维生素C在对乙酰氨基酚致小鼠急性肝脏损伤中的作用

Role of vitamin C in acute liver injury induced by acetaminophen in mice

为了探究维生素C对对乙酰氨基酚(APAP)致小鼠肝脏损伤的影响,试验将60只雄性健康昆明小鼠随机分为对照组、APAP组及维生素C干预组(VC+APAP组),除对照组外,APAP组和VC+APAP组小鼠均按体重腹腔注射350 mg/kg的APAP,按体重200μL/10 g给药,对照组小鼠注射等体积的0.9%氯化钠溶液,VC+APAP组在APAP注射前30 min和注射后12 h按体重腹腔注射30 mg/kg的维生素C。APAP处理24 h后收集小鼠血液样本,检测各组小鼠肝脏功能指标,包括丙氨酸氨基转移酶(ALT)和天冬氨酸氨基转移酶(AST);用苏木精-伊红染色法观察小鼠肝脏病理组织学变化;用原位末端标记(TUNEL)法测定肝脏细胞凋亡数;二氢乙啶(DHE)活性氧(ROS)荧光探针检测肝脏氧化应激水平;用ELISA试剂盒检测白细胞介素6(IL-6)与肿瘤坏死因子α(TNF-α)质量浓度;用免疫印迹法检测Bcl-2相关X蛋白(Bax)、B细胞淋巴瘤-2(Bcl-2)和核转录因子κB(NF-κB)蛋白的表达情况。结果表明:与APAP组相比,VC+APAP组小鼠血清中的ALT和AST水平显著降低(P<0.05),小鼠肝脏组织病理学损伤明显减轻,TUNEL阳性细胞百分率和Bax蛋白相对表达量显著降低(P<0.05),Bcl-2蛋白相对表达量显著升高(P<0.05);小鼠肝脏组织中ROS相对荧光强度和丙二醛(MDA)浓度显著降低(P<0.05),而超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性及谷胱甘肽(GSH)浓度显著升高(P<0.05);肝脏NF-κB蛋白相对表达量及TNF-α、IL-6质量浓度显著降低(P<0.05)。说明维生素C对APAP诱导的急性肝脏损伤具有保护作用,这种保护作用可能与其抑制肝细胞凋亡、降低肝脏氧化应激和炎症反应有关。

In order to investigate the effects of vitamin C on acetaminophen(APAP)-induced liver damage in mice,in this experiment,sixty healthy male Kunming mice were selected and randomly divided into control group,APAP group,and vitamin C intervention group(VC+APAP group).Apart of the control group,the remaining two groups were intraperitoneally injected with 350 mg/kg APAP(injection volume:200μL/10 g).The mice in the control group were injected with the same volume of 0.9%sodium chloride.Mice in VC+APAP group were intraperitoneally injected with 30 mg/kg VC 30 min before and 12 h after APAP treatment,respectively.Blood samples were collected after 24 h of APAP treatment,the changes of liver function indexes alanine aminotransferase(ALT)and aspartate aminotransferase(AST)in each group were detected.Hematoxylin-eosin staining method was used to observe the pathological changes of mice livers.TUNEL assay was used to detect the apoptosis.Fluorescence probe for reactive oxygen species(ROS)and colorimetry method were used to measure oxidative stress level in liver.Enzyme-linked immunosorbent assay was used to determine the concentrations of interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α).The protein expressions of BCL2 associated X protein(Bax),B cell leukemia/lymphoma 2(Bcl-2),and Nuclear factor NF-kappa-B(NF-kB)were measured by immunobloting.The results showed that,compared with APAP group,the levels of serum ALT and AST in VC+APAP group were significantly decreased(P<0.05);the histopathological injury of mouse liver was significantly alleviated;the ratio of TUNEL-positive cells and the relative protein expression of Bax were significantly decreased(P<0.05);the relative protein level of Bcl-2 was markedly increased(P<0.05).The fluorescence intensity of ROS and malondialdehyde(MDA)content were significantly decreased(P<0.05),while the contents of superoxide dismutase(SOD),catalase(CAT),and glutathione(GSH)were significantly increased(P<0.05)in VC+APAP group,which had significant differences compared with those of in APAP group(P<0.05).Compared with APAP group,the relative expression of NF-kB protein,the concentrations of TNF-αand IL-6 in VC+APAP group were significantly decreased(P<0.05).It indicated that VC had a protective effect on APAP-induced acute liver injury,which might be related to inhibiting of hepatocyte apoptosis,reducing of oxidative stress and inflammation.