scholarly journals IFN-γ mediates Paneth cell death via suppression of mTOR

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Alessandra Araujo ◽  
Alexandra Safronova ◽  
Elise Burger ◽  
Américo López-Yglesias ◽  
Shilpi Giri ◽  
...  
Keyword(s):  
Cell Death ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Paneth Cells ◽  
Cell Integrity ◽  
Host Protection ◽  
Cell Functions ◽  
Cell Death Pathways ◽  
Inflammatory Conditions ◽  
Ifn Γ

Paneth cells constitutively produce antimicrobial peptides and growth factors that allow for intestinal homeostasis, host protection and intestinal stem cell replication. Paneth cells rely heavily on the glycolytic metabolic program, which is in part controlled by the kinase complex Mechanistic target of rapamycin (mTORC1). Yet, little is known about mTOR importance in Paneth cell integrity under steady state and inflammatory conditions. Our results demonstrate that IFN-γ, a crucial mediator of the intestinal inflammation, acts directly on murine Paneth cells to alter their mitochondrial integrity and membrane potential, resulting in an mTORC1-dependent cell death mechanism distinct from canonical cell death pathways including apoptosis, necroptosis, and pyroptosis. These results were established with the purified cytokine and a physiologically relevant common Th1-inducing human parasite Toxoplasma gondii. Given the crucial role for IFN-γ, which is a cytokine frequently associated with the development of inflammatory bowel disease (IBD) and compromised Paneth cell functions, the identified mechanisms underlying mTORC1-dependent Paneth cell death downstream of IFN-γ may provide promising novel approaches for treating intestinal inflammation.

scholarly journals DOP86 An IFN-STAT1-MLKL axis drives programmed necrosis of Paneth cells in Crohn’s ileitis

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S125-S126
Author(s):  
L HARTMANN ◽  
B Siegmund ◽  
C Weidinger ◽  
C Becker ◽  
M F Neurath ◽  
...  
Keyword(s):  
Cell Death ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Signalling Pathways ◽  
Caspase 8 ◽  
Programmed Necrosis ◽  
Cell Death Pathways ◽  
Small Intestinal

Abstract Background Interferons (IFNs) are immune-modulatory cytokines expressed by epithelial and mucosal cells in response to viral and bacterial infection. Just recently, we discovered a correlation between IFN-λ expression and disease activity, including small intestinal inflammation and Paneth cell dysfunction, in human Crohn’s disease patients. On a molecular level, we uncovered that IFN-λ mediates epithelial cell death, in particular, Paneth cell death by a programmed necrosis, dependent on STAT1 activation and controlled by caspase-8. These results suggested that IFN-λ can be considered as a pathogenic cytokine in Crohn′s ileitis and should be considered as a new and promising target for future therapeutic intervention for this particular subtype of IBD. Our central question is now by which pathways interferon-regulated programmed necrosis of epithelial cells contributes to intestinal inflammation and how these mechanisms could be targeted for future therapeutic intervention. Methods We use a mouse model for Crohn’s Disease like inflammation and Paneth cell death that has a specific deletion of Caspase-8 in intestinal epithelial cells (Casp8∆IEC). We stimulate small intestinal organoids derived from Casp8∆IEC mice with IFNs in vitro and we overexpress IFN-λ in these mice in vivo by hydrodynamic tail vein injection of an IFN-λ expression vector. Furthermore, we use JAK-inhibitors to impede pharmacologically cell death pathways in the pathogenesis of intestinal inflammation in vitro and in vivo. Results We uncovered that gene expression of the cell death mediators Mlkl and Caspase-8 is dependent on IFN-λ-mediated JAK-STAT1 signalling. The non-specific pan JAK-inhibitor Tofacitinib is able to attenuate gene expression of Mlkl and Caspase-8 in vitro as well as in vivo. It prevents non-apoptotic as well as apoptotic cell death of small intestinal organoids stimulated with IFN-λ and is sufficient to prevent small intestinal tissue destruction in Casp8∆IEC mice challenged with IFN-λ. Additionally, we use the selective JAK1-inhibitor Filgotinib to limit the targeted JAK-STAT signalling pathways to only JAK1-STAT1 signalling and thus reduce side effects of the inhibitor on other signalling pathways. This had a similar effect as Tofacitinib suggesting that IFN controls MLKL-mediated cell death via JAK1. Conclusion In summary, our results indicate that targeting IFN-λ-mediated JAK-STAT1 signalling by the small-molecules Tofacitinib and Filgotinib impedes induction of Mlkl and Caspase-8-mediated cell death pathways. Therefore, JAK1 inhibitors such as Filgotinib might represent a promising novel therapy that may be sufficient to achieve efficacy particularly in Crohn′s ileitis patients who display elevated IFN-l serum levels.

Paneth Cells and their Antimicrobials in Intestinal Immunity

2018 ◽  
Vol 24 (10) ◽  
pp. 1121-1129 ◽  
Author(s):  
Timon E. Adolph ◽  
Lisa Mayr ◽  
Felix Grabherr ◽  
Herbert Tilg
Keyword(s):  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Paneth Cells ◽  
Secretory Cells ◽  
Intestinal Epithelial ◽  
Intestinal Immunity ◽  
Susceptibility To Infection ◽  
Cell Functions ◽  
Bowel Diseases ◽  
Initial Description

Since the initial description of granular-rich small-intestinal crypt-based epithelial cells in 1872, today referred to as Paneth cells, a plethora of recent studies underlined their function in intestinal homeostasis. Paneth cells are evolutionary conserved highly secretory cells that produce antimicrobials to control gut microbial communities. Moreover, Paneth cells emerged as stem cell regulators that translate environmental cues into intestinal epithelial responses. Paneth cell disturbances may instigate intestinal inflammation and provide susceptibility to infection. Altered Paneth cell functions have been associated with a variety of inflammatory disease models and were linked to human intestinal disease processes including inflammatory bowel diseases such as Crohn´s disease and ulcerative colitis. This review summarizes our current understanding of Paneth cells and their antimicrobials in health and disease.

scholarly journals Constitutive STAT5 activation regulates Paneth and Paneth-like cells to control Clostridium difficile colitis

2019 ◽  
Vol 2 (2) ◽  
pp. e201900296 ◽  
Author(s):  
Ruixue Liu ◽  
Richard Moriggl ◽  
Dongsheng Zhang ◽  
Haifeng Li ◽  
Rebekah Karns ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Cell Fate ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Cell Lineage ◽  
Niche Differentiation ◽  
Paneth Cells ◽  
Lineage Tracing ◽  
Intestinal Epithelial ◽  
Clostridium Difficile Colitis

Clostridium difficile impairs Paneth cells, driving intestinal inflammation that exaggerates colitis. Besides secreting bactericidal products to restrain C. difficile, Paneth cells act as guardians that constitute a niche for intestinal epithelial stem cell (IESC) regeneration. However, how IESCs are sustained to specify Paneth-like cells as their niche remains unclear. Cytokine-JAK-STATs are required for IESC regeneration. We investigated how constitutive STAT5 activation (Ca-pYSTAT5) restricts IESC differentiation towards niche cells to restrain C. difficile infection. We generated inducible transgenic mice and organoids to determine the effects of Ca-pYSTAT5-induced IESC lineages on C. difficile colitis. We found that STAT5 absence reduced Paneth cells and predisposed mice to C. difficile ileocolitis. In contrast, Ca-pYSTAT5 enhanced Paneth cell lineage tracing and restricted Lgr5 IESC differentiation towards pYSTAT5+Lgr5−CD24+Lyso+ or cKit+ niche cells, which imprinted Lgr5hiKi67+ IESCs. Mechanistically, pYSTAT5 activated Wnt/β-catenin signaling to determine Paneth cell fate. In conclusion, Ca-pYSTAT5 gradients control niche differentiation. Lack of pYSTAT5 reduces the niche cells to sustain IESC regeneration and induces C. difficile ileocolitis. STAT5 may be a transcription factor that regulates Paneth cells to maintain niche regeneration.

scholarly journals SOCS1 is essential for regulatory T cell functions by preventing loss of Foxp3 expression as well as IFN-γ and IL-17A production

2011 ◽  
Vol 208 (10) ◽  
pp. 2055-2067 ◽  
Author(s):  
Reiko Takahashi ◽  
Shuhei Nishimoto ◽  
Go Muto ◽  
Takashi Sekiya ◽  
Taiga Tamiya ◽  
...  
Keyword(s):  
T Cells ◽  
Inflammatory Responses ◽  
Foxp3 Expression ◽  
Treg Cells ◽  
Negative Regulator ◽  
Cytokine Signaling ◽  
Cell Integrity ◽  
Cell Functions ◽  
Ifn Γ

Regulatory T cells (Treg cells) maintain immune homeostasis by limiting inflammatory responses. SOCS1 (suppressor of cytokine signaling 1), a negative regulator of cytokine signaling, is necessary for the suppressor functions of Treg cells in vivo, yet detailed mechanisms remain to be clarified. We found that Socs1−/− Treg cells produced high levels of IFN-γ and rapidly lost Foxp3 when transferred into Rag2−/− mice or cultured in vitro, even though the CNS2 (conserved noncoding DNA sequence 2) in the Foxp3 enhancer region was fully demethylated. Socs1−/− Treg cells showed hyperactivation of STAT1 and STAT3. Because Foxp3 expression was stable and STAT1 activation was at normal levels in Ifnγ−/−Socs1−/− Treg cells, the restriction of IFN-γ–STAT1 signaling by SOCS1 is suggested to be necessary for stable Foxp3 expression. However, Ifnγ−/−Socs1−/− Treg cells had hyperactivated STAT3 and higher IL-17A (IL-17) production compared with Ifnγ−/−Socs1+/+ Treg cells and could not suppress colitis induced by naive T cells in Rag2−/− mice. In vitro experiments suggested that cytokines produced by Socs1−/− Treg cells and Ifnγ−/−Socs1−/− Treg cells modulated antigen-presenting cells for preferential Th1 and Th17 induction, respectively. We propose that SOCS1 plays important roles in Treg cell integrity and function by maintaining Foxp3 expression and by suppressing IFN-γ and IL-17 production driven by STAT1 and STAT3, respectively.

scholarly journals R-Spondin1 expands Paneth cells and prevents dysbiosis induced by graft-versus-host disease

2017 ◽  
Vol 214 (12) ◽  
pp. 3507-3518 ◽  
Author(s):  
Eiko Hayase ◽  
Daigo Hashimoto ◽  
Kiminori Nakamura ◽  
Clara Noizat ◽  
Reiki Ogasawara ◽  
...  
Keyword(s):  
Graft Versus Host Disease ◽  
Fecal Microbiota Transplantation ◽  
Paneth Cell ◽  
Fecal Microbiota ◽  
Paneth Cells ◽  
Host Disease ◽  
Graft Versus Host ◽  
Cell Functions ◽  
Therapeutic Benefits ◽  
Physiological Approach

The intestinal microbial ecosystem is actively regulated by Paneth cell–derived antimicrobial peptides such as α-defensins. Various disorders, including graft-versus-host disease (GVHD), disrupt Paneth cell functions, resulting in unfavorably altered intestinal microbiota (dysbiosis), which further accelerates the underlying diseases. Current strategies to restore the gut ecosystem are bacteriotherapy such as fecal microbiota transplantation and probiotics, and no physiological approach has been developed so far. In this study, we demonstrate a novel approach to restore gut microbial ecology by Wnt agonist R-Spondin1 (R-Spo1) or recombinant α-defensin in mice. R-Spo1 stimulates intestinal stem cells to differentiate to Paneth cells and enhances luminal secretion of α-defensins. Administration of R-Spo1 or recombinant α-defensin prevents GVHD-mediated dysbiosis, thus representing a novel and physiological approach at modifying the gut ecosystem to restore intestinal homeostasis and host–microbiota cross talk toward therapeutic benefits.

scholarly journals The Muc2 mucin coats murine Paneth cell granules and facilitates their content release and dispersion

2018 ◽  
Vol 315 (2) ◽  
pp. G195-G205 ◽  
Author(s):  
Martin Stahl ◽  
Sarah Tremblay ◽  
Marinieve Montero ◽  
Wayne Vogl ◽  
Lijun Xia ◽  
...  
Keyword(s):  
Cell Function ◽  
Paneth Cell ◽  
Goblet Cells ◽  
Paneth Cells ◽  
Phenotypic Differences ◽  
Cell Functions ◽  
First Time ◽  
Electron Lucent ◽  
Granule Release

Paneth cells are a key subset of secretory epithelial cells found at the base of small intestinal crypts. Unlike intestinal goblet cells, which secrete the mucin Muc2, Paneth cells are best known for producing an array of antimicrobial factors. We unexpectedly identified Muc2 staining localized around Paneth cell granules. Electron microscopy (EM) confirmed an electron lucent halo around these granules, which was lost in Paneth cells from Muc2-deficient (−/−) mice. EM and immunostaining for lysozyme revealed that Muc2−/− Paneth cells contained larger, more densely packed granules within their cytoplasm, and we detected defects in the transcription of key antimicrobial genes in the ileal tissues of Muc2−/− mice. Enteroids derived from the small intestine of wild-type and Muc2−/− mice revealed phenotypic differences in Paneth cells similar to those seen in vivo. Moreover, lysozyme-containing granule release from Muc2−/− enteroid Paneth cells was shown to be impaired. Surprisingly, Paneth cells within human ileal and duodenal tissues were found to be Muc2 negative. Thus Muc2 plays an important role in murine Paneth cells, suggesting links in function with goblet cells; however human Paneth cells lack Muc2, highlighting that caution should be applied when linking murine to human Paneth cell functions. NEW & NOTEWORTHY We demonstrate for the first time that murine Paneth cell granules possess a halo comprised of the mucin Muc2. The presence of Muc2 exerts an impact on Paneth cell granule size and number and facilitates the release and dispersal of antimicrobials into the mucus layer. Interestingly, despite the importance of Muc2 in murine Paneth cell function, our analysis of Muc2 in human intestinal tissues revealed no trace of Muc2 expression by human Paneth cells.

scholarly journals The RNA helicase Dhx15 mediates Wnt-induced antimicrobial protein expression in Paneth cells

2021 ◽  
Vol 118 (4) ◽  
pp. e2017432118
Author(s):  
Yalong Wang ◽  
Kaixin He ◽  
Baifa Sheng ◽  
Xuqiu Lei ◽  
Wanyin Tao ◽  
...  
Keyword(s):  
Rna Splicing ◽  
Intestinal Inflammation ◽  
Paneth Cell ◽  
Enteric Bacteria ◽  
Paneth Cells ◽  
Antimicrobial Protein ◽  
Rna Helicases

RNA helicases play roles in various essential biological processes such as RNA splicing and editing. Recent in vitro studies show that RNA helicases are involved in immune responses toward viruses, serving as viral RNA sensors or immune signaling adaptors. However, there is still a lack of in vivo data to support the tissue- or cell-specific function of RNA helicases owing to the lethality of mice with complete knockout of RNA helicases; further, there is a lack of evidence about the antibacterial role of helicases. Here, we investigated the in vivo role of Dhx15 in intestinal antibacterial responses by generating mice that were intestinal epithelial cell (IEC)-specific deficient for Dhx15 (Dhx15 f/f Villin1-cre, Dhx15ΔIEC). These mice are susceptible to infection with enteric bacteria Citrobacter rodentium (C. rod), owing to impaired α-defensin production by Paneth cells. Moreover, mice with Paneth cell-specific depletion of Dhx15 (Dhx15 f/f Defensinα6-cre, Dhx15ΔPaneth) are more susceptible to DSS (dextran sodium sulfate)-induced colitis, which phenocopy Dhx15ΔIEC mice, due to the dysbiosis of the intestinal microbiota. In humans, reduced protein levels of Dhx15 are found in ulcerative colitis (UC) patients. Taken together, our findings identify a key regulator of Wnt-induced α-defensins in Paneth cells and offer insights into its role in the antimicrobial response as well as intestinal inflammation.

scholarly journals Entamoeba histolyticaAlters Ileal Paneth Cell Functions in Intact and Muc2 Mucin Deficiency

2018 ◽  
Vol 86 (7) ◽  
pp. e00208-18 ◽  
Author(s):  
Eduardo R. Cobo ◽  
Ravi Holani ◽  
France Moreau ◽  
Kiminori Nakamura ◽  
Tokiyoshi Ayabe ◽  
...  
Keyword(s):  
Entamoeba Histolytica ◽  
Host Defense ◽  
Cysteine Proteinase ◽  
Paneth Cell ◽  
Active Form ◽  
Paneth Cells ◽  
Content Type ◽  
Cell Functions ◽  
Pathogen Invasion ◽  
And Function

ABSTRACTEnteric α-defensins, termed cryptdins (Crps) in mice, and lysozymes secreted by Paneth cells contribute to innate host defense in the ileum. Antimicrobial factors, including lysozymes and β-defensins, are often embedded in luminal glycosylated colonic Muc2 mucin secreted by goblet cells that form the protective mucus layer critical for gut homeostasis and pathogen invasion. In this study, we investigated ileal innate immunity againstEntamoeba histolytica, the causative agent of intestinal amebiasis, by inoculating parasites in closed ileal loops inMuc2+/+andMuc2−/−littermates and quantifying Paneth cell localization (lysozyme expression) and function (Crp secretion). Relative toMuc2+/+littermates,Muc2−/−littermates showed a disorganized mislocalization of Paneth cells that was diffusely distributed, with elevated lysozyme secretion in the crypts and on villi in response toE. histolytica. Inhibition ofE. histolyticaGal/GalNAc lectin (Gal-lectin) binding with exogenous galactose andEntamoeba histolyticacysteine proteinase 5 (EhCP5)-negativeE. histolyticahad no effect on parasite-induced erratic Paneth cell lysozyme synthesis. Although the basal ileal expression ofCrpgenes was unaffected inMuc2−/−mice in response toE. histolytica, there was a robust release of proinflammatory cytokines and Crp peptide secretions in luminal exudates that was also present in the colon. Interestingly,E. histolytica-secreted cysteine proteinases cleaved the proregion of Crp4 but not the active form. These findings define Muc2 mucin as an essential component of ileal barrier function that regulates the localization and function of Paneth cells critical for host defense against microbes.

scholarly journals Mycobacterium tuberculosis Protein PE6 (Rv0335c), a Novel TLR4 Agonist, Evokes an Inflammatory Response and Modulates the Cell Death Pathways in Macrophages to Enhance Intracellular Survival

2021 ◽  
Vol 12 ◽  
Author(s):  
Neha Sharma ◽  
Mohd Shariq ◽  
Neha Quadir ◽  
Jasdeep Singh ◽  
Javaid A. Sheikh ◽  
...  
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Proinflammatory Cytokines ◽  
Intracellular Survival ◽  
Immune Defense ◽  
Innate Immune ◽  
Bafilomycin A1 ◽  
Potent Inducer ◽  
Cell Functions ◽  
Cell Death Pathways

Mycobacterium tuberculosis (M. tb) is an intracellular pathogen that exploits moonlighting functions of its proteins to interfere with host cell functions. PE/PPE proteins utilize host inflammatory signaling and cell death pathways to promote pathogenesis. We report that M. tb PE6 protein (Rv0335c) is a secretory protein effector that interacts with innate immune toll-like receptor TLR4 on the macrophage cell surface and promotes activation of the canonical NFĸB signaling pathway to stimulate secretion of proinflammatory cytokines TNF-α, IL-12, and IL-6. Using mouse macrophage TLRs knockout cell lines, we demonstrate that PE6 induced secretion of proinflammatory cytokines dependent on TLR4 and adaptor Myd88. PE6 possesses nuclear and mitochondrial targeting sequences and displayed time-dependent differential localization into nucleus/nucleolus and mitochondria, and exhibited strong Nucleolin activation. PE6 strongly induces apoptosis via increased production of pro-apoptotic molecules Bax, Cytochrome C, and pcMyc. Mechanistic details revealed that PE6 activates Caspases 3 and 9 and induces endoplasmic reticulum-associated unfolded protein response pathways to induce apoptosis through increased production of ATF6, Chop, BIP, eIF2α, IRE1α, and Calnexin. Despite being a potent inducer of apoptosis, PE6 suppresses innate immune defense strategy autophagy by inducing inhibitory phosphorylation of autophagy initiating kinase ULK1. Inversely, PE6 induces activatory phosphorylation of autophagy master regulator MtorC1, which is reflected by lower conversion of autophagy markers LC3BI to LC3BII and increased accumulation of autophagy substrate p62 which is also dependent on innate immune receptor TLR4. The use of pharmacological agents, rapamycin and bafilomycin A1, confirms the inhibitory effect of PE6 on autophagy, evidenced by the reduced conversion of LC3BI to LC3BII and increased accumulation of p62 in the presence of rapamycin and bafilomycin A1. We also observed that PE6 binds DNA, which could have significant implications in virulence. Furthermore, our analyses reveal that PE6 efficiently binds iron to likely aid in intracellular survival. Recombinant Mycobacterium smegmatis (M. smegmatis) containing pe6 displayed robust growth in iron chelated media compared to vector alone transformed cells, which suggests a role of PE6 in iron acquisition. These findings unravel novel mechanisms exploited by PE6 protein to subdue host immunity, thereby providing insights relevant to a better understanding of host–pathogen interaction during M. tb infection.

scholarly journals Escherichia coli K12 Upregulates Programmed Cell Death Ligand 1 (PD-L1) Expression in Gamma Interferon-Sensitized Intestinal Epithelial Cells via the NF-κB Pathway

2020 ◽  
Vol 89 (1) ◽  
pp. e00618-20 ◽  
Author(s):  
Seul A. Lee ◽  
Yiming Wang ◽  
Fang Liu ◽  
Stephen M. Riordan ◽  
Lu Liu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Death ◽  
Programmed Cell Death ◽  
Bacterial Species ◽  
Intestinal Epithelial ◽  
Content Type ◽  
E Coli ◽  
Inflammatory Conditions ◽  
Ifn Γ ◽  
Ht 29

ABSTRACTProgrammed cell death ligand-1 (PD-L1) is an immune checkpoint protein which is used by tumor cells for immune evasion. PD-L1 is upregulated in inflamed intestinal tissues. The intestinal tract is colonized by millions of bacteria, most of which are commensal bacterial species. We hypothesized that under inflammatory conditions, some commensal bacterial species contribute to increased PD-L1 expression in intestinal epithelium and examined this hypothesis. Human intestinal epithelial HT-29 cells with and without interferon (IFN)-γ sensitization were incubated with six strains of four enteric bacterial species. The mRNA and protein levels of PD-L1 in HT-29 cells were examined using quantitative real-time PCR and flow cytometry, respectively. The levels of interleukin (IL)-1β, IL-18, IL-6, IL-8, and tumor necrosis factor (TNF)-α secreted by HT-29 cells were measured using enzyme-linked immunosorbent assay. Apoptosis of HT-29 cells was measured using a caspase 3/7 assay. We found that Escherichia coli K12 significantly upregulated both PD-L1 mRNA and protein in IFN-γ-sensitized HT-29 cells. E. coli K12 induced the production of IL-8 in HT-29 cells, however, IL-8 did not affect HT-29 PD-L1 expression. Inhibition of the nuclear factor-kappa B pathway significantly reduced E. coli K12-induced PD-L1 expression in HT-29 cells. The other two E. coli strains and two enteric bacterial species did not significantly affect PD-L1 expression in HT-29 cells. Enterococcus faecalis significantly inhibited PD-L1 expression due to induction of cell death. Data from this study suggest that some gut bacterial species have the potential to affect immune function under inflammatory conditions via upregulating epithelial PD-L1 expression.

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