCG regulating adipose tissue oxidative stress and inflammation by targeting Nrf2.Methods:3T3-L1 preadipocytes were induced to differentiate into mature adipocytes.The following two states were simulated: 1)The basal state was treated with(0?g/ml),1?g/ml,10?g/ml,50?g/ml EGCG for differentiation of mature 3T3-L1 adipocytes for24 hours;LPS-induced inflammation state of adipocytes: LPS contrast group(0 ?g/ml EGCG),1 ?g/ml,10 ?g/ml,50?g/ml EGCG pretreatment for 2h,then plus 1?g/mL LPS intervention 24 h.1.Detection of oxidative stress levels:respectively,The activities of superoxide dismutase(SOD),glutathione(GSH)and malondialdehyde(MDA)in the different concentrations of EGCG on the two states above of the 3T3-L1 adipocyteswere measured by TBA,hydroxylamine and spectrophotometry.2.Detection of inflammation levels:1)The levels of IL-6,TNF-? and MCP-1 in the supernatant of basal and LPS-induced3T3-L1 adipocytes in different concentrations of EGCG were detected by enzyme-linked immunosorbent assay(ELISA).2)Real-time fluorescent quantitative PCR was used to detect the expression of IL-6,TNF-a and MCP-1 mRNA in 3T3-L1 adipocytes of EGCG in different concentrations.3.Nrf2 and HO-1 mRNA expression in 3T3-L1 adipocytes were detected by real-time fluorescence quantitative PCR with different concentrations of EGCG.Results:1.Whether the basal or LPS-induced state,1?g/ml ~ 50?g/ml EGCG intervention significantly decrease the content of MDA of mature 3T3-L1 adipocytes,and remarkably increase the SOD and GSH activity(P <0.05),and showed a dose-dependent relationship,The effect of 50?g/ml EGCG on the three oxidative stress indexes was the most significant.The results of 50?g /ml concentration in the basal state were compared with those of the blank group.50?g /ml compared with the blank group as follows: MDA 0.84 ± 0.37 vs 3.01 ± 0.99;SOD 143.17 ± 14.18 vs 68.69 ± 5.32;GSH 308.12 ± 85.16 vs 148.87 ± 14.65;LPS induced 50?g /ml concentration group compared with LPS control group as follows: MDA 1.30 ±0.05 vs 9.81 ± 0.10,SOD 188.35 ± 13.87 vs 46.12 ± 3.44;GSH 366.60 ±60.40 vs 121.20 ± 0.31.2.Whether the basal or LPS-induced state,1?g/ml ~ 50?g/ml EGCG intervention on mature 3T3-L1 adipocytes inflammatory factors IL-6,TNF-a and MCP-1 mRNA expression and secretion were significantly decreased(P <0.05),and there was a dose-dependent relationship.The effect of 50?g / ml EGCG was the most obvious:the concentration of 50?g/ml in basal state was compared with the blank group the results were as follows: IL-6 272.59 ± 6.61 vs 216.73 ± 22.81,MCP-1 1021.68± 150.09 vs 2134.83 ± 210.42,TNF-a 4.01 ± 1.03 vs 10.33 ± 1.18;LPS induced 50?g / ml concentration group compared with LPS control group as follows:IL-6 107.94 ± 26.22 vs 272.59 ± 6.61,MCP-1 1009.93 ± 37.34 vs 2299.46 ±93.99,TNF-a 5.53 ± 2.07 vs 18.51 ± 0.49.3.Whether the basal or LPS-induced state,the expression of Nrf2 and HO-1 mRNA in mature 3T3-L1 adipocytes was significantly increased by 10?g/ml ~ 50?g/ml EGCG(P <0.05).And showed a certain dose-dependent relationship.And 50?g/ml EGCG intervention effect is most obvious.Conclusion:EGCG inhibited the oxidative stress and inflammatory effects of 3T3-L1 adipocytes and LPS-induced 3T3-L1 adipocytes.The effect of EGCG on oxidative stress and inflammation of 3T3-L1 adipocytes may be related to up-regulation of Nrf2/ HO-1.