Background:Enterotoxigenic Escherichia coli(ETEC)K88 commonly colonize in the small intestine and keep releasing enterotoxins to impair the intestinal barrier function and trigger inflammatory reaction.Although Lactob...Background:Enterotoxigenic Escherichia coli(ETEC)K88 commonly colonize in the small intestine and keep releasing enterotoxins to impair the intestinal barrier function and trigger inflammatory reaction.Although Lactobacillus salivarius(L.salivarius)has been reported to enhance intestinal health,it remains to be seen whether there is a functional role of L.salivarius in intestinal inflammatory response in intestinal porcine epithelial cell line(IPEC-J2)when stimulated with ETEC K88.In the present study,IPEC-J2 cells were first treated with L.salivarius followed by the stimulation of ETEC K88 for distinct time period.ETEC K88 adherent status,pattern recognition receptors(PRRs)mRNA,mitogen-activated protein kinase(MAPK)and nuclear factor-κB(NF-κB)activation,the release of pro-inflammation cytokines and cell integrity were examined.Results:Aside from an inhibited adhesion of ETEC K88 to IPEC-J2 cells,L.salivarius was capable of remarkably attenuating the expression levels of interleukin(IL)-1β,tumor necrosis factor-α(TNF-α),IL-8,Toll-like receptor(TLR)4,nucleotide-binding oligomerization domain(NOD)-like receptor pyrin domain-containing protein(NLRP)3 and NLRP6.This alternation was accompanied by a significantly decreased phosphorylation of p38 MAPK and p65 NF-κB during ETEC K88 infection with L.salivarius pretreatment.Western blot analysis revealed that L.salivarius increased the expression levels of zona occludens 1(ZO-1)and occludin(P<0.05)in ETEC K88-infected IPEC-J2 cells.Compared with ETEC K88-infected groups,the addition of L.salivarius as well as extra inhibitors for MAPKs and NF-κB to ETEC K88-infected IPEC-J2 cells had the capability to reduce pro-inflammatory cytokines.Conclusions:Collectively,our results suggest that L.salivarius might reduce inflammation-related cytokines through attenuating phosphorylation of p38 MAPK and blocking the NF-κB signaling pathways.Besides,L.salivarius displayed a potency in the enhancement of IPEC-J2 cell integrity.展开更多
Background:Necroptosis and pyroptosis are newly identified forms of programmed cell death,which play a vital role in development of many gastrointestinal disorders.Although plant polyphenols have been reported to prot...Background:Necroptosis and pyroptosis are newly identified forms of programmed cell death,which play a vital role in development of many gastrointestinal disorders.Although plant polyphenols have been reported to protect intestinal health,it is still unclear whether there is a beneficial role of plant polyphenols in modulating necroptosis and pyroptosis in intestinal porcine epithelial cell line(IPEC-1)infected with enterotoxigenic Escherichia coli(ETEC)K88.This research was conducted to explore whether plant polyphenols including protocatechuic acid(PCA)and quercetin(Que),attenuated inflammation and injury of IPEC-1 caused by ETEC K88 through regulating necroptosis and pyroptosis signaling pathways.Methods:IPEC-1 cells were treated with PCA(40μmol/L)or Que(10μmol/L)in the presence or absence of ETEC K88.Results:PCA and Que decreased ETEC K88 adhesion and endotoxin level(P<0.05)in cell supernatant.PCA and Que increased cell number(P<0.001)and decreased lactate dehydrogenases(LDH)activity(P<0.05)in cell supernatant after ETEC infection.PCA and Que improved transepithelial electrical resistance(TEER)(P<0.001)and reduced fluorescein isothiocyanate-labeled dextran(FD4)flux(P<0.001),and enhanced membrane protein abundance of occludin,claudin-1 and ZO-1(P<0.05),and rescued distribution of these tight junction proteins(P<0.05)after ETEC infection.PCA and Que also declined cell necrosis ratio(P<0.05).PCA and Que reduced mRNA abundance and concentration of tumor necrosis factor-α(TNF-α),interleukin(IL)-6 and IL-8(P<0.001),and down-regulated gene expression of toll-like receptors 4(TLR4)and its downstream signals(P<0.001)after ETEC infection.PCA and Que down-regulated protein abundance of total receptor interacting protein kinase 1(t-RIP1),phosphorylated-RIP1(p-RIP1),p-RIP1/t-RIP1,t-RIP3,p-RIP3,mixed lineage kinase domain-like protein(MLKL),p-MLKL,dynamin-related protein 1(DRP1),phosphoglycerate mutase 5(PGAM5)and high mobility group box 1(HMGB1)(P<0.05)after ETEC infection.Moreover,PCA and Que reduced protein abundance of nod-like receptor protein 3(NLRP3),nod-like receptors family CARD domain-containing protein 4(NLRC4),apoptosis-associated speck-like protein containing a CARD(ASC),gasdermin D(GSDMD)and caspase-1(P<0.05)after ETEC infection.Conclusions:In general,our data suggest that PCA and Que are capable of attenuating ETEC-caused intestinal inflammation and damage via inhibiting necroptosis and pyroptosis signaling pathways.展开更多
基金funded by the Tianjin Natural Science Foundation(18JCYBJC30000)the National Natural Science Foundation of China(31702147)the Tianjin“131”Innovative Talents Team(20180338).
文摘Background:Enterotoxigenic Escherichia coli(ETEC)K88 commonly colonize in the small intestine and keep releasing enterotoxins to impair the intestinal barrier function and trigger inflammatory reaction.Although Lactobacillus salivarius(L.salivarius)has been reported to enhance intestinal health,it remains to be seen whether there is a functional role of L.salivarius in intestinal inflammatory response in intestinal porcine epithelial cell line(IPEC-J2)when stimulated with ETEC K88.In the present study,IPEC-J2 cells were first treated with L.salivarius followed by the stimulation of ETEC K88 for distinct time period.ETEC K88 adherent status,pattern recognition receptors(PRRs)mRNA,mitogen-activated protein kinase(MAPK)and nuclear factor-κB(NF-κB)activation,the release of pro-inflammation cytokines and cell integrity were examined.Results:Aside from an inhibited adhesion of ETEC K88 to IPEC-J2 cells,L.salivarius was capable of remarkably attenuating the expression levels of interleukin(IL)-1β,tumor necrosis factor-α(TNF-α),IL-8,Toll-like receptor(TLR)4,nucleotide-binding oligomerization domain(NOD)-like receptor pyrin domain-containing protein(NLRP)3 and NLRP6.This alternation was accompanied by a significantly decreased phosphorylation of p38 MAPK and p65 NF-κB during ETEC K88 infection with L.salivarius pretreatment.Western blot analysis revealed that L.salivarius increased the expression levels of zona occludens 1(ZO-1)and occludin(P<0.05)in ETEC K88-infected IPEC-J2 cells.Compared with ETEC K88-infected groups,the addition of L.salivarius as well as extra inhibitors for MAPKs and NF-κB to ETEC K88-infected IPEC-J2 cells had the capability to reduce pro-inflammatory cytokines.Conclusions:Collectively,our results suggest that L.salivarius might reduce inflammation-related cytokines through attenuating phosphorylation of p38 MAPK and blocking the NF-κB signaling pathways.Besides,L.salivarius displayed a potency in the enhancement of IPEC-J2 cell integrity.
基金provided by National Key R&D Program of China(2022YFD1300403)National Natural Science Foundation of China(No.U22A20517,32272906,and 31802070)Wuhan Science and Technology Bureau(No.2022020801010391)。
文摘Background:Necroptosis and pyroptosis are newly identified forms of programmed cell death,which play a vital role in development of many gastrointestinal disorders.Although plant polyphenols have been reported to protect intestinal health,it is still unclear whether there is a beneficial role of plant polyphenols in modulating necroptosis and pyroptosis in intestinal porcine epithelial cell line(IPEC-1)infected with enterotoxigenic Escherichia coli(ETEC)K88.This research was conducted to explore whether plant polyphenols including protocatechuic acid(PCA)and quercetin(Que),attenuated inflammation and injury of IPEC-1 caused by ETEC K88 through regulating necroptosis and pyroptosis signaling pathways.Methods:IPEC-1 cells were treated with PCA(40μmol/L)or Que(10μmol/L)in the presence or absence of ETEC K88.Results:PCA and Que decreased ETEC K88 adhesion and endotoxin level(P<0.05)in cell supernatant.PCA and Que increased cell number(P<0.001)and decreased lactate dehydrogenases(LDH)activity(P<0.05)in cell supernatant after ETEC infection.PCA and Que improved transepithelial electrical resistance(TEER)(P<0.001)and reduced fluorescein isothiocyanate-labeled dextran(FD4)flux(P<0.001),and enhanced membrane protein abundance of occludin,claudin-1 and ZO-1(P<0.05),and rescued distribution of these tight junction proteins(P<0.05)after ETEC infection.PCA and Que also declined cell necrosis ratio(P<0.05).PCA and Que reduced mRNA abundance and concentration of tumor necrosis factor-α(TNF-α),interleukin(IL)-6 and IL-8(P<0.001),and down-regulated gene expression of toll-like receptors 4(TLR4)and its downstream signals(P<0.001)after ETEC infection.PCA and Que down-regulated protein abundance of total receptor interacting protein kinase 1(t-RIP1),phosphorylated-RIP1(p-RIP1),p-RIP1/t-RIP1,t-RIP3,p-RIP3,mixed lineage kinase domain-like protein(MLKL),p-MLKL,dynamin-related protein 1(DRP1),phosphoglycerate mutase 5(PGAM5)and high mobility group box 1(HMGB1)(P<0.05)after ETEC infection.Moreover,PCA and Que reduced protein abundance of nod-like receptor protein 3(NLRP3),nod-like receptors family CARD domain-containing protein 4(NLRC4),apoptosis-associated speck-like protein containing a CARD(ASC),gasdermin D(GSDMD)and caspase-1(P<0.05)after ETEC infection.Conclusions:In general,our data suggest that PCA and Que are capable of attenuating ETEC-caused intestinal inflammation and damage via inhibiting necroptosis and pyroptosis signaling pathways.
