Shigella-induced necrosis and apoptosis of U937 cells and J774 macrophages

Microbiology ◽  
2003 ◽  
Vol 149 (9) ◽  
pp. 2513-2527 ◽  
Author(s):  
Takashi Nonaka ◽  
Taku Kuwabara ◽  
Hitomi Mimuro ◽  
Asaomi Kuwae ◽  
Shinobu Imajoh-Ohmi
Keyword(s):  
Cell Death ◽  
Cell Membrane ◽  
Host Cell ◽  
Caspase 3 ◽  
Cytochalasin D ◽  
Host Cells ◽  
U937 Cells ◽  
Host Cell Membrane ◽  
Caspase 1 ◽  
Infected Cells

It is currently unclear whether Shigella kills its phagocytic host cells by apoptosis or necrosis. This study shows that rapid necrosis ensues in macrophage-like cell lines (U937 cells differentiated by all-trans-retinoic acid and J774 cells) infected with the Shigella flexneri strain YSH6000. The infected cells rapidly lose membrane integrity, a typical feature of necrosis, as indicated by the release of the cytoplasmic lactate dehydrogenase and the exposure of phosphatidylserine (PS) associated with the rapid uptake of propidium iodide (PI). The infected cells exhibit DNA fragmentation without nuclear condensation, and substantial involvement of either caspase-3/-7 or caspase-1 was not detected, which is also contrary to what is normally observed in apoptosis. Cytochalasin D potently inhibited Shigella-induced cell death, indicating that only internalized Shigella can cause necrosis. Osmoprotectants such as polyethylene glycols could suppress cell death, suggesting that insertion of a pore by Shigella into the host cell membrane induces the necrosis. The pore was estimated to be 2·87±0·4 nm in diameter. Shigella was also found to be able to induce apoptosis but only in one of the lines tested and under specific conditions, namely U937 cells differentiated with interferon-γ (U937IFN). Caspase-3/-7 but not caspase-1 activation was observed in these infected cells and the exposure of PS occurred without the uptake of PI. An avirulent Shigella strain, wild-type Shigella killed with gentamicin, and even Escherichia coli strain JM109, could also induce apoptosis in U937IFN cells, and cytochalasin D could not prevent apoptosis. It appears therefore that Shigella-induced apoptosis of U937IFN cells is unrelated to Shigella pathogenicity and does not require bacterial internalization. Thus, Shigella can induce rapid necrosis of macrophage-like cells in a virulence-related manner by forming pores in the host cell membrane while some cells can be killed through apoptosis in a virulence-independent fashion.

scholarly journals Characterization of Stages of Hepatozoon americanum and of Parasitized Canine Host Cells

2005 ◽  
Vol 42 (6) ◽  
pp. 788-796 ◽  
Author(s):  
C. A. Cummings ◽  
R. J. Panciera ◽  
K. M. Kocan ◽  
J. S. Mathew ◽  
S. A. Ewing
Keyword(s):  
Cell Membrane ◽  
Host Cell ◽  
Striated Muscle ◽  
Parasitophorous Vacuole ◽  
Protozoan Parasite ◽  
Host Cells ◽  
Blood Leukocytes ◽  
Host Cell Membrane ◽  
Asexual Multiplication ◽  
Hepatozoon Americanum

American canine hepatozoonosis is caused by Hepatozoon americanum, a protozoan parasite, the definitive host of which is the tick, Amblyomma maculatum. Infection of the dog follows ingestion of ticks that harbor sporulated H. americanum oocysts. Following penetration of the intestinal mucosa, sporozoites are disseminated systemically and give rise to extensive asexual multiplication in cells located predominantly in striated muscle. The parasitized canine cells in “onion skin” cysts and in granulomas situated within skeletal muscle, as well as those in peripheral blood leukocytes (PBL), were identified as macrophages by use of fine structure morphology and/or immunohistochemical reactivity with macrophage markers. Additionally, two basic morphologic forms of the parasite were observed in macrophages of granulomas and PBLs. The forms were presumptively identified as merozoites and gamonts. The presence of a “tail” in some gamonts in PBLs indicated differentiation toward microgametes. Recognition of merozoites in PBLs supports the contention that hematogenously redistributed merozoites initiate repeated asexual cycles and could explain persistence of infection for long periods in the vertebrate host. Failure to clearly demonstrate a host cell membrane defining a parasitophorous vacuole may indicate that the parasite actively penetrates the host cell membrane rather than being engulfed by the host cell, as is characteristic of some protozoans.

scholarly journals The NADase-Negative Variant of the Streptococcus pyogenes Toxin NAD + Glycohydrolase Induces JNK1-Mediated Programmed Cellular Necrosis

mBio ◽  
2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Sukantha Chandrasekaran ◽  
Michael G. Caparon
Keyword(s):  
Cell Death ◽  
Epithelial Cells ◽  
Host Cell ◽  
Streptococcus Pyogenes ◽  
Host Cells ◽  
Programmed Necrosis ◽  
Nad Glycohydrolase ◽  
Streptolysin O ◽  
Different Types ◽  
Infected Cells

ABSTRACT Virulence factors are often multifunctional and contribute to pathogenesis through synergistic mechanisms. For the human pathogen Streptococcus pyogenes , two factors that act synergistically are the S. pyogenes NAD + glycohydrolase (SPN) and streptolysin O (SLO). Through distinct mechanisms, SLO forms pores in host cell membranes and translocates SPN into the host cell cytosol. Two natural variants of SPN exist, one that exhibits NADase activity and one that lacks this function, and both versions are translocated and act in concert with SLO to cause an accelerated death response in epithelial cells. While NADase + SPN is known to trigger a metabolic form of necrosis through the depletion of NAD + , the mechanism by which NADase − SPN induces cell death was unknown. In the studies described here, we examined the pathway of NADase − cell death through analysis of activation patterns of mitogen-activated protein kinases (MAPKs). S. pyogenes infection resulted in activation of members of three MAPK subfamilies (p38, ERK, and JNK). However, only JNK was activated in an SLO-specific manner. NADase − SPN induced necrosis in HeLa epithelial cells associated with depolarization of mitochondrial membranes, activation of NF-κB, and the generation of reactive oxygen species. Remarkably, RNA interference (RNAi) silencing of JNK protected cells from NADase − -SPN-mediated necrosis, suggesting that NADase − SPN triggers a form of programmed necrosis dependent on JNK signaling. Taken together, these data demonstrate that SPN acts with SLO to elicit necrosis through two different mechanisms depending on its NADase activity, i.e., metabolic (NADase + ) or programmed (NADase − ), leading to distinct inflammatory profiles. IMPORTANCE Many bacterial pathogens produce toxins that alter how infected host cells interact with the immune system. For Streptococcus pyogenes , two toxins, a NAD + glycohydrolase (SPN) and streptolysin O (SLO), act in combination to cause infected cells to die. However, there are two natural forms of SPN, and these variants cause dying cells to produce different types of signaling molecules. The NADase + form of SPN kills cells by depleting reserves of NAD + and cellular energy. The other form of SPN lacks this activity (NADase − ); thus, the mechanism by which this variant induces toxicity was unknown. Here, we show that infected cells recognize NADase − SPN through a specific signaling molecule called JNK, which causes these cells to undergo a form of cellular suicide known as programmed necrosis. This helps us to understand how different forms of toxins alter host cell signaling to help S. pyogenes cause different types of diseases.

scholarly journals Knock Out of Cell Death Pathway Components Results in Differential Caspase Expression in Response to HCV Infection

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 144
Author(s):  
Hannah L. Wallace ◽  
Lingyan Wang ◽  
Cassandra Davidson ◽  
Vipin Chelakkot ◽  
Michael Grant ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Death ◽  
Cross Talk ◽  
Caspase 3 ◽  
Caspase Activation ◽  
Knock Out ◽  
Caspase 1 ◽  
Cell Death Pathway ◽  
Infected Cells ◽  
Active Caspase

Introduction: Pyroptosis (inflammatory programmed cell death) is induced after the activation of an inflammasome, ultimately resulting in pore formation and cell lysis. One factor in the pathology associated with chronic hepatitis C virus (HCV) infection is non-inflammatory caspase-3-mediated apoptosis. Our lab has found both apoptosis and pyroptosis occurring in HCV-infected Huh-7.5 cells. In the context of some viral infections, pyroptosis is beneficial to the virus; for others, pyroptosis is believed to represent an innate antiviral response. This study aimed to test the effects of knocking out components of the inflammasome pathway on caspase activation in HCV-infected cells. Methods: FAM-FLICA (Carboxyfluorescein - Fluorochrome Inhibitor of Caspases) probes or antibodies were used to visualize active caspase-1 and active caspase-3 in vitro. Huh-7.5 cells with components of the pyroptotic or apoptotic pathways knocked out (NLRP3, GSDM-D or caspase-3) were used to determine the effects of their absence on the virus and caspase activation using confocal microscopy and flow cytometry. Results: Increased levels of caspase-1 were consistently observed in HCV-infected cells compared to those in uninfected cells, and these levels increased with subsequent days post-infection. The inhibition of inflammasome activation using knock out cell lines induced the differential activation of caspase-1 and caspase-3, with the inhibition of pyroptosis, resulting in a trend towards greater expression of caspase-3, indicative of apoptosis. The inhibition of NLRP3 did not fully stop caspase-1 activation, but it was decreased. The flow cytometry results revealed a small sub-set of cells positive for both caspase-1 and caspase-3. Conclusions: These data confirm the occurrence of pyroptosis in HCV-infected cells and demonstrate the involvement of the NLRP3 inflammasome, although other inflammasome sensors might be involved. Since the inhibition of one cell death pathway resulted in the increased activation of the other, along with the presence of double-positive cells, there may be cross-talk between apoptotic and pyroptotic pathways; the role of this cross-talk during infection remains to be elucidated.

scholarly journals Induction of Epithelial Cell Death Including Apoptosis by Enteropathogenic Escherichia coli Expressing Bundle-Forming Pili

2001 ◽  
Vol 69 (12) ◽  
pp. 7356-7364 ◽  
Author(s):  
Maan Abul-Milh ◽  
Ying Wu ◽  
Bedy Lau ◽  
Clifford A. Lingwood ◽  
Debora Barnett Foster
Keyword(s):  
Escherichia Coli ◽  
Cell Death ◽  
Host Cell ◽  
Morphological Changes ◽  
Laboratory Strain ◽  
Host Cells ◽  
Host Cell Membrane ◽  
Infantile Diarrhea ◽  
Enterocyte Effacement ◽  
Colonic Cells

ABSTRACT Infection with enteropathogenic Escherichia coli (EPEC) is a major cause of severe infantile diarrhea, particularly in parts of the developing world. The bundle-forming pilus (BFP) of EPEC is an established virulence factor encoded on the EPEC adherence factor plasmid (EAF) and has been implicated in both localized adherence to host cells and bacterial autoaggregation. We investigated the role of BFP in the ability of EPEC binding to kill host epithelial cells. BFP-expressing strains killed all three cell lines tested, comprising HEp-2 (laryngeal), HeLa (cervical), and Caco-2 (colonic) cells. Analysis of phosphatidylserine expression, internucleosomal cleavage of host cell DNA, and morphological changes detected by electron microscopy indicated evidence of apoptosis. The extent of cell death was significantly greater for BFP-expressing strains, including E2348/69, a wild-type clinical isolate, as well as for a laboratory strain, HB101, transformed with a bfp-carrying plasmid. Strains which did not express BFP induced significantly less cell death, including a bfpA disruptional mutant of E2348/69, EAF plasmid-cured E2348/69, HB101, and HB101 complemented with the locus of enterocyte effacement pathogenicity island. These results indicate a direct correlation between BFP expression and induction of cell death, including apoptosis, an event which may involve the targeting of host cell membrane phosphatidylethanolamine.

Three-Dimensional Imaging of Toxoplasma gondii–Host Cell Interactions within the Parasitophorous Vacuole

2004 ◽  
Vol 10 (5) ◽  
pp. 580-585 ◽  
Author(s):  
Heide Schatten ◽  
Hans Ris
Keyword(s):  
Toxoplasma Gondii ◽  
Cell Membrane ◽  
Host Cell ◽  
Parasitophorous Vacuole ◽  
Three Dimensional ◽  
Host Cells ◽  
Preparation Technique ◽  
Host Cell Membrane ◽  
Three Dimensional Imaging ◽  
Structural Interactions

The protozoan parasite Toxoplasma gondii is a representative of apicomplexan parasites that invades host cells through an unconventional motility mechanism. During host cell invasion it forms a specialized membrane-surrounded compartment that is called the parasitophorous vacuole. The interactions between the host cell and parasite membranes are complex and recent studies have revealed in more detail that both the host cell and the parasite membrane contribute to the formation of the parasitophorous vacuole. By using our a new specimen preparation technique that allows three-dimensional imaging of thick-sectioned internal cell structures with high-resolution, low-voltage field emission scanning electron microscopy, we were able to visualize continuous structural interactions of the host cell membrane with the parasite within the parasitophorous vacuole. Fibrous and tubular material extends from the host cell membrane and is connected to parasite membrane components. Shorter protrusions are also elaborated from the parasite. Several of these shorter fine protrusions connect to the fibrous material of the host cell membrane. The elaborate network may be used for modifications of the parasitophorous vacuole membrane that will allow utilization of nutrients from the host cell by the parisite while it is being protected from host cell attacks. The structural interactions between parasite and host cells undergo time-dependent changes, and a fission pore is the most prominent structure left connecting the parasite with the host cell. The fission pore is anchored in the host cell by thick structural components of unknown nature. The new information gained with this technique includes structural details of fibrous and tubular material that is continuous between the parasite and host cell and can be imaged in three dimensions. We present this technique as a tool to investigate more fully the complex structural interactions of the host cell and the parasite residing in the parasitophorous vacuole.

scholarly journals Subverting the mechanisms of cell death: flavivirus manipulation of host cell responses to infection

2018 ◽  
Vol 46 (3) ◽  
pp. 609-617 ◽  
Author(s):  
Elisa Vicenzi ◽  
Isabel Pagani ◽  
Silvia Ghezzi ◽  
Sarah L. Taylor ◽  
Timothy R. Rudd ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Surface ◽  
Host Cell ◽  
Host Cells ◽  
Zikv Infection ◽  
Cell Responses ◽  
Dead End ◽  
Disease Aetiology ◽  
Infected Cells ◽  
Host Cell Surface

Viruses exploit host metabolic and defence machinery for their own replication. The flaviviruses, which include Dengue (DENV), Yellow Fever (YFV), Japanese Encephalitis (JEV), West Nile (WNV) and Zika (ZIKV) viruses, infect a broad range of hosts, cells and tissues. Flaviviruses are largely transmitted by mosquito bites and humans are usually incidental, dead-end hosts, with the notable exceptions of YFV, DENV and ZIKV. Infection by flaviviruses elicits cellular responses including cell death via necrosis, pyroptosis (involving inflammation) or apoptosis (which avoids inflammation). Flaviviruses exploit these mechanisms and subvert them to prolong viral replication. The different effects induced by DENV, WNV, JEV and ZIKV are reviewed. Host cell surface proteoglycans (PGs) bearing glycosaminoglycan (GAG) polysaccharides — heparan/chondroitin sulfate (HS/CS) — are involved in initial flavivirus attachment and during the expression of non-structural viral proteins play a role in disease aetiology. Recent work has shown that ZIKV-infected cells are protected from cell death by exogenous heparin (a GAG structurally similar to host cell surface HS), raising the possibility of further subtle involvement of HS PGs in flavivirus disease processes. The aim of this review is to synthesize information regarding DENV, WNV, JEV and ZIKV from two areas that are usually treated separately: the response of host cells to infection by flaviviruses and the involvement of cell surface GAGs in response to those infections.

New permeability pathways induced by the malarial parasite in the membrane of its host erythrocyte: Potential routes for targeting of drugs into infected cells

1987 ◽  
Vol 7 (6) ◽  
pp. 455-463 ◽  
Author(s):  
Hagai Ginsburg ◽  
Wilfred D. Stein
Keyword(s):  
Cell Membrane ◽  
Host Cell ◽  
Malarial Parasite ◽  
Structural Defects ◽  
Transport Systems ◽  
Host Cell Membrane ◽  
Transport Pathways ◽  
Infected Cells ◽  
New Permeability Pathways

Malarial parasites propagate asexually inside the erythrocytes of their vertebrate host. Six hours after invasion, the permeability of the host cell membrane to anions and small nonelectrolytes starts to increase and reaches its peak as the parasite matures. This increased permeability differs from the native transport systems of the normal erythrocyte in its solute selectivity pattern, its enthalpy of activation and its susceptibility to inhibitors, suggesting the appearance of new transport pathways. A biophysical analysis of the permeability data indicates that the selectivity barrier discriminates between permeants according to their hydrogen bonding capacity and has solubilization properties compared to those of iso-butanol. The new permeability pathways could result from structural defects caused in the host cell membrane by the insertion of parasite-derived polypeptides. It is suggested that the unique transport properties of the new pathways be used to target drugs into infected cells, to affect the parasite either directly or through the modulation of the intraerythrocytic environment. The feasibility of drug targeting is demonstrated in in vitro cultures of the human malarial parasite Plasmodium falciparum.

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.

Association of matrix protein of respiratory syncytial virus with the host cell membrane of infected cells

2003 ◽  
Vol 149 (1) ◽  
pp. 199-210 ◽  
Author(s):  
A. Marty ◽  
J. Meanger ◽  
J. Mills ◽  
B. Shields ◽  
R. Ghildyal
Keyword(s):  
Respiratory Syncytial Virus ◽  
Cell Membrane ◽  
Host Cell ◽  
Matrix Protein ◽  
Host Cell Membrane ◽  
Infected Cells ◽  
Syncytial Virus

scholarly journals Candidate screening of host cell membrane proteins involved in SARS-CoV-2 entry

2020 ◽  
Author(s):  
Norihiro Kotani ◽  
Takanari Nakano
Keyword(s):  
Membrane Proteins ◽  
Cell Membrane ◽  
Host Cell ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Host Cells ◽  
Host Cell Membrane ◽  
Cell Membrane Proteins ◽  
Cell Membrane Protein ◽  
Spike Proteins

ABSTRACTCOVID-19 represents a real threat to the global population, and understanding the biological features of the causative virus (SARS-CoV-2) is imperative to aid in mitigating this threat. Analyses of proteins such as primary receptors and co-receptors (co-factors) that are involved in SARS-CoV-2 entry into host cells will provide important clues to help control the virus. Here, we identified host cell membrane protein candidates that were present in proximity to the attachment sites of SARS-CoV-2 spike proteins through the use of proximity labeling and proteomics analysis. The identified proteins represent candidate key factors that may be required for viral entry. Our results indicated that a number of membrane proteins, including DPP4, Cadherin-17, and CD133, were identified to co-localize with cell membrane-bound SARS-CoV-2 spike proteins in Caco-2 cells that were used to expand the SARS-CoV-2 virion. We anticipate that the information regarding these protein candidates will be utilized for the future development of vaccines and antiviral agents against SARS-CoV-2.

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