α-酮戊二酸二甲酯预处理对犬脂肪间充质干细胞氧化应激损伤的保护作用

doi:10.16431/j.cnki.1671-7236.2021.12.042

Abstract

Abstract: The aim of the present study was to explore the effect of dimethyl α-ketoglutarate (DMKG) on antioxidative stress injury of canine adipose-derived mesenchymal stem cells(ADSCs).The canine ADSCs were treated with DMEM/F12 basic medium which containing hydrogen peroxide (H₂O₂) at concentrations of 0, 0.2, 0.4, 0.6, 0.8 and 1.0 mmol/L.After 1 h, the cell survival rate was detected by CCK-8 assay to screen the optimal concentration for building canine ADSCs oxidative stress model.In this experiment, the well-growing passage 3 canine ADSCs were divided into three groups: Blank control group, model group and DMKG group.The blank control group was cultured normally without treatment.In the model group, canine ADSCs were treated with the best concentration of H₂O₂ solution for 1 h. In DMKG group, canine ADSCs were pretreated with 1 mmol/L DMKG for 24 h, and then treated with the best concentration of H₂O₂ solution for 1 h.After the culture of each group, the cell survival rate of each group was detected by CCK-8 assay, the content of reduced glutathione was detected by visible spectrophotometry, the level of reactive oxygen was detected by DCFH-DA fluorescent probe, the apoptosis rate was detected by Annexin V/PI double staining, and the mRNA expression levels of superoxide dismutase(SOD), catalase (CAT), anti-apoptotic gene Bcl-2, pro-apoptotic genes Bax and Cleaved-Caspase3 were detected by Real-time quantitative PCR.The results showed that compared with the model group, DMKG pretreatment could effectively reduce the production of reactive oxygen species (P<0.05), increase the cell survival rate (P<0.01), and decrease the rate of cell apoptosis (P<0.05).Meanwhile, DMKG pretreatment could increase the content of reduced glutathione (P<0.01), up-regulate the expression levels of antioxidant enzyme-related genes SOD1, SOD2 (P<0.01)and CAT (P<0.05), down-regulate the expression levels of Cleaved-Caspase3 gene (P<0.05)and increase the rate of Bcl-2/Bax (P<0.05).In conclusion, DMKG pretreatment could improve the ability of anti-oxidation and anti-apoptosis of canine ADSCs, so as to increase the survival of canine ADSCs under oxidative stress.

Key words: canine adipose-derived mesenchymal stem cells(ADSCs); dimethyl α-ketoglutarate(DMKG); oxidative stress; cell apoptosis

CLC Number:

S852.1

YAO Jiawei, HUANG Yujie, HUANG Lizhen, RUAN Huimin, CHEN Zhisheng, WANG Bingyun, CHEN Shengfen, ZHANG Hui, BAI Yinshan, LIU Canying, JI Huiqin. Protective Effect of Dimethyl α-Ketoglutarate Pretreatment on Oxidative Stress in Canine Adipose-derived Mesenchymal Stem Cells[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(12): 4735-4743.

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Protective Effect of Dimethyl α-Ketoglutarate Pretreatment on Oxidative Stress in Canine Adipose-derived Mesenchymal Stem Cells

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