scholarly journals Staphylococcus aureus α-toxin induces acid sphingomyelinase release from a human endothelial cell line

2021 ◽  
Author(s):  
David Krones ◽  
Marcel Rühling ◽  
Katrin Anne Becker ◽  
Tobias C. Kunz ◽  
Carolin Sehl ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Plasma Membrane ◽  
Extracellular Fluid ◽  
Fluid Phase ◽  
Cell Types ◽  
Acid Sphingomyelinase ◽  
Host Cells ◽  
Membrane Repair ◽  
Membrane Area

AbstractStaphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (α-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal α-toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus α-toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal α-toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.

scholarly journals Staphylococcus aureus α-Toxin Induces Acid Sphingomyelinase Release From a Human Endothelial Cell Line

2021 ◽  
Vol 12 ◽  
Author(s):  
David Krones ◽  
Marcel Rühling ◽  
Katrin Anne Becker ◽  
Tobias C. Kunz ◽  
Carolin Sehl ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Plasma Membrane ◽  
Extracellular Fluid ◽  
Fluid Phase ◽  
Cell Types ◽  
Acid Sphingomyelinase ◽  
Host Cells ◽  
Membrane Repair ◽  
Membrane Area

Staphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (α-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal α-toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus α-toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal α-toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.

scholarly journals Staphylococcus aureus Alpha-Toxin Disrupts Endothelial-Cell Tight Junctions via Acid Sphingomyelinase and Ceramide

2017 ◽  
Vol 86 (1) ◽  
Author(s):  
Katrin Anne Becker ◽  
Björn Fahsel ◽  
Hannes Kemper ◽  
Joelina Mayeres ◽  
Cao Li ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Tight Junctions ◽  
Molecular Mechanisms ◽  
Acid Sphingomyelinase ◽  
Host Cells ◽  
Lung Edema ◽  
Alpha Toxin ◽  
Content Type

ABSTRACTStaphylococcus aureus(S. aureus) infections are among the most common and severe infections, garnering notoriety in an era of increasing resistance to antibiotics. It is therefore important to define molecular mechanisms by which this pathogen attacks host cells. Here, we demonstrate that alpha-toxin, one of the major toxins ofS. aureus, induces activation of acid sphingomyelinase and concomitant release of ceramide in endothelial cells treated with the toxin. Activation of acid sphingomyelinase by alpha-toxin is mediated via ADAM10. Infection experiments employing alpha-toxin-deficientS. aureusand the corresponding wild-type strain reveal that activation of acid sphingomyelinase in endothelial cells requires alpha-toxin expression by the pathogen. Activation of acid sphingomyelinase is linked to degradation of tight junctions in endothelial cellsin vitro, which is blocked by pharmacological inhibition of acid sphingomyelinase. Most importantly, alpha-toxin induces severe degradation of tight junctions in the lung and causes lung edemain vivo, which is prevented by genetic deficiency of acid sphingomyelinase. These data indicate a novel and important role of the acid sphingomyelinase/ceramide system for the endothelial response to toxins and provide a molecular link between alpha-toxin and the degradation of tight junctions. The data also suggest that inhibition of acid sphingomyelinase may provide a novel treatment option to prevent lung edema caused byS. aureusalpha-toxin.

scholarly journals Trypanosoma cruzi subverts the sphingomyelinase-mediated plasma membrane repair pathway for cell invasion

2011 ◽  
Vol 208 (5) ◽  
pp. 909-921 ◽  
Author(s):  
Maria Cecilia Fernandes ◽  
Mauro Cortez ◽  
Andrew R. Flannery ◽  
Christina Tam ◽  
Renato A. Mortara ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Trypanosoma Cruzi ◽  
Host Cell ◽  
Cell Invasion ◽  
Acid Sphingomyelinase ◽  
Host Cells ◽  
Cell Contact ◽  
Membrane Repair ◽  
Membrane Injury ◽  
Plasma Membrane Repair

Upon host cell contact, the protozoan parasite Trypanosoma cruzi triggers cytosolic Ca2+ transients that induce exocytosis of lysosomes, a process required for cell invasion. However, the exact mechanism by which lysosomal exocytosis mediates T. cruzi internalization remains unclear. We show that host cell entry by T. cruzi mimics a process of plasma membrane injury and repair that involves Ca2+-dependent exocytosis of lysosomes, delivery of acid sphingomyelinase (ASM) to the outer leaflet of the plasma membrane, and a rapid form of endocytosis that internalizes membrane lesions. Host cells incubated with T. cruzi trypomastigotes are transiently wounded, show increased levels of endocytosis, and become more susceptible to infection when injured with pore-forming toxins. Inhibition or depletion of lysosomal ASM, which blocks plasma membrane repair, markedly reduces the susceptibility of host cells to T. cruzi invasion. Notably, extracellular addition of sphingomyelinase stimulates host cell endocytosis, enhances T. cruzi invasion, and restores normal invasion levels in ASM-depleted cells. Ceramide, the product of sphingomyelin hydrolysis, is detected in newly formed parasitophorous vacuoles containing trypomastigotes but not in the few parasite-containing vacuoles formed in ASM-depleted cells. Thus, T. cruzi subverts the ASM-dependent ceramide-enriched endosomes that function in plasma membrane repair to infect host cells.

scholarly journals Plasma membrane integrity in health and disease: significance and therapeutic potential

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Catarina Dias ◽  
Jesper Nylandsted
Keyword(s):  
Plasma Membrane ◽  
Therapeutic Potential ◽  
Calcium Influx ◽  
Membrane Integrity ◽  
Cell Types ◽  
Physical Parameters ◽  
Membrane Repair ◽  
Plasma Membrane Integrity ◽  
Repair Mechanisms ◽  
And Function

AbstractMaintenance of plasma membrane integrity is essential for normal cell viability and function. Thus, robust membrane repair mechanisms have evolved to counteract the eminent threat of a torn plasma membrane. Different repair mechanisms and the bio-physical parameters required for efficient repair are now emerging from different research groups. However, less is known about when these mechanisms come into play. This review focuses on the existence of membrane disruptions and repair mechanisms in both physiological and pathological conditions, and across multiple cell types, albeit to different degrees. Fundamentally, irrespective of the source of membrane disruption, aberrant calcium influx is the common stimulus that activates the membrane repair response. Inadequate repair responses can tip the balance between physiology and pathology, highlighting the significance of plasma membrane integrity. For example, an over-activated repair response can promote cancer invasion, while the inability to efficiently repair membrane can drive neurodegeneration and muscular dystrophies. The interdisciplinary view explored here emphasises the widespread potential of targeting plasma membrane repair mechanisms for therapeutic purposes.

scholarly journals Structural, Molecular, and Functional Alterations of the Blood-Brain Barrier during Epileptogenesis and Epilepsy: A Cause, Consequence, or Both?

2020 ◽  
Vol 21 (2) ◽  
pp. 591 ◽  
Author(s):  
Wolfgang Löscher ◽  
Alon Friedman
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Defense Mechanism ◽  
Extracellular Fluid ◽  
Cell Types ◽  
Therapeutic Interventions ◽  
Brain Barrier ◽  
Transport Systems ◽  
Blood Brain ◽  
The Brain

The blood-brain barrier (BBB) is a dynamic, highly selective barrier primarily formed by endothelial cells connected by tight junctions that separate the circulating blood from the brain extracellular fluid. The endothelial cells lining the brain microvessels are under the inductive influence of neighboring cell types, including astrocytes and pericytes. In addition to the anatomical characteristics of the BBB, various specific transport systems, enzymes and receptors regulate molecular and cellular traffic across the BBB. While the intact BBB prevents many macromolecules and immune cells from entering the brain, following epileptogenic brain insults the BBB changes its properties. Among BBB alterations, albumin extravasation and diapedesis of leucocytes from blood into brain parenchyma occur, inducing or contributing to epileptogenesis. Furthermore, seizures themselves may modulate BBB functions, permitting albumin extravasation, leading to activation of astrocytes and the innate immune system, and eventually modifications of neuronal networks. BBB alterations following seizures are not necessarily associated with enhanced drug penetration into the brain. Increased expression of multidrug efflux transporters such as P-glycoprotein likely act as a ‘second line defense’ mechanism to protect the brain from toxins. A better understanding of the complex alterations in BBB structure and function following seizures and in epilepsy may lead to novel therapeutic interventions allowing the prevention and treatment of epilepsy as well as other detrimental neuro-psychiatric sequelae of brain injury.

scholarly journals Lysosomal Fusion Is Essential for the Retention of Trypanosoma cruzi Inside Host Cells

2004 ◽  
Vol 200 (9) ◽  
pp. 1135-1143 ◽  
Author(s):  
Luciana O. Andrade ◽  
Norma W. Andrews
Keyword(s):  
Plasma Membrane ◽  
Life Cycle ◽  
Trypanosoma Cruzi ◽  
Parasitophorous Vacuole ◽  
Cell Types ◽  
Significant Fraction ◽  
Host Cells ◽  
Productive Infection ◽  
Kinase Inhibition

Trypomastigotes, the highly motile infective forms of Trypanosoma cruzi, are capable of infecting several cell types. Invasion occurs either by direct recruitment and fusion of lysosomes at the plasma membrane, or through invagination of the plasma membrane followed by intracellular fusion with lysosomes. The lysosome-like parasitophorous vacuole is subsequently disrupted, releasing the parasites for replication in the cytosol. The role of this early residence within lysosomes in the intracellular cycle of T. cruzi has remained unclear. For several other cytosolic pathogens, survival inside host cells depends on an early escape from phagosomes before lysosomal fusion. Here, we show that when lysosome-mediated T. cruzi invasion is blocked through phosophoinositide 3-kinase inhibition, a significant fraction of the internalized parasites are not subsequently retained inside host cells for a productive infection. A direct correlation was observed between the lysosomal fusion rates after invasion and the intracellular retention of trypomastigotes. Thus, formation of a parasitophorous vacuole with lysosomal properties is essential for preventing these highly motile parasites from exiting host cells and for allowing completion of the intracellular life cycle.

scholarly journals Exocytosis of acid sphingomyelinase by wounded cells promotes endocytosis and plasma membrane repair

2010 ◽  
Vol 189 (6) ◽  
pp. 1027-1038 ◽  
Author(s):  
Christina Tam ◽  
Vincent Idone ◽  
Cecilia Devlin ◽  
Maria Cecilia Fernandes ◽  
Andrew Flannery ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lysosomal Enzyme ◽  
Membrane Integrity ◽  
Acid Sphingomyelinase ◽  
Membrane Repair ◽  
Membrane Injury ◽  
Cellular Survival ◽  
Plasma Membrane Repair ◽  
Membrane Resealing ◽  
Niemann Pick

Rapid plasma membrane resealing is essential for cellular survival. Earlier studies showed that plasma membrane repair requires Ca2+-dependent exocytosis of lysosomes and a rapid form of endocytosis that removes membrane lesions. However, the functional relationship between lysosomal exocytosis and the rapid endocytosis that follows membrane injury is unknown. In this study, we show that the lysosomal enzyme acid sphingomyelinase (ASM) is released extracellularly when cells are wounded in the presence of Ca2+. ASM-deficient cells, including human cells from Niemann-Pick type A (NPA) patients, undergo lysosomal exocytosis after wounding but are defective in injury-dependent endocytosis and plasma membrane repair. Exogenously added recombinant human ASM restores endocytosis and resealing in ASM-depleted cells, suggesting that conversion of plasma membrane sphingomyelin to ceramide by this lysosomal enzyme promotes lesion internalization. These findings reveal a molecular mechanism for restoration of plasma membrane integrity through exocytosis of lysosomes and identify defective plasma membrane repair as a possible component of the severe pathology observed in NPA patients.

scholarly journals Involvement of Raft-Like Plasma Membrane Domains of Entamoeba histolytica in Pinocytosis and Adhesion

2004 ◽  
Vol 72 (9) ◽  
pp. 5349-5357 ◽  
Author(s):  
Richard C. Laughlin ◽  
Glen C. McGugan ◽  
Rhonda R. Powell ◽  
Brenda H. Welter ◽  
Lesly A. Temesvari
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Entamoeba Histolytica ◽  
Fluid Phase ◽  
Cell Types ◽  
Cysteine Proteases ◽  
Membrane Domains ◽  
Cell Surface Molecule ◽  
Physiological Relevance ◽  
Plasma Membrane Domains

ABSTRACT Lipid rafts are highly ordered, cholesterol-rich, and detergent-resistant microdomains found in the plasma membrane of many eukaryotic cells. These domains play important roles in endocytosis, secretion, and adhesion in a variety of cell types. The parasitic protozoan Entamoeba histolytica, the causative agent of amoebic dysentery, was determined to have raft-like plasma membrane domains by use of fluorescent lipid analogs that specifically partition into raft and nonraft regions of the membrane. Disruption of raft-like membrane domains in Entamoeba with the cholesterol-binding agents filipin and methyl-β-cyclodextrin resulted in the inhibition of several important virulence functions, fluid-phase pinocytosis, and adhesion to host cell monolayers. However, disruption of raft-like domains did not inhibit constitutive secretion of cysteine proteases, another important virulence function of Entamoeba. Flotation of the cold Triton X-100-insoluble portion of membranes on sucrose gradients revealed that the heavy, intermediate, and light subunits of the galactose-N-acetylgalactosamine-inhibitible lectin, an important cell surface adhesion molecule of Entamoeba, were enriched in cholesterol-rich (raft-like) fractions, whereas EhCP5, another cell surface molecule, was not enriched in these fractions. The subunits of the lectin were also observed in high-density, actin-rich fractions of the sucrose gradient. Together, these data suggest that pinocytosis and adhesion are raft-dependent functions in this pathogen. This is the first report describing the existence and physiological relevance of raft-like membrane domains in E. histolytica.

scholarly journals CD209L/L-SIGN and CD209/DC-SIGN act as receptors for SARS-CoV-2 and are differentially expressed in lung and kidney epithelial and endothelial cells

2020 ◽  
Author(s):  
Razie Amraei ◽  
Wenqing Yin ◽  
Marc A. Napoleon ◽  
Ellen L. Suder ◽  
Jacob Berrigan ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Vascular System ◽  
Virus Entry ◽  
Antiviral Drug ◽  
Cell Types ◽  
Host Cells ◽  
Human Endothelial Cells ◽  
High Importance ◽  
Lung And Kidney ◽  
Low Expression

AbstractAs the COVID-19 pandemic continues to spread, investigating the processes underlying the interactions between SARS-CoV-2 and its hosts is of high importance. Here, we report the identification of CD209L/L-SIGN and the related protein CD209/DC-SIGN as receptors capable of mediating SARS-CoV-2 entry into human cells. Immunofluorescence staining of human tissues revealed prominent expression of CD209L in the lung and kidney epithelium and endothelium. Multiple biochemical assays using a purified recombinant SARS-CoV-2 spike receptor binding domain (S-RBD) and ectopically expressed CD209L and CD209 revealed that CD209L and CD209 interact with S-RBD. CD209L contains two N-glycosylation sequons, at sites N92 and N361, but only site N92 is occupied. Removal of the N-glycosylation at this site enhances the binding of S-RBD with CD209L. CD209L also interacts with ACE2, suggesting a role for heterodimerization of CD209L and ACE2 in SARS-CoV-2 entry and infection in cell types where both are present. Furthermore, we demonstrate that human endothelial cells are permissive to SARS-CoV-2 infection and interference with CD209L activity by knockdown strategy or with soluble CD209L inhibits virus entry. Our observations demonstrate that CD209L and CD209 serve as alternative receptors for SARS-CoV-2 in disease-relevant cell types, including the vascular system. This property is particularly important in tissues where ACE2 has low expression or is absent, and may have implications for antiviral drug development.SignificanceUnderstanding the interactions between SARS-CoV-2 with host cells is of high importance. ACE2 is recognized as a major entry receptor, but SARS-CoV-2 may also employ alternative receptors for cell entry and these may hold the key to infection in tissues, where ACE2 has a low expression level or is absent. We identify CD209L/L-SIGN and CD209/DC-SIGN as receptors for SARS-CoV-2. We show that CD209L is N-glycosylated and this modification modulates the binding of CD209L with spike protein. CD209L interacts with ACE2, suggesting that CD209L and ACE2 could function as co-receptors for SARS-CoV-2 entry and infection. Human endothelial cells are permissive to SARS-CoV-2 infection. We show that interfering with CD209L activity in endothelial cells by knockdown or with introduction of soluble CD209L inhibits virus entry, suggesting a novel target for development of antiviral drugs.

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