L Liu1,2, Y Li1,2, Y He1,2, Z Wang1,2, H Zhao1,2, X Jin1,2, D Shi1,2, X Wang1,2. 1. State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, China. 2. Key Laboratory of Preventive Veterinary Medicine in Hubei Province, Hubei Provincial Institute of Veterinary Drug Control, Wuhan, China.
Abstract
AIMS: Zona occludens-1 (ZO-1) is a key regulatory tight junction protein that plays an important role in maintaining gastrointestinal health. In this study, we investigated the protective effect and regulation mechanism of the probiotic Enterococcus faecium HDRsEf1 on tight junction protein ZO-1 at the cellular and molecular levels. METHODS AND RESULTS: We established lipopolysaccharide (LPS)-induced intestinal epithelial cell injury model and detected the protective effect of HDRsEf1 on ZO-1 in IPEC-J2 cells by real-time polymerase chain reaction and Western blot. The results showed that HDRsEf1 inhibited the downregulation of ZO-1 expression induced by LPS. HDRsEf1 stabilized the destruction of the ZO-1 structure caused by LPS in an immunofluorescence assay. Through gene overexpression and siRNA interference tests, we found that transcription factor activator protein 1 (AP-1) inhibited the level of ZO-1 expression. Silencing experiment further supported that the protective effect of HDRSEF1 might be mediated by suppression of LPS-provoked activation of apoptosis signal-regulating kinase 1 (ASK1)/mitogen-activated protein kinase kinase 7 (MKK7)/c-Jun N-terminal kinase (JNK) signalling pathways. In addition, HDRsEf1 could stabilize ZO-1 expression by increasing toll-like receptor 2 (TLR2) expression and competing with LPS for the TLR4 binding site. More interestingly, we also found that HDRsEf1 could stabilize ZO-1 expression through inhibiting the production of tumour necrosis factor-α (TNF-α) induced by LPS. CONCLUSIONS: HDRsEf1 could protect the IPEC-J2 cell against LPS induced downregulation of ZO-1 expression by inhibiting the activation of TLR2/4-mediated JNK-AP-1 and signalling cascade and the production of TNF-α. SIGNIFICANCE AND IMPACT OF THE STUDY: This study can provide a theoretical basis for probiotics to regulate the expression of intestinal tight junction proteins, and supply technical support for probiotics to prevent and treat animal intestinal infectious diseases.
AIMS: Zona occludens-1 (ZO-1) is a key regulatory tight junction protein that plays an important role in maintaining gastrointestinal health. In this study, we investigated the protective effect and regulation mechanism of the probiotic Enterococcus faecium HDRsEf1 on tight junction protein ZO-1 at the cellular and molecular levels. METHODS AND RESULTS: We established lipopolysaccharide (LPS)-induced intestinal epithelial cell injury model and detected the protective effect of HDRsEf1 on ZO-1 in IPEC-J2 cells by real-time polymerase chain reaction and Western blot. The results showed that HDRsEf1 inhibited the downregulation of ZO-1 expression induced by LPS. HDRsEf1 stabilized the destruction of the ZO-1 structure caused by LPS in an immunofluorescence assay. Through gene overexpression and siRNA interference tests, we found that transcription factor activator protein 1 (AP-1) inhibited the level of ZO-1 expression. Silencing experiment further supported that the protective effect of HDRSEF1 might be mediated by suppression of LPS-provoked activation of apoptosis signal-regulating kinase 1 (ASK1)/mitogen-activated protein kinase kinase 7 (MKK7)/c-Jun N-terminal kinase (JNK) signalling pathways. In addition, HDRsEf1 could stabilize ZO-1 expression by increasing toll-like receptor 2 (TLR2) expression and competing with LPS for the TLR4 binding site. More interestingly, we also found that HDRsEf1 could stabilize ZO-1 expression through inhibiting the production of tumour necrosis factor-α (TNF-α) induced by LPS. CONCLUSIONS: HDRsEf1 could protect the IPEC-J2 cell against LPS induced downregulation of ZO-1 expression by inhibiting the activation of TLR2/4-mediated JNK-AP-1 and signalling cascade and the production of TNF-α. SIGNIFICANCE AND IMPACT OF THE STUDY: This study can provide a theoretical basis for probiotics to regulate the expression of intestinal tight junction proteins, and supply technical support for probiotics to prevent and treat animal intestinal infectious diseases.