The apoptotic effects of soybean agglutinin were induced through three different signal pathways by down-regulating cytoskeleton proteins in IPEC-J2 cells

Abstract

Soybean agglutinin (SBA) is a main anti-nutritional factor in soybean. SBA exhibits its anti-nutritional functions by binding to intestinal epithelial cells. Keratin8 (KRT8), Keratin18 (KRT18) and Actin (ACTA) are the representative SBA-specific binding proteins. Such cytoskeletal proteins act a crucial role in different cell activities. However, limited reports reveal what the signal transduction pathway of apoptosis caused by SBA when binding to KRT8, KRT18 and ACTA. We aimed to evaluate the effects of SBA on cell apoptosis and the expression of the cytoskeletal protein (KRT8, KRT18 and ACTA), reveal the roles of these cytoskeletal proteins or their combinations on SBA-induced cell apoptosis in IPEC-J2 cell line, evaluate the influences of SBA on the mitochondria, endoplasmic reticulum stress and death receptor-mediated apoptosis signal pathway and to show the roles of KRT8, KRT18 and ACTA in different apoptosis signal pathways induced by SBA. The results showed that SBA induced the IPEC-J2 cell apoptosis and decreased the mRNA expression of KRT8, KRT18 and ACTA (p < 0.05). The degree of effect of three cytoskeleton proteins on cell apoptosis was ACTA > KRT8 > KRT18. The roles of these three cytoskeletal proteins on IPEC-J2 apoptotic rates had a certain accumulation effect. SBA up-regulated mitochondrial fission variant protein (FIS1) and fusion protein (Mfn2) promoted CytC and AIF in mitochondria to enter the cytoplasm, activated caspase-9 and caspase-3, damaged or declined mitochondrial function and reduced ATP synthesis (p < 0.05). Also, SBA up-regulated the expression of GRP78, XBP-1, eIF2α, p-eIF2α and CHOP (p < 0.05), down-regulated the expression level of ASK1 protein (p < 0.05). SBA led to the recruitment of FADD to the cytoplasmic membrane and increased the expression of FasL, resulting in caspase-8 processing. SBA up-regulated the expression level of Bax protein and decreased cytosolic Bcl-2 and Bid (p < 0.05). In addition, there was a significant negative correlation between the gene expression of cytoskeleton proteins and apoptosis, as well as the expression of key proteins of apoptosis-related signal transduction pathways. In conclusion, SBA induced the activation of the mitochondria, endoplasmic reticulum stress and the death receptor-mediated apoptosis signal pathway and the crosstalk between them. The effect of SBA on these three pathways was mainly exhibited via down-regulation of the mRNA expression of the three cytoskeletal expressions. This study elucidates the molecular mechanism and signaling pathway of SBA that lead to apoptosis from the perspective of cell biology and molecular biology and provides a new perspective on the toxicity mechanism of other food-derived anti-nutrients, medical gastrointestinal health and related cancer treatment.

Lectin-mediated, time-efficient, and high-yield sorting of different morphologically intact nephron segments

“…These harmful effects include mitogenic stimulation of different organs, induction of apoptosis and autophagy, and immunomodulatory functions [ 18 ]. For example, SBA was described to dose-dependently increase reactive oxygen species in HeLa cells [ 24 ], inhibit plasma membrane repair of damaged gut epithelial cells [ 19 ], or induce apoptosis in the naïve porcine jejunal cell line IPEC‐J2 through mitochondrial and ER stress pathways and the downregulation of cytoskeleton proteins [ 23 ]. Yet, in comparison to these studies, we do not expect such effects with our method, which is due to the low concentration of Flaggs (1–2 µg/ml vs. 20–2000 µg/ml), the low temperature during incubation and sorting (4 °C vs. 37 °C), and finally, the relatively short time frame (less than 2 h vs. 24-h incubation periods).…”

A Single-Cell Landscape of Spermioteleosis in Mice and Pigs

“…As shown in the gene expression map, the basal cell marker gene Krt8 (KRT8) was also highly expressed in the principal cells of both mice and pigs. KRT8 plays a role in maintaining cellular structural integrity and functions in signal transduction and cellular differentiation [57,58]. Basal cells can regulate electrolyte and water transport via principal cells [59].…”