AUTHOR=Wu Qiong , Zhu Yao-Hong , Xu Jin , Liu Xiao , Duan Cong , Wang Mei-Jun , Wang Jiu-Feng 

TITLE=Lactobacillus rhamnosus GR-1 Ameliorates Escherichia coli-Induced Activation of NLRP3 and NLRC4 Inflammasomes With Differential Requirement for ASC

JOURNAL=Frontiers in Microbiology

VOLUME=9

YEAR=2018

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2018.01661

DOI=10.3389/fmicb.2018.01661

ISSN=1664-302X

ABSTRACT=<p><italic>Escherichia coli</italic> is a common cause of mastitis in dairy cows. The adaptor protein apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) synergizes with caspase-1 to regulate inflammasome activation during pathogen infection. Here, the <italic>ASC</italic> gene was knocked out in bovine mammary epithelial (MAC-T) cells using clustered, regularly interspaced, short palindromic repeat (CRISPR)/CRISPR-associated (Cas)-9 technology. MAC-T cells were pre-incubated with and without <italic>Lactobacillus rhamnosus</italic> GR-1 and then exposed to <italic>E. coli</italic>. Western blot analysis demonstrated increased expression of NLRP3 and NLRC4 following <italic>E. coli</italic> infection, but this increase was attenuated by pre-incubation with <italic>L. rhamnosus</italic> GR-1, regardless of <italic>ASC</italic> knockout. Western blot and immunofluorescence analyses revealed that pre-incubation with <italic>L. rhamnosus</italic> GR-1 decreased <italic>E. coli-</italic>induced caspase-1 activation at 6 h after <italic>E. coli</italic> infection, as also observed in <italic>ASC</italic>-knockout MAC-T cells. The <italic>E. coli-</italic>induced increase in <italic>caspase-4</italic> mRNA expression was inhibited by pre-incubation with <italic>L. rhamnosus</italic> GR-1. <italic>ASC</italic> knockout diminished, but did not completely prevent, increased production of IL-1β and IL-18 and cell pyroptosis associated with <italic>E. coli</italic> infection, whereas pre-incubation with <italic>L. rhamnosus</italic> GR-1 inhibited this increase. Our data indicate that <italic>L. rhamnosus</italic> GR-1 suppresses activation of ASC-dependent NLRP3 and NLRC4 inflammasomes and production of downstream IL-lβ and IL-18 during <italic>E. coli</italic> infection. <italic>L. rhamnosus</italic> GR-1 also inhibited <italic>E. coli</italic>-induced cell pyroptosis, in part through attenuation of NLRC4 and non-canonical caspase-4 activation independently of ASC.</p>