cFLIPS regulates alternative NLRP3 inflammasome activation in human monocytes

The innate immune responses,including inflammasome activation,are paramount for host defense against pathogen infection.In contrast to canonical and noncanonical inflammasome activation,in this study,heat-killed gram-negative bacteria(HK bacteria)were identified as single-step stimulators of the NLRP3 inflammasome in human monocytes,and they caused a moderate amount of IL-1βto be released from cells.Time course experiments showed that this alternative inflammasome response was finished within a few hours.Further analysis showed that the intrinsically limited NLRP3 inflammasome activation response was due to the negative regulation of caspase-8 by the short isoform of cFLIP(cFLIPs),which was activated by NF-κB.In contrast,overexpressed cFLIPS,but not overexpressed cFLIPL,inhibited the activation of caspase-8 and the release of IL-1βin response to HK bacteria infection in human monocytes.Furthermore,we demonstrated that TAK1 activity mediated the expression of cFLIPs and was upstream and essential for the caspase-8 cleavage induced by HK bacteria in human monocytes.The functional specificity of cFLIPs and TAK1 revealed unique responses of human monocytes to a noninvasive pathogen,providing novel insights into an alternative regulatory pathway of NLRP3 inflammasome activation.

TAK1 Deficiency in Macrophages Increases Host Susceptibility to Leishmania Infection

Leishmania parasites mainly infect macrophages and may cause severe immunopathologies in their host,which are called leishman-iases.In the current work,we infected human and mouse macrophages in vitro with Leishmania major,an etiological agent of cu-taneous leishmaniasis,and found that inhibition or deletion of the transforming growth factorβ–activated kinase 1(TAK1)gene re-sulted in increased parasite loads.In vivo,following a challenge with L.major,mice with a macrophage-specific deletion of TAK1 showed increased clinical signs and higher parasite loads compared with wild-type controls.TAK1 deficiency in mouse macro-phages led to biased Th2 cell responses during the acute stage of infection,characterized by a decrease in interferon-γexpression,and increased expression of IL-4,IL-5 and IL-10.Finally,we found that,in the late stage of L.major infection,excessive Th2-related cytokines led to high arginase 1 expression in mouse tissues and a significant reduction of NO production both locally and system-ically,resulting in compromised control of Leishmania.These findings suggest that TAK1 plays a vital role in host resistance to Leish-mania infection.

Celastrol specifically inhibits the activation of NLRP3 inflammasome

Dear Editor,Inflammasomes are vital components of the innate immune system that survey microbial infections or sterile challenges in the cytosol(de Zoete et al.,2014).A typical inflammasome is composed of a sensor protein such as NOD like receptor(NLR)or AIM2 like receptor(ALR),the

Macrophage-derived IL-1α promotes sterile inflammation in a mouse model of acetaminophen hepatotoxicity

The metabolic intermediate of acetaminophen(APAP)can cause severe hepatocyte necrosis,which triggers aberrant immune activation of liver non-parenchymal cells(NPC).Overzealous hepatic inflammation determines the morbidity and mortality of APAP-induced liver injury(AILI).Interleukin-1 receptor(IL-1R)signaling has been shown to play a critical role in various inflammatory conditions,but its precise role and underlying mechanism in AILI remain debatable.Herein,we show that NLRP3 inflammasome activation of IL-1βis dispensable to AILI,whereas IL-1α,the other ligand of IL-1R1,accounts for hepatic injury by a lethal dose of APAP.Furthermore,Kupffer cells function as a major source of activated IL-1αin the liver,which is activated by damaged hepatocytes through TLR4/MyD88 signaling.Finally,IL-1αis able to chemoattract and activate CD11b^(+)Gr-1^(+) myeloid cells,mostly neutrophils and inflammatory monocytes,to amplify deteriorated inflammation in the lesion.Therefore,this work identifies that MyD88-dependent activation of IL-1αin Kupffer cells plays a central role in the immunopathogenesis of AILI and implicates that IL-1αis a promising therapeutic target for AILI treatment.

Human pathogenic fungus Trichophyton schoenleinii activates the NLRP3 inflammasome

The fungus Trichophyton schoenleinii(T.schoenleinii)is the causative agent of Trichophytosis and Tinea favosa of the scalp in certain regions of Eurasia and Africa.Hu-man innate immune system plays an important role in combating with various pathogens including fungi.The inflammasome is one of the most critical arms of host innate immunity,which is a protein complex controlling maturation of IL-1β.To clarify whether T.schoenleinii is able to activate the infl ammasome,we analyzed human monocytic cell line THP-1 for IL-1βproduction upon infec-tion with T.schoenleinii strain isolated from Tinea favosa patients,and rapid IL-1βsecretion from THP-1 cells was observed.Moreover,applying competitive inhibitors and gene specifi c silencing with shRNA,we found that T.sch-oenleinii induced IL-1βsecretion,ASC pyroptosome for-mation as well as caspase-1 activation were all dependent on NLRP3.Cathepsin B activity,ROS production and K+effl ux were required for the infl ammasome activation by T.schoenleinii.Our data thus reveal that the NLRP3 infl am-masome plays an important role in host defense against T.schoenleinii,and suggest that manipulating NLRP3 signaling can be a novel approach for control of diseases caused by T.schoenleinii infection.

Cellular localization of NLRP3 inflammasome

Infl ammasome is a large protein complex activated upon cellular stress or microbial infection,which triggers maturation of pro-inflammatory cytokines interleukin-1βand interleukin-18 through caspase-1 activation.Nod-like receptor family protein 3(NLRP3)is the most character-ized infl ammasome activated by various stimuli.However,the mechanism of its activation is unclear and its exact cellular localization is still unknown.We examined the potential co-localization of NLRP3 infl ammasome with mi-tochondria and seven other organelles under adenosine triphosphate,nigericin or monosodium urate stimulation in mouse peritoneal macrophages using confocal micros-copy approach.Our results revealed that the activated endogenous apoptosis-associated speck-like protein containing a CARD(ASC)pyroptosome forms in the cyto-plasm and co-localizes with NLRP3 and caspase-1,but not with any of the organelles screened.This study indicates that the ASC pyroptosome universally localizes within the cytoplasm rather than with any specifi c organelles.

IL-18通过共生菌依赖但炎症小体非依赖的方式维持肠道免疫稳态

炎症小体及其产物IL-18在肠道菌群监控和稳态维持方面发挥重要作用,其功能缺失会导致肠道菌群失衡并加速疾病进展,但其诱导的肠道菌群失衡对粘膜免疫系统的影响尚不清楚.本研究发现来自Il18^(-/-)小鼠的紊乱菌群以独立于炎症小体的方式诱导小肠免疫细胞的缺失.同窝喂养实验表明这些共生菌诱导小肠免疫细胞丢失的表型可转移至野生型(WT)小鼠,并通过RIP3-MLKL信号途径诱导免疫细胞死亡.共生菌组成分析在菌属水平上确定Ureaplasma和Parasutterella两类共生菌在Il18^(-/-)小鼠中富集并与小肠免疫细胞的丢失相关.此外,Il18^(-/-)小鼠的菌群失衡还导致其对李斯特菌的易感性增加.综上所述,本研究表明IL-18对共生菌的稳态至关重要且紊乱菌群可以显著地塑造免疫系统的格局.

Pannexin-1 infl uences peritoneal cavity cell population but is not involved in NLRP3 infl ammasome activation

Pannexin-1(Panx1)forms nonselective large channel in cell plasma membrane and has been shown to be associated with NLRP3 inflammasome activation,ATP release and phagocytes recruitment.In the current study,by manipulation of Panx1 expression in human myeloid cells and application of Panx1 defi cient mice,we failed to fi nd a correlation between Panx1 and NLRP3 infl ammasome activation,although an interaction between these two proteins was evident.However,in thioglycollate induced peritonitis,Panx1 defi cient mice showed much more phagocytes infiltration.Further analyses showed that mice defi cient for Panx1 exhibited enlarged F4/80^(low)Gr1-Ly6C-cell population in the peritonea.Our study thus reveals an important role for Panx1 in regulation of peritoneal cell population and peritonitis development.

The NLRP3 Inflammasome activation in human or mouse cells,sensitivity causes puzzle

SIGNAL 1 ALONE IS ENOUGH TO ACTIVATE THE NLRP3 INFLAMMAOSME IN HUMAN CELLS According to our current understanding,the NLR family,pyrin domain containing 3(NLRP3)infl ammasome activation is generally a two-step process.The fi rst step is priming,in which pathogen associated molecular patterns(PAMPs)such as LPS or pro-inflammatory cytokines such as tumor necrosis factor-α(TNF-α)induced NF-κB activation provides synthesis of pro-IL-1βand NLRP3 proteins.This priming step is considered as signal 1,which makes the cell ready for a second strike to assemble the infl ammasome.Then danger signals such as ATP and MSU provide the signal 2 that promotes formation of the NLRP3 infl ammasome and activates caspase-1.Both of these 2 steps are required in mouse macrophages for the NLRP3 inflammasome activation(Dinarello,2007).