scholarly journals Involvement of Caspase-9 in the Inhibition of Necrosis of RAW 264 Cells Infected with Mycobacterium tuberculosis

2007 ◽  
Vol 75 (6) ◽  
pp. 2894-2902 ◽  
Author(s):  
Ryosuke Uchiyama ◽  
Ikuo Kawamura ◽  
Takao Fujimura ◽  
Michiko Kawanishi ◽  
Kohsuke Tsuchiya ◽  
...  
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Host Cell ◽  
Specific Inhibitor ◽  
Host Cells ◽  
Ros Generation ◽  
Caspase 9 ◽  
Host Cell Death ◽  
Intracellular Ros ◽  
Infected Cells

ABSTRACT In order to know how caspases contribute to the intracellular fate of Mycobacterium tuberculosis and host cell death in the infected macrophages, we examined the effect of benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethane (z-VAD-fmk), a broad-spectrum caspase inhibitor, on the growth of M. tuberculosis H37Rv in RAW 264 cells. In the cells treated with z-VAD-fmk, activation of caspase-8, caspase-3/7, and caspase-9 was clearly suppressed, and DNA fragmentation of the infected cells was also reduced. Under this experimental condition, it was found that the treatment markedly inhibited bacterial growth inside macrophages. The infected cells appeared to undergo cell death of the necrosis type in the presence of z-VAD-fmk. We further found that z-VAD-fmk treatment resulted in the generation of intracellular reactive oxygen species (ROS) in the infected cells. By addition of a scavenger of ROS, the host cell necrosis was inhibited and the intracellular growth of H37Rv was significantly restored. Among inhibitors specific for each caspase, only the caspase-9-specific inhibitor enhanced the generation of ROS and induced necrosis of the infected cells. Furthermore, we found that severe necrosis was induced by infection with H37Rv but not H37Ra in the presence of z-VAD-fmk. Caspase-9 activation was also detected in H37Rv-infected cells, but H37Ra never induced such caspase-9 activation. These results indicated that caspase-9, which was activated by infection with virulent M. tuberculosis, contributed to the inhibition of necrosis of the infected host cells, presumably through suppression of intracellular ROS generation.

scholarly journals Intracellular growth of Mycobacterium tuberculosis after macrophage cell death leads to serial killing of host cells

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Deeqa Mahamed ◽  
Mikael Boulle ◽  
Yashica Ganga ◽  
Chanelle Mc Arthur ◽  
Steven Skroch ◽  
...  
Keyword(s):  
Cell Death ◽  
Mycobacterium Tuberculosis ◽  
Host Cells ◽  
Intracellular Growth ◽  
Pathogen Virulence ◽  
Host Cell Death ◽  
A Cell ◽  
Infected Cells ◽  
Serial Killing ◽  
Cell Death Cascade

A hallmark of pulmonary tuberculosis is the formation of macrophage-rich granulomas. These may restrict Mycobacterium tuberculosis (Mtb) growth, or progress to central necrosis and cavitation, facilitating pathogen growth. To determine factors leading to Mtb proliferation and host cell death, we used live cell imaging to track Mtb infection outcomes in individual primary human macrophages. Internalization of Mtb aggregates caused macrophage death, and phagocytosis of large aggregates was more cytotoxic than multiple small aggregates containing similar numbers of bacilli. Macrophage death did not result in clearance of Mtb. Rather, it led to accelerated intracellular Mtb growth regardless of prior activation or macrophage type. In contrast, bacillary replication was controlled in live phagocytes. Mtb grew as a clump in dead cells, and macrophages which internalized dead infected cells were very likely to die themselves, leading to a cell death cascade. This demonstrates how pathogen virulence can be achieved through numbers and aggregation states.

scholarly journals Pathogen induced subversion of NAD+ metabolism mediating host cell death: a target for development of chemotherapeutics

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Ayushi Chaurasiya ◽  
Swati Garg ◽  
Ashish Khanna ◽  
Chintam Narayana ◽  
Ved Prakash Dwivedi ◽  
...  
Keyword(s):  
Cell Death ◽  
Host Cell ◽  
Nicotinamide Adenine Dinucleotide ◽  
Malaria Parasite ◽  
Host Cells ◽  
Nicotinamide Adenine ◽  
Targeted Therapeutics ◽  
Nad Metabolism ◽  
Host Cell Death ◽  
Nanomolar Range

AbstractHijacking of host metabolic status by a pathogen for its regulated dissemination from the host is prerequisite for the propagation of infection. M. tuberculosis secretes an NAD+-glycohydrolase, TNT, to induce host necroptosis by hydrolyzing Nicotinamide adenine dinucleotide (NAD+). Herein, we expressed TNT in macrophages and erythrocytes; the host cells for M. tuberculosis and the malaria parasite respectively, and found that it reduced the NAD+ levels and thereby induced necroptosis and eryptosis resulting in premature dissemination of pathogen. Targeting TNT in M. tuberculosis or induced eryptosis in malaria parasite interferes with pathogen dissemination and reduction in the propagation of infection. Building upon our discovery that inhibition of pathogen-mediated host NAD+ modulation is a way forward for regulation of infection, we synthesized and screened some novel compounds that showed inhibition of NAD+-glycohydrolase activity and pathogen infection in the nanomolar range. Overall this study highlights the fundamental importance of pathogen-mediated modulation of host NAD+ homeostasis for its infection propagation and novel inhibitors as leads for host-targeted therapeutics.

scholarly journals Core N-Glycan Structures Are Critical for the Pathogenicity of Cryptococcus neoformans by Modulating Host Cell Death

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Eun Jung Thak ◽  
Su-Bin Lee ◽  
Shengjie Xu-Vanpala ◽  
Dong-Jik Lee ◽  
Seung-Yeon Chung ◽  
...  
Keyword(s):  
Cell Death ◽  
Fungal Infection ◽  
Host Cell ◽  
Host Cells ◽  
Glycan Structure ◽  
Macrophage Cell ◽  
Content Type ◽  
Host Cell Death ◽  
Intact Core

ABSTRACT Cryptococcus neoformans is a human-pathogenic fungal pathogen that causes life-threatening meningoencephalitis in immunocompromised individuals. To investigate the roles of N-glycan core structure in cryptococcal pathogenicity, we constructed mutant strains of C. neoformans with defects in the assembly of lipid-linked N-glycans in the luminal side of the endoplasmic reticulum (ER). Deletion of ALG3 (alg3Δ), which encodes dolichyl-phosphate-mannose (Dol-P-Man)-dependent α-1,3-mannosyltransferase, resulted in the production of truncated neutral N-glycans carrying five mannose residues as a major species. Despite moderate or nondetectable defects in virulence-associated phenotypes in vitro, the alg3Δ mutant was avirulent in a mouse model of systemic cryptococcosis. Notably, the mutant did not show defects in early stages of host cell interaction during infection, including attachment to lung epithelial cells, opsonic/nonopsonic phagocytosis, and manipulation of phagosome acidification. However, the ability to drive macrophage cell death was greatly decreased in this mutant, without loss of cell wall remodeling capacity. Furthermore, deletion of ALG9 and ALG12, encoding Dol-P-Man-dependent α-1,2-mannosyltransferases and α-1,6-mannosyltransferases, generating truncated core N-glycans with six and seven mannose residues, respectively, also displayed remarkably reduced macrophage cell death and in vivo virulence. However, secretion levels of interleukin-1β (IL-1β) were not reduced in the bone marrow-derived dendritic cells obtained from Asc- and Gsdmd-deficient mice infected with the alg3Δ mutant strain, excluding the possibility that pyroptosis is a main host cell death pathway dependent on intact core N-glycans. Our results demonstrated N-glycan structures as a critical feature in modulating death of host cells, which is exploited by as a strategy for host cell escape for dissemination of C. neoformans. IMPORTANCE We previously reported that the outer mannose chains of N-glycans are dispensable for the virulence of C. neoformans, which is in stark contrast to findings for the other human-pathogenic yeast, Candida albicans. Here, we present evidence that an intact core N-glycan structure is required for C. neoformans pathogenicity by systematically analyzing alg3Δ, alg9Δ, and alg12Δ strains that have defects in lipid-linked N-glycan assembly and in in vivo virulence. The alg null mutants producing truncated core N-glycans were defective in inducing host cell death after phagocytosis, which is triggered as a mechanism of pulmonary escape and dissemination of C. neoformans, thus becoming inactive in causing fatal infection. The results clearly demonstrated the critical features of the N-glycan structure in mediating the interaction with host cells during fungal infection. The delineation of the roles of protein glycosylation in fungal pathogenesis not only provides insight into the glycan-based fungal infection mechanism but also will aid in the development of novel antifungal agents.

scholarly journals Listeria monocytogenes: The Impact of Cell Death on Infection and Immunity

Pathogens ◽  
2018 ◽  
Vol 7 (1) ◽  
pp. 8 ◽  
Author(s):  
Courtney McDougal ◽  
John-Demian Sauer
Keyword(s):  
Cell Death ◽  
Listeria Monocytogenes ◽  
Host Cell ◽  
Acute Infection ◽  
Host Cells ◽  
Cell Mediated Immunity ◽  
Cell Death Pathways ◽  
Host Cell Death ◽  
The Impact

Listeria monocytogenes has evolved exquisite mechanisms for invading host cells and spreading from cell-to-cell to ensure maintenance of its intracellular lifecycle. As such, it is not surprising that loss of the intracellular replication niche through induction of host cell death has significant implications on the development of disease and the subsequent immune response. Although L. monocytogenes can activate multiple pathways of host cell death, including necrosis, apoptosis, and pyroptosis, like most intracellular pathogens L. monocytogenes has evolved a series of adaptations that minimize host cell death to promote its virulence. Understanding how L. monocytogenes modulates cell death during infection could lead to novel therapeutic approaches. In addition, as L. monocytogenes is currently being developed as a tumor immunotherapy platform, understanding how cell death pathways influence the priming and quality of cell-mediated immunity is critical. This review will focus on the mechanisms by which L. monocytogenes modulates cell death, as well as the implications of cell death on acute infection and the generation of adaptive immunity.

scholarly journals RIPK3-Dependent Necroptosis Limits PRV Replication in PK-15 Cells

2021 ◽  
Vol 12 ◽  
Author(s):  
Hongchao Gou ◽  
Zhibiao Bian ◽  
Rujian Cai ◽  
Pinpin Chu ◽  
Shuai Song ◽  
...  
Keyword(s):  
Cell Death ◽  
Pseudorabies Virus ◽  
Specific Inhibitor ◽  
Host Cells ◽  
Positive Staining ◽  
Necrotic Cell Death ◽  
Necrotic Cell ◽  
Transmission Electron Microscopy Analysis ◽  
Necrosis Factor Alpha ◽  
Infected Cells

Pigs infected by pseudorabies virus (PRV) display necrotic pathology in multiple organs. The mechanism by which PRV induces cell death is still unclear. Recently, necroptosis was identified as a programmed process dependent on the receptor interacting protein kinase 3 (RIPK3) and mixed lineage kinase-like protein (MLKL). In this study, we demonstrated that PRV induced RIPK3-dependent necroptosis in PK-15 cells. The data showed that PRV infection caused cell death with Propidium Iodide (PI)-positive staining. Transmission electron microscopy analysis indicated plasma membrane disruption in PRV-infected cells. A pan-caspase inhibitor did not prevent PRV-induced necrotic cell death. Western blot analysis indicated that caspase-3 and caspase-8 were not cleaved during PRV infection. Although the transcription of tumor necrosis factor-alpha (TNF-α) was increased by PRV infection, RIPK1 was shown to be not involved in PRV-induced necrotic cell death by use of its specific inhibitor. Further experiments indicated that the phosphorylation of RIPK3 and MLKL was upregulated in PRV-infected cells. Stable shRNA knockdown of RIPK3 or MLKL had a recovery effect on PRV-induced necrotic cell death. Meanwhile, viral titers were enhanced in RIPK3 and MLKL knockdown cells. Hence, we concluded that initiation of necroptosis in host cells plays a limiting role in PRV infection. Considering that necroptosis is an inflammatory form of programmed cell death, our data may be beneficial for understanding the necrotic pathology of pigs infected by PRV.

scholarly journals Intracellular Staphylococcus aureus employs the cysteine protease staphopain A to induce host cell death in epithelial cells

2020 ◽  
Author(s):  
Kathrin Stelzner ◽  
Tobias Hertlein ◽  
Aneta Sroka ◽  
Adriana Moldovan ◽  
Kerstin Paprotka ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Death ◽  
Epithelial Cells ◽  
Host Cell ◽  
Cysteine Protease ◽  
Host Cells ◽  
Upper Respiratory Tract ◽  
Intracellular Replication ◽  
Tissue Destruction ◽  
Host Cell Death

AbstractStaphylococcus aureus is a major human pathogen, which can invade and survive in non-professional and professional phagocytes. Intracellularity is thought to contribute to pathogenicity and persistence of the bacterium. Upon internalization by epithelial cells, cytotoxic S. aureus strains can escape from the phagosome, replicate in the cytosol and induce host cell death. Here, we identified a staphylococcal cysteine protease to induce cell death by intracellular S. aureus after translocation into the host cell cytoplasm. We demonstrated that loss of staphopain A function leads to delayed onset of host cell death and prolonged intracellular replication of S. aureus in epithelial cells. Overexpression of staphopain A in a non-cytotoxic strain facilitated intracellular killing of the host cell even in the absence of detectable intracellular replication. Moreover, staphopain A contributed to efficient colonization of the lung in a mouse pneumonia model. Our study suggests that staphopain A is utilized by S. aureus to mediate escape from the host cell and thus contributes to tissue destruction and dissemination of infection.Author SummaryStaphylococcus aureus is a well-known antibiotic-resistant pathogen that emerges in hospital and community settings and can cause a variety of diseases ranging from skin abscesses to lung inflammation and blood poisoning. The bacterium asymptomatically colonizes the upper respiratory tract and skin of about one third of the human population and takes advantage of opportune conditions, like immunodeficiency or breached barriers, to cause infection. Although S. aureus is not regarded as a professional intracellular bacterium, it can be internalized by human cells and subsequently exit the host cells by induction of cell death, which is considered to cause tissue destruction and spread of infection. The bacterial virulence factors and underlying molecular mechanisms involved in the intracellular lifestyle of S. aureus remain largely unknown. We identified a bacterial cysteine protease to contribute to host cell death mediated by intracellular S. aureus. Staphopain A induced killing of the host cell after translocation of the pathogen into the cell cytosol, while bacterial proliferation was not required. Further, the protease enhanced survival of the pathogen during lung infection. These findings reveal a novel, intracellular role for the bacterial protease staphopain A.

scholarly journals EnteropathogenicE. coli-induced barrier function alteration is not a consequence of host cell apoptosis

2008 ◽  
Vol 294 (5) ◽  
pp. G1165-G1170 ◽  
Author(s):  
V. K. Viswanathan ◽  
Andrew Weflen ◽  
Athanasia Koutsouris ◽  
Jennifer L. Roxas ◽  
Gail Hecht
Keyword(s):  
Cell Death ◽  
Tight Junction ◽  
Host Cell ◽  
Barrier Function ◽  
Type Iii Secretion ◽  
Host Cells ◽  
Epithelial Barrier ◽  
Type Iii ◽  
Barrier Disruption ◽  
Host Cell Death

Enteropathogenic Escherichia coli (EPEC) is a diarrheagenic pathogen that perturbs intestinal epithelial function. Many of the alterations in the host cells are mediated by effector molecules that are secreted directly into epithelial cells by the EPEC type III secretion system. The secreted effector molecule EspF plays a key role in redistributing tight junction proteins and altering epithelial barrier function. EspF has also been shown to localize to mitochondria and trigger membrane depolarization and eventual host cell death. The relationship, if any, between EspF-induced host cell death and epithelial barrier disruption is presently not known. Site-directed mutation of leucine 16 (L16E) of EspF impairs both mitochondrial localization and consequent host cell death. Although the mutation lies within a region critical for type III secretion, EspF(L16E) is secreted efficiently from EPEC. Despite its inability to promote cell death, EspF(L16E) was not impaired for tight junction alteration or barrier disruption. Consistent with this, the pan-caspase inhibitor Q-VD-OPH, despite reducing EPEC-induced host cell death, had no effect on infection-mediated barrier function alteration. Thus EPEC alters the epithelial barrier independent of its ability to induce host cell death.

scholarly journals Host Cell Death due to EnteropathogenicEscherichia coli Has Features of Apoptosis

1999 ◽  
Vol 67 (5) ◽  
pp. 2575-2584 ◽  
Author(s):  
John K. Crane ◽  
Swastika Majumdar ◽  
Donald F. Pickhardt
Keyword(s):  
Cell Death ◽  
Host Cell ◽  
Propidium Iodide ◽  
Tyrosine Kinases ◽  
Enteric Bacteria ◽  
Host Cells ◽  
Watery Diarrhea ◽  
Iodide Uptake ◽  
Host Cell Death

ABSTRACT Enteropathogenic Escherichia coli (EPEC) is a cause of prolonged watery diarrhea in children in developing countries. The ability of EPEC to kill host cells was investigated in vitro in assays using two human cultured cell lines, HeLa (cervical) and T84 (colonic). EPEC killed epithelial cells as assessed by permeability to the vital dyes trypan blue and propidium iodide. In addition, EPEC triggered changes in the host cell, suggesting apoptosis as the mode of death; such changes included early expression of phosphatidylserine on the host cell surface and internucleosomal cleavage of host cell DNA. Genistein, an inhibitor of tyrosine kinases, and wortmannin, an inhibitor of host phosphatidylinositol 3-kinase, markedly increased EPEC-induced cell death and enhanced the features of apoptosis. EPEC-induced cell death was contact dependent and required adherence of live bacteria to the host cell. A quantitative assay for EPEC-induced cell death was developed by using the propidium iodide uptake method adapted to a fluorescence plate reader. With EPEC, the rate and extent of host cell death were less that what has been reported forSalmonella, Shigella, and Yersinia, three other genera of enteric bacteria known to cause apoptosis. However, rapid apoptosis of the host cell may not favor the pathogenic strategy of EPEC, a mucosa-adhering, noninvasive pathogen.

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