β-Conglycinin-Induced Intestinal Porcine Epithelial Cell Damage via the Nuclear Factor κB/Mitogen-Activated Protein Kinase Signaling Pathway

Abstract

Soybean allergy is a serious health risk to humans and animals; β-conglycinin is the primary antigenic protein in soybean. Intestinal porcine epithelial (IPEC-J2) cells were used as an in vitro physiological model of the intestinal epithelium to study the effects of different concentrations of soybean antigen protein β-conglycinin to identify the involved signaling pathways. The cells were divided into eight groups and either untreated or treated with different concentrations of β-conglycinin, pyrrolidine dithiocarbamate (PDTC), N ω -nitro-L-arginine methyl ester hydrochloride (L-NAME), SP600125, and SB202190 either alone or in combination. The cells were incubated with 1, 5, and 10 mg•mL −1 β-conglycinin or 5 mg•mL −1 β-conglycinin and 1 μmol•L −1 nuclear factor κB (NF-κB) inhibitor (PDTC), inducible nitric oxide synthase inhibitor (L-NAME), c-Jun N-terminal kinase (JNK) inhibitor (SP600125), and p38 inhibitor (SB202190) for 24 h, separately; controls were left untreated. The mRNA, protein, and phosphorylation levels of NF-κB, p38, and JNK were higher in the treated groups than in the control group. β-Conglycinin decreased tight junction distribution, destroyed the cytoskeleton of IPEC-J2 cells, and caused cell death. After the addition of the inhibitors, β-conglycinin-induced IPEC-J2 cell damage was significantly reduced. β-Conglycinin caused damage to IPEC-J2 cells via the mitogen-activated protein kinase/NF-κB signaling pathway. The results of this study are crucial for exploring the mechanisms underlying allergic reactions caused by soybean antigen proteins.

Soybean antigen protein induces caspase-3/mitochondrion-regulated apoptosis in IPEC-J2 cells

“…Bcl-2 is an important anti-apoptotic protein (Pan et al, 2018), and several kinases, such as ERK1/2, p38-MAPK, and JNK, are known to phosphorylate Bcl-2 (Mc Gee et al, 2004;Rosini et al, 2000). Our previous research have confirmed that soybean antigenic proteins cause changes in MAPK pathway-related proteins such as p-ERK1/2, p-p38 and p-JNK (Peng et al, 2019). Bcl-2 inhibits apoptosis by regulating the release of cytochrome c (cyt-c) and other proteins from mitochondria, whereas Bax promotes cell death by permeabilizing the outer mitochondrial membrane (Dlugosz et al, 2006).…”

“…After fixation, samples were washed in sodium cacodylate (0.1 M, pH 7.4) and CaCl 2 buffer (3 mM), then post-fixed with 1% osmium tetroxide in buffer for 30 min and en bloc stained in 2% uranyl acetate overnight at 0°C. TEM experiments were carried out as described in our previous study (Peng et al, 2019).…”

Glycinin-induced porcine IPEC-J2 cells damage via the NF-κB/MAPK signaling pathway

“…TJs are generally known to create continuous cell-cell contacts, which form paracellular barriers and pores that determine their permeability and form an intercellular barrier, sealing off the intercellular space between adherent epithelial cells. ZO proteins, such as ZO-1, maintain the complex integrity of TJs primarily by linking claudins, occludin, and cytoskeletal proteins [25,27,28]. Overall, the TJ-cytoskeleton structure is an essential part of the intestinal barrier.…”

“…The severity of an allergic reaction to glycinin depends on its concentration; higher concentrations induce a more severe allergic reaction [9]. Our previous research showed that glycinin causes IPEC-J2 cell damage [25]. Therefore, we evaluated different concentrations of glycinin in this study.…”

Porcine circovirus type 2 exploits JNK-mediated disruption of tight junctions to facilitate Streptococcus suis translocation across the tracheal epithelium

“…Numerous signaling pathways have been reported to be involved in the assembly, disassembly, and maintenance of TJ; these include protein kinase C [14], Rho GTPase [15], myosin light chain kinase [16] and mitogen-activated protein kinase (MAPK) signaling pathways [17]. The MAPK pathway is able to regulate the expression of TJ in different epithelial cells, and the most extensive families of MAPK include p38, ERK1/2, and JNK [18][19][20][21].…”