| Objective(1)By using serum and pathology index to evaluate the clinical efficacy of Astragaloside Ⅳ in the treatment of nonalcoholic fatty liver disease(NAFLD)in rats.(2)To investigate the effect and its possible mechanism of Astragaloside Ⅳ on toll like receptor 4(TLR4),nuclear factor-kappa B(NF-κB),myeloid differentiation factor 88(My D88)levels in hepatic tissue and inflammatory factor levels in serum in rats with NAFLD.Methods(1)Sixty three male SD rats were selected,and randomly divided into high fat diet group and control(CON)group.There were 53 rats in the high fat diet group fed with high fat diet,and 10 rats in the CON group fed with normal diet.After 12 weeks,3rats from high fat diet group were confirmed NAFLD rat modle through observing the pathological changes of rat liver.Then the remaining 50 model rats in high fat diet group were randomly divided into 5 equal groups,model(MOD)group,Astragaloside Ⅳ low dose(AS-Ⅳ-L)group,Astragaloside Ⅳ middle dose(AS-Ⅳ-M)group,Astragaloside Ⅳ high dose(AS-Ⅳ-H)group and Polyene phosphatidylcholine(PPC)group.Intervention group and MOD group were given different dose Astragaloside Ⅳ,Polyene phosphatidylcholine or saline by intragastric adminstration respectively everyday for 4 weeks.(2)After 16 weeks of treatment,the body weight and liver index of all rats were tested.The pathological changes of liver tissue were observed by optical microscope after HE staining,and the NAFLD activity score(NAS)of liver tissue in each group was calculated.(3)Alanine aminotransferase(ALT),aspartate aminotransferase(AST),fasting blood glucose(FBG),triglyceride(TG)and total cholesterol(TC)of serum were tested by fully automatic biochemical analyser.(4)Enzyme linked immunosorbent assay(Elisa)was used to measure the concentration of serum fasting insulin(FINS),total anti-oxidation competence(T-AOC),superoxide dismutase(SOD),glutathione peroxidase(GSH-PX),malondialdehyde(MDA),interleukin-6(IL-6),interleukin-8(IL-8),interleukin-10(IL-10)and tumor necrosis factor-α(TNF-a).(5)Western blot was used to determine the expression of TLR4,My D88 and NF-κBp65 protein in liver tissue of rats.(6)Real-time quantitative RT-PCR was used to determine the expression of TLR4 m RNA,My D88 m RNA and NF-κBp65 m RNA in liver tissue of rats.(7)T test was used to compare the two groups of data.LSD-t was used to compare each group.Results(1)Compared with the MOD group,all of the Astragaloside Ⅳ groups were able to reduce the body weight of rats fed on high fat diet,but only the AS-Ⅳ-H group was statistically significant difference(584.45 ± 31.11 vs 533.67 ± 28.27,P<0.01).Compared with the MOD group,AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group could improve the liver index of NAFLD model rats.The difference was statistically significant(4.28 ± 0.32 vs 3.72 ± 0.08,P<0.01;4.28 ± 0.32 vs 3.69 ± 0.18,P<0.01;4.28 ± 0.32 vs 3.48 ± 0.21,P<0.01).(2)Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could reduce serum ALT level in NAFLD model rats.The difference was statistically significant(115.22 ± 24.65 vs 98.37 ± 10.4,P<0.05;115.22 ± 24.65 vs 92.00 ± 17.63,P<0.01).Compared with the MOD group,AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group could reduce the serum AST level of NAFLD model rats(224.86 ±32.23 vs 198.14 ± 30.05,P<0.01;224.86 ± 32.23 vs 181.71 ± 28.77,P<0.01;224.86± 32.23 vs 170.62 ± 16.30,P<0.01).Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could reduce serum TG level in NAFLD model rats.The difference was statistically significant(2.59 ± 0.31 vs 2.33 ± 0.41,P<0.05;2.59 ±0.31 vs 2.02 ± 0.22,P<0.01).Compared with the MOD group,the serum TC level in all of the Astragaloside Ⅳ groups was decreased,but the difference was no statistically significance(P>0.05).(3)Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could reduce serum FBG level in NAFLD model rats.The difference was statistically significant(18.21 ± 1.55 vs 16.51 ± 2.04,P<0.05;18.21 ± 1.55 vs 15.71 ± 1.78,P<0.01).Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could reduce serum FINS level in NAFLD model rats(7.35 ± 0.87 vs 6.35 ± 1.28,P<0.05;7.35 ± 0.87 vs 5.40 ± 0.77,P<0.01).AS-Ⅳ-M group and AS-Ⅳ-H group were able to decrease HOMA-IR in NAFLD model rats.The difference is statistically significant compared with the MOD group(5.97 ± 1.06 vs 4.75 ± 1.46,P<0.05;5.97 ± 1.06 vs3.81 ± 0.88,P<0.01).(4)Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could increase serum T-AOC level in NAFLD model rats(7.99 ± 0.60 vs 9.82 ± 1.17,P<0.01;7.99 ± 0.60 vs 10.78 ± 1.43,P<0.01).AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group could increase serum GSH-PX level in NAFLD model rats compared with the MOD group(172.85 ± 21.06 vs 214.15 ± 20.62,P<0.01;172.85 ± 21.06 vs251.35 ± 22.56,P<0.01;172.85 ± 21.06 vs 297.05 ± 27.48,P<0.01).Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could decrease serum MDA level in NAFLD model rats(6.84 ± 0.96 vs 6.30 ± 0.78,P<0.05;6.84 ± 0.96 vs 5.23 ±1.07,P<0.01).AS-Ⅳ-H group could significantly decrease serum SOD level in rats with high fat diet compared with the MOD group(170.63 ± 28.14 vs 144.75 ± 20.67,P<0.05).(5)Compared with the MOD group,AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group could decrease the NAS score of liver tissue in NAFLD model rats.The difference was statistically significant(7.3 ± 0.68 vs 6.3 ± 0.78,P<0.01;7.3 ± 0.68 vs5.1 ± 1.37,P<0.01;7.3 ± 0.68 vs 4.1 ± 1.51,P<0.01).(6)Compared with the MOD group,AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group could significantly decrease serum TNF-α level in NAFLD model rats(4.23 ± 1.25 vs 3.06 ± 0.97,P<0.05;4.23 ± 1.25 vs 2.81 ± 0.70,P<0.01;4.23 ±1.25 vs 2.10 ± 0.34,P<0.01).AS-Ⅳ-M group and AS-Ⅳ-H group could significantly decrease serum IL-6 level in NAFLD model rats compared with the MOD group(164.81 ± 26.29 vs 131.76 ± 22.35,P<0.01;164.81 ± 26.29 vs 125.45 ± 22.67,P<0.01).Compared with the MOD group,AS-Ⅳ-M group and AS-Ⅳ-H group could significantly decrease serum IL-8 level in NAFLD model rats(1.53 ± 0.44 vs 1.10 ±0.17,P<0.05;1.53 ± 0.44 vs 0.92 ± 0.24,P<0.01).AS-Ⅳ-M group and AS-Ⅳ-H group could significantly increase serum IL-10 level in NAFLD model rats(21.47 ±3.25 vs 32.11 ± 5.07,P<0.01;21.47 ± 3.25 vs 38.96 ± 8.71,P<0.01).(7)Compared with the MOD group,the expression of TLR4,My D88 and NF-κBp65protein in liver tissue of NAFLD model rats were significantly decreased in AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group(P<0.01).(8)The expression of TLR4 m RNA,My D88 m RNA and NF-κBp65 m RNA in liver tissue of NAFLD model rats were significantly decreased in AS-Ⅳ-L group,AS-Ⅳ-M group and AS-Ⅳ-H group.There were significant difference compared with the MOD group(P<0.05).Conclusions(1)Astragaloside Ⅳ could significantly reduce the body weight,improve hyperlipidemia,decrease the level of serum ALT and AST,improve IR and oxidative stress,decrease the liver index,reduce hepatic steatosis and lipids deposition in liver tissue,improve the pathological changes of liver of NAFLD rats induced by high fat diet.It possibly can be a promissing drug in the treatment of NAFLD.(2)Astragaloside Ⅳ inhibits the level of IL-6,IL-8,TNF-α and increases the level of IL-10 by down-regulating the expression of TLR4,My D88 and NF-κB in the liver of NAFLD rats,thus exerting the protective effect on NAFLD. |
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