维生素C对建鲤肠上皮细胞氧化损伤保护作用的研究

Protective Effect of Vitamin C on Oxidative Damage in Intestinal Epithelial Cells of Jian Carp(Cyprinus carpio var.Jian)

建鲤肠上皮细胞(IEC)在24孔培养板中原代培养72 h并更换无血清DMEM培养液继续培养12 h后,随机分为7组,每组4个重复。除其中1组作为空白对照外,另外6组均加入100μmol/L的过氧化氢(H2O2)以建立IEC氧化应激模型。相同条件下继续培养16 h后,将培养液分别更换为含0(空白对照组)、0、4、10、16、22和28 mg/L维生素C的无血清DMEM培养液,相同条件下继续培养72 h后取样测定指标,以考察不同维生素C浓度对H2O2诱导的建鲤IEC氧化损伤的保护作用。结果显示:1)100μmol/L H2O2处理导致IEC存活数量减少,仅形成一些小细胞集落,并显著降低IEC中超氧化物歧化酶、过氧化氢酶、谷胱甘肽过氧化物酶、谷胱甘肽硫转移酶和谷胱甘肽还原酶活性以及谷胱甘肽和蛋白质含量(P<0.05),提高IEC中丙二醛及蛋白质羰基含量以及培养液中乳酸脱氢酶活性(P<0.05),使建鲤IEC产生了氧化应激;2)维生素C显著提高IEC的抗羟自由基能力和抗超氧阴离子自由基能力(P<0.05),缓解由H2O2导致的建鲤IEC氧化损伤。综上可见,维生素C可提高建鲤IEC抗氧化能力,有效保护IEC免受氧化损伤。

This experiment was conducted to investigate the protective effect of different concentrations of vitamin C on hydrogen peroxide(H2O2)-induced oxidative damage in intestinal epithelial cells(IEC) of Jian carp.In 24-well culture plates,the Jian carp IEC were primarily cultured for 72 h,and then continued to culture in serum-free DMEM culture medium.After 12 h further culture,the IEC were randomly divided into 7 groups with 4 replicates per group.One of the 7 groups was set as blank control,and the other groups were all supplemented with 100 μmol/L H2O2 in culture medium in order to establish oxidative stress model.After 16 h further culture in the same condition,the culture medium was changed into the DMEM culture medium which contained 0(blank control group),0,4,10,16,22 and 28 mg/L vitamin C,respectively.The samples were collected to measure the indices after 72 h vitamin C culture.The results showed as follows: 1) the 100 μmol/L H2O2 treatment led to less survival number and small colony of IEC;significantly decreased the activities of superoxide dismutase(SOD),catalase(CAT),glutathione peroxidase(GPX),glutathione S-transferase(GST) and glutathion reductase(GR),and reduced the contents of glutathione(GSH) and protein(P<0.05);significantly increased the contents of malonaldehyde(MDA) and protein carbonyl(PCA),and improved the lactate dehydrogenase(LDH) activity in culture medium(P<0.05);caused oxidative stress of Jian carp IEC.2) Vitamin C significantly increased the anti-hydroxyl free radical ability(AHA) and anti-superoxide anion free radical ability(ASA)(P<0.05),and recovered the oxidation damage in IEC of Jian carp induced by H2O2.In summary,vitamin C can improve the antioxidant ability of the Jian carp IEC,and effectively protect the IEC from oxidative damage.