地锦草水提物在IPEC-J2细胞中的抗炎与抗氧化作用

doi:10.16431/j.cnki.1671-7236.2023.02.028

Abstract

Abstract: 【Objective】 In order to reveal the anti-inflammatory and antioxidation effects of Euphorbiae humifusae (EH) in swine intestinal epithelial cells (IPEC-J2).【Method】 Different concentrations of EH aqueous extract (0,5,10,50,125,200 μg/mL) were used to treat IPEC-J2 cells for 12 hours.The viability of IPEC-J2 cells was detected by CCK-8 method,determining the optimal concentration of EH treatment.IPEC-J2 cells were randomly divided into control group (CT),lipopolysaccharide group (LPS),EH+LPS group (ELP),with three replicates in each group.The cells in CT group were cultured in DMEM medium containing 10 % fetal bovine serum.The cells in LPS group were treated with 5 μg/mL LPS.The cells in ELP group were co-cultured with 5 μg/mL LPS and EH,cells in each group was treated for 12 hours,collecting the cells and supernatant.The expression of interleukin-6 (IL-6),tumor necrosis factor α (TNF-α),kelch like epichlorohydrin related protein 1 (Keap1),nuclear factor E2 related factor 2 (Nrf2),heme oxygenase-1 (HO-1) mRNA were detected by real time fluorescent quantitative PCR,and the content of malondialdehyde (MDA),superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in the supernatant were detected by ELISA,and the reactive oxygen species (ROS) level in the cells was detected by chemical fluorescence method.【Result】 Compared with EH untreated cells,5,50 μg/mL EH increased significantly the viability of IPEC-J2 cells (P＜0.05).EH concentration was at 10 μg/mL,the increased cell viability was very significant (P＜0.01).When EH concentration was at 125 μg/mL,IPEC-J2 cell viability decreased,there was no statistical significance (P＞0.05).EH concentration is at 200 μg/mL,the decreased cell viability was extremely significant (P＜0.01).The concentration of EH treatment was 10 μg/mL in subsequent experiment.Compared with the CT group,in LPS group,the IL-6 and TNF-α genes mRNA expression in IPEC-J2 cells were extremely significantly increased (P＜0.01),and the content of MDA in the supernatant and the fluorescence intensity of cells ROS were extremely significantly increased (P＜0.01).The expression of Keap1 ,Nrf2 and HO-1 genes mRNA were extremely significantly reduced (P＜0.01),the content of antioxidant enzymes SOD and GSH-Px were extremely significantly decreased (P＜0.01).Compared with LPS group,in ELP group,the mRNA expression of IL-6 gene was extremely significantly reduced (P＜0.01),the mRNA expression of TNF-α gene was significantly decreased (P＜0.05).The content of MDA in the supernatant and the fluorescence intensity of cells ROS were extremely significant decreased (P＜0.01).The mRNA expression of Keap1 ,Nrf2 and HO-1 genes were significantly up-regulated (P＜0.01),the content of GSH-Px was increased extremely significant (P＜0.01).The content of SOD was also significantly increased (P＜0.05).【Conclusion】 EH aqueous extract played an anti-inflammatory and antioxidant role in IPEC-J2 cells by activating the Keap1/Nrf2 signal,increasing the contents of antioxidant enzymes SOD and GSH-Px, activating the expression of downstream target genes HO-1,and 10 μg/mL EH water extract had the best effect.

Key words: Euphorbiae humifusae aqueous extract; IPEC-J2 cells; LPS; oxidative stress; Keap1/Nrf2 signaling

CLC Number:

S852.2

CHEN Jin, XIE Jinghao, HAN Yingqian, YANG Yanbin, WANG Yueying, LI Heping. Anti-inflammatory and Antioxidant Effects of Euphorbiae humifusae Aqueous Extract in IPEC-J2 Cells[J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(2): 704-712.

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Anti-inflammatory and Antioxidant Effects of Euphorbiae humifusae Aqueous Extract in IPEC-J2 Cells

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