Biology subjects

Cohen, H.

Publications and source records attributed to Cohen, H..

24 records · Page 2Linked to original sources

bioRxiv2022

T cell response following anti COVID-19 BNT162b2 vaccination is maintained against the SARS-CoV-2 Omicron B.1.1.529 variant of concern

The progression of the COVID-19 pandemic leads to the emergence of variants of concern (VOC), which may compromise the efficacy of the currently administered vaccines. Antigenic drift can potentially bring about a reduced protective T cell immunity and consequently to more severe disease manifestations. To assess this possibility, the T cell responses to the wild-type, Wuhan-1 SARS-CoV-2 ancestral spike protein and Omicron B.1.1.529 spike protein were compared. Accordingly, peripheral blood mononuclear cells (PBMC) were collected from 8 healthy volunteers 4-5 months following a third vaccination with BNT162b2, and stimulated with overlapping peptide libraries representing the spike of either the ancestral or Omicron SARS-CoV- 2 virus variants. Quantification of the specific T cells was carried out by a fluorescent ELISPOT assay, monitoring interferon-gamma (IFNg), interleukin-10 (IL-10) and interleukin-4 (IL-4) secreting cells. For all the examined individuals, comparable level of reactivity to both forms of spike protein were determined. In addition, a dominant Th1 response was observed, manifested mainly by IFNg secreting cells and only limited numbers of IL-10 and IL-4 secreting cells. The data demonstrates a stable T cell activity to the emerging Omicron variant in the tested individuals, therefore the protective immunity to the variant following BNT162b2 vaccination is not significantly affected.

immunology↗

bioRxiv2021

Intracellular Salmonella Paratyphi A is motile and differs in the expression of flagella-chemotaxis, SPI-1 and carbon utilization pathways in comparison to Intracellular S. Typhimurium

Although Salmonella Typhimurium (STM) and Salmonella Paratyphi A (SPA) belong to the same phylogenetic species, share large portion of their genome and express many common virulence factors, they differ vastly in their host specificity, the immune response they elicit, and the clinical manifestations they cause. In this work, we compared for the first time their intracellular trascriptomic architecture and cellular phenotypes during epithelial cell infection. While transcription induction of many metal transport systems, purines, biotin, PhoPQ and SPI-2 regulons was common in both intracellular SPA and STM, we identified 234 differentially expressed genes that showed distinct expression patterns in intracellular SPA vs. STM. Surprisingly, clear expression differences were found in SPI-1, motility and chemotaxis, and carbon (mainly citrate, galactonate and ethanolamine) utilization pathways, indicating that these pathways are regulated and possibly function differently, during their intracellular phase. Moreover, we show that the induction of flagella genes by intracellular SPA leads to cytosolic motility, a conserved trait specific to SPA. To the best of our knowledge, this is the first report of a flagellum-dependent intracellular motility of any Salmonella serovar in living host cells. Importantly, we demonstrate that the elevated expression of SPI-1 and motility genes by intracellular SPA results in increased invasiveness of SPA, following exit from host cells. We propose that such changes prime SPA towards new cycles of host cell infection and contribute to the ability of SPA to disseminate beyond the intestinal lamina propria of the human host, during enteric fever. IMPORTANCESalmonella enterica is a ubiquitous, facultative intracellular animal and human pathogen. Although non-typhoidal Salmonella (NTS) and typhoidal Salmonella serovars belong to the same phylogenetic species and share many virulence factors, the disease they cause in humans is very different. While the underlying mechanisms for these differences are not fully understood, one possible reason expected to contribute to their different pathogenicity is a distinct expression pattern of genes involved in host-pathogen interactions. Here, we compared the global gene expression and the intracellular behavior, during epithelial cell infection of S. Paratyphi A (SPA) and S. Typhimurium (STM), as prototypical serovars of typhoidal and NTS, respectively. Interestingly, we identified different expression patterns in key virulence and metabolic pathways, together with intracellular motility and increased invasiveness of SPA, following exit from infected cells. We hypothesize that these differences are pivotal to the invasive and systemic disease developed following SPA infection in humans.

microbiology↗

bioRxiv2021

Nonclinical Safety and Immunogenicity of an rVSV-ΔG-SARS-CoV-2-S vaccine in mice, hamsters, rabbits and pigs

rVSV-{Delta}G-SARS-CoV-2-S is a clinical stage (Phase 2) replication competent recombinant vaccine against SARS-CoV-2. Nonclinical safety, immunogenicity and efficacy studies were conducted in 4 animal species, using multiple dose levels (up to 108 PFU/animal) and various dosing regimens. There were no treatment related mortalities in any study, or any noticeable clinical signs. Compared to unvaccinated controls, hematology and biochemistry parameters were unremarkable and no adverse histopathological findings gave cause for safety concern in any of the studies. There was no viral shedding in urine, nor viral RNA detected in whole blood or serum samples 7 days post vaccination. The rVSV-{Delta}G-SARS-CoV-2-S vaccine immune response gave rise to neutralizing antibodies, cellular immune response, and increased lymphocytic cellularity in the spleen germinal centers and regional lymph node. No evidence for neurovirulence was found in C57BL/6 immune competent mice or in highly sensitive IFNAR KO mice. Vaccine virus replication and distribution in K18 hACE2 transgenic mice showed a gradual clearance from the vaccination site with no vaccine virus recovered from the lungs. The rVSV-{Delta}G-SARS-CoV-2-S vaccine was well tolerated locally and systemically and elicited an effective immunogenic response up to the highest dose tested, supporting further clinical development.

pharmacology and toxicology↗

bioRxiv2020

Oxidative stress pathogenically remodels the cardiac myocyte cytoskeleton via structural alterations to the microtubule lattice

In the failing heart, the cardiac myocyte microtubule network is remodeled, which increases cellular stiffness and disrupts contractility, contributing to heart failure and death. However, the origins of this deleterious cytoskeletal reorganization are unknown. We now find that oxidative stress, a condition characteristic of failing heart cells, leads to cysteine oxidation of microtubules. Further, our electron and fluorescence microscopy experiments revealed regions of structural damage within the oxidized microtubule lattice. These damaged regions led to the lengthening, realignment, and acetylation of dynamic microtubules within cardiac myocytes. Thus, we found that oxidative stress acts inside of cardiac myocytes to facilitate a dramatic, pathogenic shift from a dynamic, multifaceted microtubule network into a highly acetylated, longitudinally aligned, and static microtubule network. Our results demonstrate how a disease condition characterized by oxidative stress can trigger a molecular oxidation event, which propagates a toxic cellular-scale transformation of the cardiac myocyte microtubule network.Competing Interest StatementThe authors have declared no competing interest.View Full Text

cell biology↗

bioRxiv2020

Proteomic analysis of necroptotic extracellular vesicles

Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.

cell biology↗

bioRxiv2019

Vibrio pore-forming leukocidin activates pyroptotic cell death via the NLRP3 inflammasome

Cell death mechanisms are central to combat infectious microbes and to drive pathological inflammation. One such mechanism, the inflammasome, controls infection through either activation of caspase-1 and the subsequent secretion of the mature pro-inflammatory cytokine, interleukin 1{beta} (IL-1{beta}), or by stopping the dissemination of intracellular pathogens by inducing pyroptotic cell death in infected cells. Hemolysins, which are pore-forming toxins (PFTs), target the host cell plasma membrane by producing pores with different diameters. These pores alter the permeability of the target membrane, often leading to cell death. We previously discovered a functional and potent pore-forming, leukocidin domain-containing hemolysin produced by the Gram-negative marine bacterium Vibrio proteolyticus (V. proteolyticus), termed VPRH. Although leukocidin domains are found in other known PFTs, VPRH constitutes a distinct, understudied class within the leukocidin superfamily. Since PTFs of other pathogens were shown to induce cell death by activating the inflammasome pathway, we hypothesized that VPRH-induced cell death is mediated by direct activation of the inflammasome in mammalian immune host cells. Indeed, we found that VPRH induced a two-step cell death in primary macrophages. The first, a rapid step, was mediated by activating the NLRP3 inflammasome, leading to caspase-1 activation and GSDMD cleavage that resulted in IL-1{beta} secretion and pyroptotic cell death. The second step was independent of the inflammasome; however, its mechanism remains unknown. This study sets the foundation for better understanding the immunological consequences of inflammasome activation by a new leukocidin class of toxins.

immunology↗