Research on IPEC-J2 Cell Injury Induced by 11S Globulin via Signal Pathway of Nuclear Factor-&kappa;B, Inducible Nitric Oxide Synthase, c-Jun N-Terminal Kinase and p38 Mitogen Activated Protein Kinase

WANG Lei; SUN Zhifeng; PENG Chenglu; DING Hongyan; WANG Zhi; LI Siting; WAGN Chengzhi; LI Yu; WANG Xichun; WU Jinjie

doi:10.3969/j.issn.1006-267x.2021.03.047

2021 , Vol. 33 >Issue 3: 1663 - 1674

DOI: https://doi.org/10.3969/j.issn.1006-267x.2021.03.047

Research on IPEC-J2 Cell Injury Induced by 11S Globulin via Signal Pathway of Nuclear Factor-κB, Inducible Nitric Oxide Synthase, c-Jun N-Terminal Kinase and p38 Mitogen Activated Protein Kinase

WANG Lei ,
SUN Zhifeng ,
PENG Chenglu ,
DING Hongyan ,
WANG Zhi ,
LI Siting ,
WAGN Chengzhi ,
LI Yu ,
WANG Xichun ,
WU Jinjie

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College of Animal Science and Technology, Anhui Agricultural University, Hefei 230061, China

Received date: 2020-08-09

Online published: 2021-03-18

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Abstract

The present study aimed to analyze the discrepancies in 11S glycinin induced porcine intestinal epithelial cell (IPEC-J2 cell) injury via signal pathways of nuclear factor-κB (NF-κB), inducible nitric oxide synthase (iNOS), c-Jun N-terminal kinase (JNK) and p38 mitogen activated protein kinase (p38 MAPK) using cell culture technology in vitro. The experiment was randomly divided into 6 groups: group A (control group) was no added; group B was added 5 mg/mL 11S glycinin; groups C, D, E and F were pretreated with 1 μmol/L NF-κB inhibitor of pyrrolidine dithiocarbamte (PDTC), iNOS inhibitor of Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME), JNK inhibitor of SP600125) and p38 MAPK inhibitor of SB202190, respectively, and then were added 5 mg/mL 11S glycinin. After 24 hours culture, cell viability was detected by CCK-8. The contents of nitric oxide (NO), tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ) and interleukin-10 (IL-10) were measured by enzyme-linked immunosorbent assay (ELISA) method. The morphology of cells and nuclei were observed by hematoxylin-eosin (HE)staining. The ultrastructure of cells was observed by transmission electron microscope. The mRNA relative expression levels of NF-κB, iNOS, JNK and p38 MAPK were determined by real-time quantitative PCR. The protein expression levels of NF-κB, iNOS, JNK and p38 MAPK were detected by Western blot. The results showed as follows: 1) compared with group A, the cell viability of group B was significantly decreased (P<0.01); compared with group B, the cell viability of groups C, D, E and F were significantly increased (P<0.01), and the cell viability of group C was significantly higher than that of groups D, E and F (P<0.05). 2) Compared with group A, the contents of TNF-α, INF-γ and NO of group B were significantly increased (P<0.01), but the IL-10 content was significantly decreased (P<0.01); compared with group B, the contents of TNF-α, INF-γ and NO of groups C, D, E and F were significantly decreased (P<0.01), but the IL-10 content was significantly increased (P<0.01). 3) Compared with group A, the protein expression levels and the mRNA relative expression levels of NF-κB, iNOS, JNK and p38 MAPK of group B were significantly increased (P<0.01). 4) Compared with group B, the mRNA relative expression levels of NF-κB, iNOS, JNK and p38 MAPK of group C and F were significantly decreased (P<0.05 or P<0.01), the mRNA relative expression levels of NF-κB, iNOS and JNK of group D were significantly decreased (P<0.01), the mRNA relative expression levels of NF-κB, JNK and p38 MAPK of group E were significantly decreased (P<0.05 or P<0.01). 5) Compared with group B, the protein expression levels of NF-κB, iNOS, JNK and p38 MAPK of group C ,E and F were significantly decreased (P<0.01), the protein expression levels of NF-κB, iNOS and JNK of group D were significantly decreased (P<0.01). 6) Observation of the HE staining and transmission electron microscope showed cells damage, cytoplasmic vacuolation and aggregation of nuclear chromatin in group B, the injury of cells were inhibited in groups C, D, E and F, and the integrity of cell structure and morphology in group C was better than that in groups D, E and F. In conclusion, the 11S glycinin can induce IPEC-J2 cell damage via JNK/p38 MAPK/NF-κB/iNOS signaling pathways, and the NF-κB signaling pathway plays a critical role in the process of inducing cell damage.

Key words： glycinin; IPEC-J2 cells; signal pathway; cell damage

Cite this article

WANG Lei , SUN Zhifeng , PENG Chenglu , DING Hongyan , WANG Zhi , LI Siting , WAGN Chengzhi , LI Yu , WANG Xichun , WU Jinjie . Research on IPEC-J2 Cell Injury Induced by 11S Globulin via Signal Pathway of Nuclear Factor-κB, Inducible Nitric Oxide Synthase, c-Jun N-Terminal Kinase and p38 Mitogen Activated Protein Kinase[J]. Chinese Journal of Animal Nutrition, 2021 , 33(3) : 1663 -1674 . DOI: 10.3969/j.issn.1006-267x.2021.03.047

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