scholarly journals Toxoplasma gondii Rhoptry 16 Kinase Promotes Host Resistance to Oral Infection and Intestinal Inflammation Only in the Context of the Dense Granule Protein GRA15

2013 ◽  
Vol 81 (6) ◽  
pp. 2156-2167 ◽  
Author(s):  
Kirk D. C. Jensen ◽  
Kenneth Hu ◽  
Ryan J. Whitmarsh ◽  
Musa A. Hassan ◽  
Lindsay Julien ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Host Resistance ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Chronic Phase ◽  
Parasite Burden ◽  
Oral Infection ◽  
Dense Granule ◽  
Intermediate Hosts ◽  
Content Type

ABSTRACTToxoplasma gondiitransmission between intermediate hosts is dependent on the ingestion of walled cysts formed during the chronic phase of infection. Immediately following consumption, the parasite must ensure survival of the host by preventing adverse inflammatory responses and/or by limiting its own replication. Since theToxoplasmasecreted effectors rhoptry 16 kinase (ROP16) and dense granule 15 (GRA15) activate the JAK-STAT3/6 and NF-κB signaling pathways, respectively, we explored whether a particular combination of these effectors impacted intestinal inflammation and parasite survivalin vivo. Here we report that expression of the STAT-activating version of ROP16 in the type II strain (strain II+ROP16I) promotes host resistance to oral infection only in the context of endogenous GRA15 expression. Protection was characterized by a lower intestinal parasite burden and dampened inflammation. Host resistance to the II+ROP16Istrain occurred independently of STAT6 and the T cell coinhibitory receptors B7-DC and B7-H1, two receptors that are upregulated by ROP16. In addition, coexpression of ROP16 and GRA15 enhanced parasite susceptibility within tumor necrosis factor alpha/gamma interferon-stimulated macrophages in a STAT3/6-independent manner. Transcriptional profiling of infected STAT3- and STAT6-deficient macrophages and parasitized Peyer's patches from mice orally challenged with strain II+ROP16Isuggested that ROP16 activated STAT5 to modulate host gene expression. Consistent with this supposition, the ROP16 kinase induced the sustained phosphorylation and nuclear localization of STAT5 inToxoplasma-infected cells. In summary, only the combined expression of both GRA15 and ROP16 promoted host resistance to acute oral infection, andToxoplasmamay possibly target the STAT5 signaling pathway to generate protective immunity in the gut.

scholarly journals Impact of Engineered Expression of Mitochondrial Association Factor 1b on Toxoplasma gondii Infection and the Host Response in a Mouse Model

mSphere ◽  
2018 ◽  
Vol 3 (5) ◽  
Author(s):  
Elizabeth D. English ◽  
Jon P. Boyle
Keyword(s):  
Toxoplasma Gondii ◽  
Chronic Infection ◽  
Acute Infection ◽  
Chronic Phase ◽  
Parasite Burden ◽  
Cytokine Signaling ◽  
Parasite Protein ◽  
Content Type ◽  
Life Threatening ◽  
The Impact

ABSTRACT The opportunistic intracellular parasite Toxoplasma gondii causes a lifelong chronic infection capable of reactivating in immunocompromised individuals, which can lead to life-threatening complications. Following invasion of the host cell, host mitochondria associate with the parasitophorous vacuole membrane. This phenotype is T. gondii strain specific and is mediated by expression of a host mitochondrial association-competent (HMA+) paralog of the parasite protein mitochondrial association factor 1 (MAF1b). Previous work demonstrated that expression of MAF1b in strains that do not normally associate with host mitochondria increases their fitness during acute infection in vivo. However, the impact of MAF1b expression during chronic T. gondii infection is unclear. In this study, we assess the impact of MAF1b expression on cyst formation and cytokine production in mice. Despite generally low numbers of cysts generated by the in vitro culture-adapted strains used in this study, we find that parasites expressing MAF1b have higher numbers of cysts in the brains of chronically infected mice and that MAF1b+ cyst burden significantly increases during the course of chronic infection. Consistent with this, mice infected with MAF1b+ parasites have higher levels of the serum cytokines RANTES and VEGF (vascular endothelial growth factor) at day 57 postinfection, although this could be due to higher parasite burden at this time point rather than direct manipulation of these cytokines by MAF1b. Overall these data indicate that MAF1b expression may also be important in determining infection outcome during the chronic phase, either by directly altering the cytokine/signaling environment or by increasing proliferation during the acute and/or chronic phase. IMPORTANCE The parasite Toxoplasma gondii currently infects approximately one-third of the world’s population and causes life-threatening toxoplasmosis in individuals with undeveloped or weakened immune systems. Current treatments are unable to cure T. gondii infection, leaving infected individuals with slow-growing tissue cysts for the remainder of their lives. Previous work has shown that expression of the parasite protein mitochondrial association factor 1 (MAF1b) is responsible for the association of T. gondii parasites with host mitochondria and provides a selective advantage during acute infection. Here we examine the impact of MAF1b expression during chronic T. gondii infection. We find that mice infected with MAF1b-expressing parasites have higher cyst burden and cytokine levels than their wild-type counterparts. A better understanding of the genes involved in establishing and maintaining chronic infection will aid in discovering effective therapeutics for chronically infected individuals.

scholarly journals RIPK3 Facilitates Host Resistance to Oral Toxoplasma gondii Infection

2021 ◽  
Vol 89 (5) ◽  
Author(s):  
Patrick W. Cervantes ◽  
Bruno Martorelli Di Genova ◽  
Billy Joel Erazo Flores ◽  
Laura J. Knoll
Keyword(s):  
Cell Death ◽  
Toxoplasma Gondii ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Parasite Burden ◽  
Kinase Domain ◽  
Content Type ◽  
Protection Mechanism ◽  
Cell Death Pathway ◽  
Toxoplasma Gondii Infection

ABSTRACT Toxoplasma gondii infection activates pattern recognition receptor (PRR) pathways that drive innate inflammatory responses to control infection. Necroptosis is a proinflammatory cell death pathway apart from the innate immune response that has evolved to control pathogenic infection. In this study, we further defined the role of Z-DNA binding protein 1 (ZBP1) as a PRR and assessed its contribution to necroptosis as a host protection mechanism to T. gondii infection. We found that ZBP1 does not induce proinflammatory necroptosis cell death, and ZBP1 null mice have reduced survival after oral T. gondii infection. In contrast, mice deleted in receptor-interacting serine/threonine-protein kinase 3 (RIPK3−/−), a central mediator of necroptosis, have significantly improved survival after oral T. gondii infection without a reduction in parasite burden. The physiological consequences of RIPK3 activity did not show any differences in intestine villus immunopathology, but RIPK3−/− mice showed higher immune cell infiltration and edema in the lamina propria. The contribution of necroptosis to host survival was clarified with mixed-lineage kinase domain-like pseudokinase null (MLKL−/−) mice. We found MLKL−/− mice succumbed to oral T. gondii infection the same as wild-type mice, indicating necroptosis-independent RIPK3 activity impacts host survival. These results provide new insights on the impacts of proinflammatory cell death pathways as a mechanism of host defense to oral T. gondii infection.

scholarly journals A Salmonella Virulence Factor Activates the NOD1/NOD2 Signaling Pathway

mBio ◽  
2011 ◽  
Vol 2 (6) ◽  
Author(s):  
A. Marijke Keestra ◽  
Maria G. Winter ◽  
Daisy Klein-Douwel ◽  
Mariana N. Xavier ◽  
Sebastian E. Winter ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Content Type ◽  
Type Iii

ABSTRACTThe invasion-associated type III secretion system (T3SS-1) ofSalmonella entericaserotype Typhimurium (S. Typhimurium) activates the transcription factor NF-κB in tissue culture cells and induces inflammatory responses in animal models through unknown mechanisms. Here we show that bacterial delivery or ectopic expression of SipA, a T3SS-1-translocated protein, led to the activation of the NOD1/NOD2 signaling pathway and consequent RIP2-mediated induction of NF-κB-dependent inflammatory responses. SipA-mediated activation of NOD1/NOD2 signaling was independent of bacterial invasionin vitrobut required an intact T3SS-1. In the mouse colitis model, SipA triggered mucosal inflammation in wild-type mice but not in NOD1/NOD2-deficient mice. These findings implicate SipA-driven activation of the NOD1/NOD2 signaling pathway as a mechanism by which the T3SS-1 induces inflammatory responsesin vitroandin vivo.IMPORTANCESalmonella entericaserotype Typhimurium (S. Typhimurium) deploys a type III secretion system (T3SS-1) to induce intestinal inflammation and benefits from the ensuing host response, which enhances growth of the pathogen in the intestinal lumen. However, the mechanisms by which the T3SS-1 triggers inflammatory responses have not been resolved. Here we show that the T3SS-1 effector protein SipA induces NF-κB activation and intestinal inflammation by activating the NOD1/NOD2 signaling pathway. These data suggest that the T3SS-1 escalates innate responses through a SipA-mediated activation of pattern recognition receptors in the host cell cytosol.

scholarly journals Infection-Induced Intestinal Dysbiosis Is Mediated by Macrophage Activation and Nitrate Production

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Shuai Wang ◽  
Ayah El-Fahmawi ◽  
David A. Christian ◽  
Qun Fang ◽  
Enrico Radaelli ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Oral Infection ◽  
Nitrate Respiration ◽  
Characteristic Change ◽  
Intestinal Immunity ◽  
Parasite Control ◽  
Shotgun Metagenomics ◽  
Content Type ◽  
Distal Small Intestine ◽  
Ifn Γ

ABSTRACT Oral infection of C57BL/6J mice with Toxoplasma gondii results in a marked bacterial dysbiosis and the development of severe pathology in the distal small intestine that is dependent on CD4+ T cells and interferon gamma (IFN-γ). This dysbiosis and bacterial translocation contribute to the development of ileal pathology, but the factors that support the bloom of bacterial pathobionts are unclear. The use of microbial community profiling and shotgun metagenomics revealed that Toxoplasma infection induces a dysbiosis dominated by Enterobacteriaceae and an increased potential for nitrate respiration. In vivo experiments using bacterial metabolic mutants revealed that during this infection, host-derived nitrate supports the expansion of Enterobacteriaceae in the ileum via nitrate respiration. Additional experiments with infected mice indicate that the IFN-γ/STAT1/iNOS axis, while essential for parasite control, also supplies a pool of nitrate that serves as a source for anaerobic respiration and supports overgrowth of Enterobacteriaceae. Together, these data reveal a trade-off in intestinal immunity after oral infection of C57BL/6J mice with T. gondii, in which inducible nitric oxide synthase (iNOS) is required for parasite control, while this host enzyme is responsible for specific modification of the composition of the microbiome that contributes to pathology. IMPORTANCE Toxoplasma gondii is a protozoan parasite and a leading cause of foodborne illness. Infection is initiated when the parasite invades the intestinal epithelium, and in many host species, this leads to intense inflammation and a dramatic disruption of the normal microbial ecosystem that resides in the healthy gut (the so-called microbiome). One characteristic change in the microbiome during infection with Toxoplasma—as well as numerous other pathogens—is the overgrowth of Escherichia coli or similar bacteria and a breakdown of commensal containment leading to seeding of peripheral organs with gut bacteria and subsequent sepsis. Our findings provide one clear explanation for how this process is regulated, thereby improving our understanding of the relationship between parasite infection, inflammation, and disease. Furthermore, our results could serve as the basis for the development of novel therapeutics to reduce the potential for harmful bacteria to bloom in the gut during infection.

scholarly journals Immediate Interferon Gamma Induction Determines Murine Host Compatibility Differences between Toxoplasma gondii and Neospora caninum

2020 ◽  
Vol 88 (4) ◽  
Author(s):  
Rachel S. Coombs ◽  
Matthew L. Blank ◽  
Elizabeth D. English ◽  
Yaw Adomako-Ankomah ◽  
Ifeanyi-Chukwu Samuel Urama ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Immune Responses ◽  
Interferon Gamma ◽  
Neospora Caninum ◽  
Ectopic Expression ◽  
Parasite Burden ◽  
Interleukin 12 ◽  
Content Type ◽  
Ifn Γ ◽  
And Control

ABSTRACT Rodents are critical for the transmission of Toxoplasma gondii to the definitive feline host via predation, and this relationship has been extensively studied as a model for immune responses to parasites. Neospora caninum is a closely related coccidian parasite of ruminants and canines but is not naturally transmitted by rodents. We compared mouse innate immune responses to N. caninum and T. gondii and found marked differences in cytokine levels and parasite growth kinetics during the first 24 h postinfection (hpi). N. caninum-infected mice produced significantly higher levels of interleukin-12 (IL-12) and interferon gamma (IFN-γ) by as early as 4 hpi, but the level of IFN-γ was significantly lower or undetectable in T. gondii-infected mice during the first 24 hpi. “Immediate” IFN-γ and IL-12p40 production was not detected in MyD88−/− mice. However, unlike IL-12p40−/− and IFN-γ−/− mice, MyD88−/− mice survived N. caninum infections at the dose used in this study. Serial measures of parasite burden showed that MyD88−/− mice were more susceptible to N. caninum infections than wild-type (WT) mice, and control of parasite burdens correlated with a pulse of serum IFN-γ at 3 to 4 days postinfection in the absence of detectable IL-12. Immediate IFN-γ was partially dependent on the T. gondii mouse profilin receptor Toll-like receptor 11 (TLR11), but the ectopic expression of N. caninum profilin in T. gondii had no impact on early IFN-γ production or parasite proliferation. Our data indicate that T. gondii is capable of evading host detection during the first hours after infection, while N. caninum is not, and this is likely due to the early MyD88-dependent recognition of ligands other than profilin.

scholarly journals Coimmunoprecipitation with MYR1 Identifies Three Additional Proteins within the Toxoplasma gondii Parasitophorous Vacuole Required for Translocation of Dense Granule Effectors into Host Cells

mSphere ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Alicja M. Cygan ◽  
Terence C. Theisen ◽  
Alma G. Mendoza ◽  
Nicole D. Marino ◽  
Michael W. Panas ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Protein Translocation ◽  
Affinity Purification ◽  
Parasitophorous Vacuole ◽  
Dense Granule ◽  
Effector Proteins ◽  
Host Cells ◽  
Content Type ◽  
Cyclin E1 ◽  
Infected Cells

ABSTRACT Toxoplasma gondii is a ubiquitous, intracellular protozoan that extensively modifies infected host cells through secreted effector proteins. Many such effectors must be translocated across the parasitophorous vacuole (PV), in which the parasites replicate, ultimately ending up in the host cytosol or nucleus. This translocation has previously been shown to be dependent on five parasite proteins: MYR1, MYR2, MYR3, ROP17, and ASP5. We report here the identification of several MYR1-interacting and novel PV-localized proteins via affinity purification of MYR1, including TGGT1_211460 (dubbed MYR4), TGGT1_204340 (dubbed GRA54), and TGGT1_270320 (PPM3C). Further, we show that three of the MYR1-interacting proteins, GRA44, GRA45, and MYR4, are essential for the translocation of the Toxoplasma effector protein GRA16 and for the upregulation of human c-Myc and cyclin E1 in infected cells. GRA44 and GRA45 contain ASP5 processing motifs, but like MYR1, processing at these sites appears to be nonessential for their role in protein translocation. These results expand our understanding of the mechanism of effector translocation in Toxoplasma and indicate that the process is highly complex and dependent on at least eight discrete proteins. IMPORTANCE Toxoplasma is an extremely successful intracellular parasite and important human pathogen. Upon infection of a new cell, Toxoplasma establishes a replicative vacuole and translocates parasite effectors across this vacuole to function from the host cytosol and nucleus. These effectors play a key role in parasite virulence. The work reported here newly identifies three parasite proteins that are necessary for protein translocation into the host cell. These results significantly increase our knowledge of the molecular players involved in protein translocation in Toxoplasma-infected cells and provide additional potential drug targets.

scholarly journals CCR5 Is Involved in Interruption of Pregnancy in Mice Infected with Toxoplasma gondii during Early Pregnancy

2017 ◽  
Vol 85 (9) ◽  
Author(s):  
Maki Nishimura ◽  
Kousuke Umeda ◽  
Masayuki Suwa ◽  
Hidefumi Furuoka ◽  
Yoshifumi Nishikawa
Keyword(s):  
Toxoplasma Gondii ◽  
Early Pregnancy ◽  
Parasite Burden ◽  
Wild Type ◽  
Necrosis Factor Alpha ◽  
Content Type ◽  
Pregnant Mice ◽  
Implantation Sites ◽  
Factor Alpha ◽  
Embryo Loss

ABSTRACT Toxoplasmosis can cause abortion in pregnant humans and other animals; however, the mechanism of abortion remains unknown. C-C chemokine receptor type 5 (CCR5) is essential for host defense against Toxoplasma gondii infection. To investigate the relationship between CCR5 and abortion in toxoplasmosis, we inoculated wild-type and CCR5-deficient (CCR5−/−) mice with T. gondii tachyzoites intraperitoneally on day 3 of pregnancy (embryonic day 3 [E3]). The pregnancy rate decreased as pregnancy progressed in infected wild-type mice. Histopathologically, no inflammatory lesions were observed in the fetoplacental tissues. Although wild-type mice showed a higher parasite burden at the implantation sites than did CCR5−/− mice at E6 (3 days postinfection [dpi]), T. gondii antigen was detected only in the uterine tissue and not in the fetoplacental tissues. At E8 (5 dpi), the embryos in infected wild-type mice showed poor development compared with those of infected CCR5−/− mice, and apoptosis was observed in poorly developed embryos. Compared to uninfected mice, infected wild-type mice showed increased CCR5 expression at the implantation site at E6 and E8. Furthermore, analyses of mRNA expression in the uterus of nonpregnant and pregnant mice suggested that a lack of the CCR5 gene and the downregulation of tumor necrosis factor alpha (TNF-α) and CCL3 expression at E6 (3 dpi) are important factors for the maintenance of pregnancy following T. gondii infection. These results suggested that CCR5 signaling is involved in embryo loss in T. gondii infection during early pregnancy and that apoptosis is associated with embryo loss rather than direct damage to the fetoplacental tissues.

scholarly journals Targeted Disruption of Toxoplasma gondii Serine Protease Inhibitor 1 Increases Bradyzoite Cyst FormationIn Vitroand Parasite Tissue Burden in Mice

2011 ◽  
Vol 80 (3) ◽  
pp. 1156-1165 ◽  
Author(s):  
Viviana Pszenny ◽  
Paul H. Davis ◽  
Xing W. Zhou ◽  
Christopher A. Hunter ◽  
Vern B. Carruthers ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Protease Inhibitor ◽  
Host Cell ◽  
Serine Protease ◽  
Parasitophorous Vacuole ◽  
Parasite Burden ◽  
Serine Protease Inhibitor ◽  
Genomic Locus ◽  
Content Type

As an intracellular protozoan parasite,Toxoplasma gondiiis likely to exploit proteases for host cell invasion, acquisition of nutrients, avoidance of host protective responses, escape from the parasitophorous vacuole, differentiation, and other activities.T. gondiiserine protease inhibitor 1 (TgPI1) is the most abundantly expressed protease inhibitor in parasite tachyzoites. We show here that alternative splicing produces twoTgPI1 isoforms, both of which are secreted via dense granules into the parasitophorous vacuole shortly after invasion, become progressively more abundant over the course of the infectious cycle, and can be detected in the infected host cell cytoplasm. To investigateTgPI1 function, the endogenous genomic locus was disrupted in the RH strain background. ΔTgPI1 parasites replicate normally as tachyzoites but exhibit increased bradyzoite gene transcription and labeling of vacuoles withDolichos bifloruslectin under conditions promotingin vitrodifferentiation. The differentiation phenotype can be partially complemented by eitherTgPI1 isoform. Mice infected with the ΔTgPI1 mutant display ∼3-fold-increased parasite burden in the spleen and liver, and thisin vivophenotype is also complemented by eitherTgPI1 isoform. These results demonstrate thatTgPI1 influences both parasite virulence and bradyzoite differentiation, presumably by inhibiting parasite and/or host serine proteases.

scholarly journals Guanylate Binding Proteins Enable Rapid Activation of Canonical and Noncanonical Inflammasomes in Chlamydia-Infected Macrophages

2015 ◽  
Vol 83 (12) ◽  
pp. 4740-4749 ◽  
Author(s):  
Ryan Finethy ◽  
Ine Jorgensen ◽  
Arun K. Haldar ◽  
Marcel R. de Zoete ◽  
Till Strowig ◽  
...  
Keyword(s):  
Host Resistance ◽  
Binding Proteins ◽  
Inflammatory Responses ◽  
Inflammasome Activation ◽  
Fast Kinetics ◽  
Content Type ◽  
Chlamydia Muridarum ◽  
Cell Cytosol ◽  
Ifn Γ ◽  
Mediate Cell

Interferon (IFN)-inducible guanylate binding proteins (GBPs) mediate cell-autonomous host resistance to bacterial pathogens and promote inflammasome activation. The prevailing model postulates that these two GBP-controlled activities are directly linked through GBP-dependent vacuolar lysis. It was proposed that the rupture of pathogen-containing vacuoles (PVs) by GBPs destroyed the microbial refuge and simultaneously contaminated the host cell cytosol with microbial activators of inflammasomes. Here, we demonstrate that GBP-mediated host resistance and GBP-mediated inflammatory responses can be uncoupled. We show that PVs formed by the rodent pathogenChlamydia muridarum, so-called inclusions, remain free of GBPs and thatC. muridarumis impervious to GBP-mediated restrictions on bacterial growth. Although GBPs neither bind toC. muridaruminclusions nor restrictC. muridarumgrowth, we find that GBPs promote inflammasome activation inC. muridarum-infected macrophages. We demonstrate thatC. muridaruminfections induce GBP-dependent pyroptosis through both caspase-11-dependent noncanonical and caspase-1-dependent canonical inflammasomes. Among canonical inflammasomes, we find thatC. muridarumand the human pathogenChlamydia trachomatisactivate not only NLRP3 but also AIM2. Our data show that GBPs support fast-kinetics processing and secretion of interleukin-1β (IL-1β) and IL-18 by the NLRP3 inflammasome but are dispensable for the secretion of the same cytokines at later times postinfection. Because IFN-γ fails to induce IL-1β transcription, GBP-dependent fast-kinetics inflammasome activation can drive the preferential processing of constitutively expressed IL-18 in IFN-γ-primed macrophages in the absence of prior Toll-like receptor stimulation. Together, our results reveal that GBPs control the kinetics of inflammasome activation and thereby shape macrophage responses toChlamydiainfections.

scholarly journals Fidaxomicin and OP-1118 Inhibit Clostridium difficile Toxin A- and B-Mediated Inflammatory Responses via Inhibition of NF-κB Activity

2017 ◽  
Vol 62 (1) ◽  
Author(s):  
Hon Wai Koon ◽  
Jiani Wang ◽  
Caroline C. Mussatto ◽  
Christina Ortiz ◽  
Elaine C. Lee ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Human Colon ◽  
Toxin A ◽  
Primary Metabolite ◽  
Necrosis Factor Alpha ◽  
Toxin B ◽  
Content Type ◽  
Tnf Α

ABSTRACTClostridium difficilecauses diarrhea and colitis by releasing toxin A and toxin B. In the human colon, both toxins cause intestinal inflammation and stimulate tumor necrosis factor alpha (TNF-α) expression via the activation of NF-κB. It is well established that the macrolide antibiotic fidaxomicin is associated with reduced relapses ofC. difficileinfection. We showed that fidaxomicin and its primary metabolite OP-1118 significantly inhibited toxin A-mediated intestinal inflammation in micein vivoand toxin A-induced cell roundingin vitro. We aim to determine whether fidaxomicin and OP-1118 possess anti-inflammatory effects against toxin A and toxin B in the human colon and examine the mechanism of this response. We used fresh human colonic explants, NCM460 human colonic epithelial cells, and RAW264.7 mouse macrophages to study the mechanism of the activity of fidaxomicin and OP-1118 against toxin A- and B-mediated cytokine expression and apoptosis. Fidaxomicin and OP-1118 dose-dependently inhibited toxin A- and B-induced TNF-α and interleukin-1β (IL-1β) mRNA expression and histological damage in human colonic explants. Fidaxomicin and OP-1118 inhibited toxin A-mediated NF-κB phosphorylation in human and mouse intestinal mucosae. Fidaxomicin and OP-1118 also inhibited toxin A-mediated NF-κB phosphorylation and TNF-α expression in macrophages, which was reversed by the NF-κB activator phorbol myristate acetate (PMA). Fidaxomicin and OP-1118 prevented toxin A- and B-mediated apoptosis in NCM460 cells, which was reversed by the addition of PMA. PMA reversed the cytoprotective effect of fidaxomicin and OP-1118 in toxin-exposed human colonic explants. Fidaxomicin and OP-1118 inhibitC. difficiletoxin A- and B-mediated inflammatory responses, NF-κB phosphorylation, and tissue damage in the human colon.

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