Clostridium perfringens beta2 toxin induced in vitro oxidative damage and its toxic assessment in porcine small intestinal epithelial cell lines

Luo, Ruirui; Yang, Qiaoli; Huang, Xiaoyu; Yan, Zunqiang; Gao, Xiaoli; Wang, Wei; Xie, Kaihui; Wang, Pengfei; Gun, Shuangbao

doi:10.1016/j.gene.2020.144999

Cited by 27 publications

(25 citation statements)

References 31 publications

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“…Tight junction proteins, including claudin proteins, zonula occludens proteins and occludin, are the main components providing barrier function in epithelial cells and maintain epithelial permeability and integrity [ 39 ]. Oxidative stress-induced alterations in tight junction protein levels can reduce the viability of porcine intestinal epithelial cells [ 40 ]. In the present study, we found that H 2 O 2 exposure decreased the expression levels of ZO-1, ZO-2, occludin and claudin-1 in IPEC-J2 cells and decreased the viability of these cells, consistent with previous research [ 41 ].…”

Section: Discussionmentioning

confidence: 99%

Pyrroloquinoline quinone regulates the redox status in vitro and in vivo of weaned pigs via the Nrf2/HO-1 pathway

Huang

Fan

Han

et al. 2021

J Animal Sci Biotechnol

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Background Oxidative stress is a main cause of piglet gut damage and diarrhea. Pyrroloquinoline quinone (PQQ), is a novel redox cofactor with antioxidant properties. However, the effect and mechanism that PQQ supplementation decreases oxidative injury in weaned pigs is not understood. Therefore, the aim of this study is to confirm the effect of PQQ on regulating redox status in weaned pigs and the mechanism for antioxidant function by porcine intestinal epithelial cell line (IPEC-J2) challenged with H2O2. Results Experiment 1, 144 Duroc × Landrace × Yorkshire pigs (weaned at 28 d) were allocated to four groups: received a basal diet (control) and diets supplemented with 0.15%, 0.30% and 0.45% PQQ, respectively. On d 28, growth performance, diarrhea incidence and redox factors were measured. Experiment 2, IPEC-J2 were treated with or without PQQ in the presence or absence of H2O2 for indicated time points. Experiment 3, IPEC-J2 were transfected with or without Nrf2 siRNA, then treated according to Experiment 2. The cell viability, redox factors, protein of tight junctions and Nrf2 pathway were determined. In vivo, PQQ supplementation demonstrated dose-related improvements in average daily gain, and gain to feed ratio (Linear P < 0.05). During d 0–28, compared to controls, 0.45% PQQ supplementation for pigs decreased diarrhea incidence and MDA content in liver and jejunum, and increased concentration of SOD in liver; 0.3% PQQ supplementation decreased ileal and liver MDA concentration; and 0.15% PQQ supplementation decreased ileal MDA concentration (P < 0.05). In vitro, compared to cells cultured with H2O2, pre-treatment with PQQ increased cell viability, tight junction proteins expression including ZO-1, ZO-2, Occludin and Claudin-1; and decreased ROS concentration and level of Caspase-3 (P < 0.05); as well as upregulated the ratio of Bcl-2 to Bax and protein expression of nuclear Nrf2, HO-1. Notably, Nrf2 knockdown by transfection with Nrf2 siRNA largely abrogated the positive effects of PQQ pretreatment on H2O2-induced intracellular changes. Conclusions PQQ administration attenuated oxidative stress in weaned pigs which is associated with activation of Nrf2/HO-1 pathway.

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Section: Discussionmentioning

confidence: 99%

Pyrroloquinoline quinone regulates the redox status in vitro and in vivo of weaned pigs via the Nrf2/HO-1 pathway

Huang

Fan

Han

et al. 2021

J Animal Sci Biotechnol

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“…The extraction and purification of CPB2 toxin were performed according to the method described by Luo et al (2020) . Briefly, a recombinant plasmid containing the CPB2 gene was successfully constructed and transformed into E. coli BL21 competent cells.…”

Section: Methodsmentioning

confidence: 99%

“…IPEC-J2 cells were seeded in 6-well plates at a density of 1×10 5 cells/mL and cultured overnight (24 h). Then, cells in three wells were each treated with 20 μg/mL CPB2 (treatment group) while cells in the other three wells were not exposed to toxin (control group) ( Gao et al, 2020 ; Luo et al, 2020 ). Total RNA was extracted by using TRIzol reagent according to the manufacturer’s instructions (Invitrogen, Carlsbad, United States), and the m 6 A content was detected with an m 6 A RNA methylation quantification kit (Epigentek, Farmingdale, United States).…”

Section: Methodsmentioning

confidence: 99%

Comprehensive Analysis of Transcriptome-wide m6A Methylome Upon Clostridium perfringens Beta2 Toxin Exposure in Porcine Intestinal Epithelial Cells by m6A Sequencing

Zhang

Yang

et al. 2021

Front. Genet.

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Piglet diarrhea is a swine disease responsible for serious economic impacts in the pig industry. Clostridium perfringens beta2 toxin (CPB2), which is a major toxin of C. perfringens type C, may cause intestinal diseases in many domestic animals. N6-methyladenosine (m6A) RNA methylation plays critical roles in many immune and inflammatory diseases in livestock and other animals. However, the role of m6A methylation in porcine intestinal epithelial (IPEC-J2) cells exposed to CPB2 has not been studied. To address this issue, we treated IPEC-J2 cells with CPB2 toxin and then quantified methylation-related enzyme expression by RT-qPCR and assessed the m6A methylation status of the samples by colorimetric N6-methyladenosine quantification. The results showed that the methylation enzymes changed to varying degrees while the m6A methylation level increased (p < 0.01). On this basis, we performed N6-methyladenosine sequencing (m6A-seq) and RNA sequencing (RNA-seq) to examine the detailed m6A modifications and gene expression of the IPEC-J2 cells following CPB2 toxin exposure. Our results indicated that 1,448 m6A modification sites, including 437 up-regulated and 1,011 down-regulated, differed significantly between CPB2 toxin exposed cells and non-exposed cells (p < 0.05). KEGG pathway analysis results showed that m6A peaks up-regulated genes (n = 394) were mainly enriched in cancer, Cushing syndrome and Wnt signaling pathways, while m6A peaks down-regulated genes (n = 920) were mainly associated with apoptosis, small cell lung cancer, and the herpes simplex virus 1 infection signaling pathway. Furthermore, gene expression (RNA-seq data) analysis identified 1,636 differentially expressed genes (DEGs), of which 1,094 were up-regulated and 542 were down-regulated in the toxin exposed group compared with the control group. In addition, the down-regulated genes were involved in the Hippo and Wnt signaling pathways. Interestingly, the combined results of m6A-seq and RNA-seq identified genes with up-regulated m6A peaks but with down-regulated expression, here referred to as “hyper-down” genes (n = 18), which were mainly enriched in the Wnt signaling pathway. Therefore, we speculate that the genes in the Wnt signaling pathway may be modified by m6A methylation in CPB2-induced IPEC-J2 cells. These findings provide new insights enabling further exploration of the mechanisms underlying piglet diarrhea caused by CPB2 toxin.

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“…Moreover, the dysregulation of tight junction protein integrity enhances intestinal barrier permeability [77]. It is worth noting that virulence factors, which are produced by C. perfringens, could effectively impair tight junctions [15,78]. In this study, L. plantarum Lac16 preincubation attenuated C. perfringens-induced disruption of mucus production.…”

Section: Discussionmentioning

confidence: 60%

“…It is worth mentioning that antibiotics play a critical role in the prevention and clinical treatment of diseases caused by C. perfringens [13,14]. However, under the situation of feed antibiotics prohibition, C. perfringens infection has become an important problem, hindering the development of the pig industry [13,15,16]. Nowadays, many feed additives are used in animal husbandry to prevent C. perfringens infection, such as probiotics, prebiotics, symbiotics, essential oils, and herbs [17][18][19][20].…”

Section: Introductionmentioning

confidence: 99%

Protective Effects of Lactobacillus plantarum Lac16 on Clostridium perfringens Infection-Associated Injury in IPEC-J2 Cells

Zhou¹,

Wang²,

Wang³

et al. 2021

IJMS

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Clostridium perfringens (C. perfringens) causes intestinal injury through overgrowth and the secretion of multiple toxins, leading to diarrhea and necrotic enteritis in animals, including pigs, chickens, and sheep. This study aimed to investigate the protective effects of Lactobacillus plantarum (L. plantarum) Lac16 on C. perfringens infection-associated injury in intestinal porcine epithelial cell line (IPEC-J2). The results showed that L. plantarum Lac16 significantly inhibited the growth of C. perfringens, which was accompanied by a decrease in pH levels. In addition, L. plantarum Lac16 significantly elevated the mRNA expression levels of host defense peptides (HDPs) in IPEC-J2 cells, decreased the adhesion of C. perfringens to IPEC-J2 cells, and attenuated C. perfringens-induced cellular cytotoxicity and intestinal barrier damage. Furthermore, L. plantarum Lac16 significantly suppressed C. perfringens-induced gene expressions of proinflammatory cytokines and pattern recognition receptors (PRRs) in IPEC-J2 cells. Moreover, L. plantarum Lac16 preincubation effectively inhibited the phosphorylation of p65 caused by C. perfringens infection. Collectively, probiotic L. plantarum Lac16 exerts protective effects against C. perfringens infection-associated injury in IPEC-J2 cells.

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Clostridium perfringens beta2 toxin induced in vitro oxidative damage and its toxic assessment in porcine small intestinal epithelial cell lines

Cited by 27 publications

References 31 publications

Pyrroloquinoline quinone regulates the redox status in vitro and in vivo of weaned pigs via the Nrf2/HO-1 pathway

Pyrroloquinoline quinone regulates the redox status in vitro and in vivo of weaned pigs via the Nrf2/HO-1 pathway

Comprehensive Analysis of Transcriptome-wide m6A Methylome Upon Clostridium perfringens Beta2 Toxin Exposure in Porcine Intestinal Epithelial Cells by m6A Sequencing

Protective Effects of Lactobacillus plantarum Lac16 on Clostridium perfringens Infection-Associated Injury in IPEC-J2 Cells

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