| Objectives:There has been increasing concern regarding deleterious effects of environmental pollution on ecological environment and human health.Perfluorooctane sulfonate(PFOS)is a persistent organic pollutant that has been shown to cause multiple toxicity.In this study,we observed the hepatotoxicity of PFOS in mice.Furthermore,we investigated the protective effect of naringin(Nar)on PFOS-induced liver injury through determining pathologic changes and serum enzyme activities.Moreover,we measured the expression of nuclear factor erythroid 2-related factor 2(Nrf2),antioxidant enzyme genes,nuclear factor-κB(NF-κB),inflammatory factors and apoptosis-related proteins to reveal whether the protection of Nar against PFOS-induced hepatic injury is involved in the regulation of oxidative stress,inflammatory response and apoptosis.These results will provide a new target and an antioxidant intervention strategy of biologically active natural substances for the prevention and treatment of hepatotoxic damage caused by perfluorinated compounds.Methods:Thirty-two healthy male Kunming mice were randomly divided into four groups:control,PFOS(10 mg/kg/day),Nar(100 mg/kg/day)and PFOS(10 mg/kg/day)+ Nar(100 mg/kg/day),with 8 mice in each group.After animals were treated once daily by oral gavage for 21 consecutive days,hepatic histopathological changes and serum alanine aminotransferase(ALT),aspartate aminotransferase(AST)and lactate dehydrogenase(LDH)activities were observed in all the four groups.The production of malondialdehyde(MDA)and hydrogen peroxide(H2O2),activity of superoxide dismutase(SOD)and content of glutathione(GSH)in liver tissue homogenates were measured to evaluate the oxidative stress status.Antioxidant enzyme genes SOD,heme oxygenase-1(HO-1)and catalase(CAT)were detected by real-time PCR.The expression of Nrf2,NF-Κb,p65,p-p65,p53,Bax and Bcl-2 proteins were determined by Western blotting.The activity of caspase-3 in liver homogenate was determined by colorimetry.The levels of interleukin 6(IL-6)and tumor necrosis factor-α(TNF-α)were measured by ELISA.Results:(1)Nar reduced serum enzyme activities in mice exposed to PFOS:Exposure to PFOS for 21 days resulted in severe liver injury,which was characterized by significant increase in AST,ALT and LDH activities and relative liver weight(P < 0.05).However,in mice treated with PFOS,the addition of Nar restored elevated liver enzyme levels and reduced relative liver weight(P < 0.05).There was no significant difference in serum biochemical indexes of liver function between the control group and the Nar treatment group(P > 0.05).(2)Nar improved liver histopathological changes caused by PFOS:PFOS administration caused obvious histopathological changes in the liver of mice,including inflammatory cell infiltration,structural disorder,marked edema,cytoplasmic vacuolation and focal necrosis.However,PFOS-induced pathological changes were significantly reduced by Nar supplementation.Mice treated with Nar alone showed normal liver structure as the untreated mice.(3)Nar inhibited PFOS-induced hepatic oxidative stress:Compared with the control group,PFOS exposure increased the generation of MDA and H2O2 and reduced SOD activity and GSH content in the liver of mice(P <0.05).However,Nar supplementation decreased MDA and H2O2 levels and increased SOD activity and GSH content in mice exposed to PFOS(P < 0.05).Compared with control mice,Nar treatment alone did not affect liver MDA and H2O2 production,but significantly increased GSH content(P > 0.05).In addition,PFOS treatment markedly suppressed the expression of Nrf2 and its downstream antioxidant enzyme genes SOD,CAT and HO-1 in the liver(P < 0.05).However,PFOS-induced down-regulation of Nrf2 and antioxidant enzyme genes were significantly alleviated by simultaneous treatment with Nar(P < 0.05).Compared with control group,the levels of Nrf2 protein and HO-1,CAT and SOD m RNA were dramatically increased by Nar treatment alone in the liver of mice(P <0.05).(4)Nar alleviated hepatocellular apoptosis caused by PFOS:PFOS challenge for 21 days led to apoptotic liver injury,which was manifested as increased expression of pro-apoptotic proteins p53 and Bax and decreased expression of anti-apoptotic protein Bcl-2(P < 0.05).Furthermore,PFOS exposure resulted in a significant increase in caspase-3 activity in the liver of mice(P < 0.05).However,the elevated p53 and Bax expression and caspase-3 activity and the declined Bcl-2 expression were obviously inhibited by combined treatment with Nar in the liver of PFOS-exposed mice(P < 0.05).Compared with control group,Nar treatment alone significantly suppressed p53 expression(P < 0.05),but had no effect on Bax and bcl-2 expression and caspase-3 activation(P > 0.05).(5)Nar alleviated PFOS-induced inflammatory liver injury:After exposure to PFOS for 21 days,the expression of p-p65,TNF-α and IL-6 in the liver was significantly increased compared with the control group(P < 0.05).However,compared with PFOS group,Nar treatment obviously inhibited p65 phosphorylation and reduced the production of TNF-α and IL-6 in the liver(P < 0.05).Nar treatment alone did not alter the hepatic levels of p-p65,TNF-α and IL-6(P >0.05).Conclusion:(1)PFOS can induce oxidative stress,apoptosis and inflammation,resulting in hepatotoxic injury in mice.(2)Nar treatment can relieve PFOS-caused hepatotoxicity by alleviating oxidative stress,inflammatory response and apoptosis.(3)The mechanisms of Nar action may involve the up-regulation of Nrf2 expression and the inhibition of NF-κB activation. |
