| ObjectiveTo create a rat model of Non-alcoholic Fatty Liver Disease (NAFLD) by fedwith fat-rich diet, and observe the changing of rats’ weight, liver weight, liver index,insulin resistance (IR), bloodfat, hepatic enzyme, the number of Treg and theexpression of Treg-associated cytokines (IL-10、TGF-β1) during the process offorming NAFLD, the association of each marker with hepatic steatosis, and theassociation of Treg and their associated cytokines (IL-10、TGF-β1) with IR wereanalyzed. and the possible immunological mechanism of Treg discussed, which wouldlay a foundation for further study on the immunotherapy of NAFLD.MethodsForty-eight contemporary male SD rats weighed160±10g were randomlydivided into two groups, which respectively nanmed fat-rich group and control group(24rats for each group). Control group was fed with normal diet while fat-rich groupfed with fat-rich diet (88%normal diet+2%cholesterol+10%lard). At the end of4th、8th、16thweek,8rats in control group and fat-rich group were respectively selectedrandomly and gaven into the experiment. Serum FBS, ALT, AST, TG and TC weremeasured by colorimetric method, FINS was measured by chemiluminescence assay,and FIRI was caculated. The number of peripheral blood Treg were detected by FCMlivers were dissoeiated and weighted, and liver index were caculated. Color andtexture of liver were recorded. Liver tissue were sunk into10%formaldehydesolution and prepared to do pathological observation. Foxp3, IL-10and TGF-β1geneexpression were detected by Real-time PCR. Results1FG had a significantly elevated levels of body weight, liver weight and liver index incomparison with NG at the end of4thweek (p=0.030,0.012,0.012), and moresignificantly elevated levels at the end of8thand16thweek(p=0.002,0.005,0.008.p=0.001,0.001,0.001)2FG had a significantly elevated levels of serurn ALT, AST, TC, TG in comparisonwith NG at the homologns week (p=0.001,0.001,0.002,0.001. p=0.001,0.001,0.001,0.002. p=0.000,0.000,0.000, p=0.000)3There is no change in the level of FBS during feeding. And all rats of FG had nosignificantly difference levels of FBS in comparison with NG (p=0.621,0.282,0.195). FINS and FIRI were gradually increased in the progress by feeding fatrich-diet, there were no significant differences between two groups at the end of the4thweek（p=0.572,0.474）, at the end of the8thand16thweek，there were significantdifferences between two groups (p=0.001,0.001. p=0.001, p=0.001)4The expression of Foxp3, IL-10in FG were significant decreased in comparison withNG at the end of the4th,8thand16thweek (p=0.002,0.002. p=0.000,0.000. p=0.001,0.001). In comparison with NG, there is no significant difference in theexpression of TGF-β1at the end of4thweek (p=0.355). But the expression ofTGF-β1were significant increased in comparison with NG at the end of8thand16thweek (p=0.001,0.000)5The proportion of peripheral blood CD4+CD25+Foxp3+Treg/CD4+T in FG weresignificantly decreased in comparison with NG at the end of4th,8thand16thweek(p=0.002,0.003,0.000)6The hepatic steatosis had appeared since the4thweek, then the moderate fatty liverdisease was developed at the end of the8thweek, and serious one at the end of the16thweek.7In FG, there was no linear correlative relation between FBS and hepatic steatosis (r=0.489), however body weight, liver weight, liver index, TC, TG, ALT, AST, FINS,FIRI and TGF-β1were all positively correlated with hepatic steatosis (r=0.968, 0.963,0.921,0.950,0.926,0.945,0.924,0.966,0.979,0.877), and FIRI coefficientcorrelation most heavy among them(r=0.979). But CD4+CD25+Foxp3+Treg, Foxp3,IL-10were negatively correlated with hepatic steatosis (r=-0.963,-0.914,-0.928),and CD4+CD25+Foxp3+Treg coefficient correlation most heavy among them (r=-0.963)8Foxp3and IL-10were all positively correlated with CD4+CD25+Foxp3+Treg/CD4+T(r=0.872and0.839respectively),but TGF-β1were negatively correlated withCD4+CD25+Foxp3+Treg/CD4+T (r=-0.910)9TGF-β1was positively correlated with FIRI (r=0.924), CD4+CD25+Foxp3+Treg,Foxp3and IL-10were all negatively correlated with FIRI (r=-0.935,-0.891,-0.936)Conclusion1NAFLD with IR model was sueeessfully developed in SD rats fed with fat-rich dietfor8weeks2The reduction in the number of Treg can lead to dysfunction in immune of livertissue, and increase the level of IR, which may be the key factor leading tooccurreuce and development of NAFLD3IL-10secreted by treg plays a protective role in the development of NAFLD,butTGF-β1plays a promoting role in it4Detection of Treg in peripheral blood may has a certain clinical significance fordiagnosis of NAFLD, and the change of the number of Treg may conduce to impedeoccurreuce and development of NAFLD... |
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