scholarly journals Temporal omics analysis in Syrian hamsters unravel cellular effector responses to moderate COVID-19

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Geraldine Nouailles ◽  
Emanuel Wyler ◽  
Peter Pennitz ◽  
Dylan Postmus ◽  
Daria Vladimirova ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cytotoxic T Cells ◽  
Transcriptional Response ◽  
Productive Infection ◽  
Cellular Mechanisms ◽  
Syrian Hamsters ◽  
Effector Functions ◽  
Specific Effector ◽  
Early Expression ◽  
Cell Type Specific

AbstractIn COVID-19, immune responses are key in determining disease severity. However, cellular mechanisms at the onset of inflammatory lung injury in SARS-CoV-2 infection, particularly involving endothelial cells, remain ill-defined. Using Syrian hamsters as a model for moderate COVID-19, we conduct a detailed longitudinal analysis of systemic and pulmonary cellular responses, and corroborate it with datasets from COVID-19 patients. Monocyte-derived macrophages in lungs exert the earliest and strongest transcriptional response to infection, including induction of pro-inflammatory genes, while epithelial cells show weak alterations. Without evidence for productive infection, endothelial cells react, depending on cell subtypes, by strong and early expression of anti-viral, pro-inflammatory, and T cell recruiting genes. Recruitment of cytotoxic T cells as well as emergence of IgM antibodies precede viral clearance at day 5 post infection. Investigating SARS-CoV-2 infected Syrian hamsters thus identifies cell type-specific effector functions, providing detailed insights into pathomechanisms of COVID-19 and informing therapeutic strategies.

scholarly journals Longitudinal omics in Syrian hamsters integrated with human data unravel cellular effector responses to moderate COVID-19

2021 ◽  
Author(s):  
Geraldine Nouailles ◽  
Emanuel Wyler ◽  
Peter Pennitz ◽  
Dylan Postmus ◽  
Daria Vladrimirova ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cytotoxic T Cells ◽  
Transcriptional Response ◽  
Productive Infection ◽  
Cellular Mechanisms ◽  
Syrian Hamsters ◽  
Effector Functions ◽  
Specific Effector ◽  
Early Expression ◽  
Cell Type Specific

Abstract In COVID-19, immune responses are key in determining disease severity. However, cellular mechanisms at the onset of inflammatory lung injury in SARS-CoV-2 infection, particularly involving endothelial cells, remain ill-defined. Using Syrian hamsters as model for moderate COVID-19, we conducted a detailed longitudinal analysis of systemic and pulmonary cellular responses, and corroborated it with datasets from COVID-19 patients. Monocyte-derived macrophages in lungs exerted the earliest and strongest transcriptional response to infection, including induction of pro-inflammatory genes, while epithelial cells showed weak activation. Without evidence for productive infection, endothelial cells reacted, depending on cell subtypes, by strong and early expression of anti-viral, pro-inflammatory, and T cell recruiting genes. Recruitment of cytotoxic T cells as well as emergence of IgM antibodies preceded viral clearance at day 5 post infection. Investigating SARS-CoV-2 infected Syrian hamsters can thus identify cell type-specific effector functions, provide detailed insights into pathomechanisms of COVID-19, and inform therapeutic strategies.

scholarly journals Vascular Inflammation Is Associated with Loss of Aquaporin 1 Expression on Endothelial Cells and Increased Fluid Leakage in SARS-CoV-2 Infected Golden Syrian Hamsters

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 639
Author(s):  
Lisa Allnoch ◽  
Georg Beythien ◽  
Eva Leitzen ◽  
Kathrin Becker ◽  
Franz-Josef Kaup ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Vascular Inflammation ◽  
Intercellular Junctions ◽  
Edema Formation ◽  
Vascular Integrity ◽  
Syrian Hamsters ◽  
Aquaporin 1 ◽  
Vascular Walls ◽  
Transmission Electron ◽  
Mock Infection

Vascular changes represent a characteristic feature of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection leading to a breakdown of the vascular barrier and subsequent edema formation. The aim of this study was to provide a detailed characterization of the vascular alterations during SARS-CoV-2 infection and to evaluate the impaired vascular integrity. Groups of ten golden Syrian hamsters were infected intranasally with SARS-CoV-2 or phosphate-buffered saline (mock infection). Necropsies were performed at 1, 3, 6, and 14 days post-infection (dpi). Lung samples were investigated using hematoxylin and eosin, alcian blue, immunohistochemistry targeting aquaporin 1, CD3, CD204, CD31, laminin, myeloperoxidase, SARS-CoV-2 nucleoprotein, and transmission electron microscopy. SARS-CoV-2 infected animals showed endothelial hypertrophy, endothelialitis, and vasculitis. Inflammation mainly consisted of macrophages and lower numbers of T-lymphocytes and neutrophils/heterophils infiltrating the vascular walls as well as the perivascular region at 3 and 6 dpi. Affected vessels showed edema formation in association with loss of aquaporin 1 on endothelial cells. In addition, an ultrastructural investigation revealed disruption of the endothelium. Summarized, the presented findings indicate that loss of aquaporin 1 entails the loss of intercellular junctions resulting in paracellular leakage of edema as a key pathogenic mechanism in SARS-CoV-2 triggered pulmonary lesions.

Platelets and Endothelial Cells Act in Concert to Delay Thrombolysis – Evidence from an In Vitro Model of the Human Occlusive Thrombus

1998 ◽  
Vol 79 (03) ◽  
pp. 602-608 ◽  
Author(s):  
W. G. Jerome ◽  
S. Handt ◽  
R. R. Hantgan
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
In Vitro Model ◽  
Umbilical Vein ◽  
Plasminogen Activator Inhibitor ◽  
Genetically Engineered ◽  
Cellular Mechanisms ◽  
Vitro Model ◽  
Activator Inhibitor

SummaryThe molecular and cellular mechanisms that over a period of hours render a human thrombus progressively resistant to fibrinolysis have been probed with a novel in vitro model. The kinetics of clot formation and fibrinolysis were monitored by laser light scattering with platelet-rich model thrombi contained in cylindrical flow chambers. In selected experiments, human umbilical vein endothelial cells were also cultured to confluence on the inner walls of these “glass blood vessels”. Following an “aging” period (0.5, 2 or 4 h), each thrombus was gently perfused with a bolus of plasminogen/recombinant tissue plasminogen activator to induce fibrinolysis. Platelets delayed lysis of 2 h-aged thrombi by ~70% and (non-stimulated) endothelial cells by ~30%, compared to cell-free control clots. However, even greater lytic delays (~260%) resulted when both vascular cells were present in the same 2 h-aged thrombus. In contrast, rapid lysis was consistently achieved with R298E,R299E t-PA, a genetically engineered plasminogen activator that is insensitive to inhibition by plasminogen activator inhibitor type 1. These observations suggest platelets and endothelial cells act in concert to enrich the fibrin scaffold of an aging human thrombus in plasminogen activator inhibitor. We propose that the presence of both platelets and endothelial cells may contribute to progressive thrombolytic resistance.

Intracellular redox status affects transplasma membrane electron transport in pulmonary arterial endothelial cells

2002 ◽  
Vol 282 (1) ◽  
pp. L36-L43 ◽  
Author(s):  
Marilyn P. Merker ◽  
Robert D. Bongard ◽  
Nicholas J. Kettenhofen ◽  
Yoshiyuki Okamoto ◽  
Christopher A. Dawson
Keyword(s):  
Endothelial Cells ◽  
Electron Transport ◽  
Control Level ◽  
Redox Status ◽  
Cellular Mechanisms ◽  
Control Activity ◽  
Transplasma Membrane Electron Transport ◽  
Cell Extracts ◽  
Pulmonary Arterial ◽  
Arterial Endothelial Cells

Pulmonary arterial endothelial cells possess transplasma membrane electron transport (TPMET) systems that transfer intracellular reducing equivalents to extracellular electron acceptors. As one aspect of determining cellular mechanisms involved in one such TPMET system in pulmonary arterial endothelial cells in culture, glycolysis was inhibited by treatment with iodoacetate (IOA) or by replacing the glucose in the cell medium with 2-deoxy-d-glucose (2-DG). TPMET activity was measured as the rate of reduction of the extracellular electron acceptor polymer toluidine blue O polyacrylamide. Intracellular concentrations of NADH, NAD+, NADPH, and NADP+ were determined by high-performance liquid chromatography of KOH cell extracts. IOA decreased TPMET activity to 47% of control activity concomitant with a decrease in the NADH/NAD+ ratio to 34% of the control level, without a significant change in the NADPH/NADP+ ratio. 2-DG decreased TPMET activity to 53% of control and decreased both NADH/NAD+ and NADPH/NADP+ ratios to 51% and 55%, respectively, of control levels. When lactate was included in the medium along with the inhibitors, the effects of IOA and 2-DG on both TPMET activity and the NADPH/NADP+ ratios were prevented. The results suggest that cellular redox status is a determinant of pulmonary arterial endothelial cell TPMET activity, with TPMET activity more highly correlated with the poise of the NADH/NAD+redox pair.

scholarly journals Activation Associated ERK1/2 Signaling Impairments in CD8+ T Cells Co-Localize with Blunted Polyclonal and HIV-1 Specific Effector Functions in Early Untreated HIV-1 Infection

PLoS ONE ◽  
2013 ◽  
Vol 8 (10) ◽  
pp. e77412 ◽  
Author(s):  
Timothy Q. Crawford ◽  
Fredrick M. Hecht ◽  
Christopher D. Pilcher ◽  
Lishomwa C. Ndhlovu ◽  
Jason D. Barbour
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Effector Functions ◽  
Specific Effector ◽  
Hiv 1

scholarly journals Estimation of nitric oxide generation by endothelial cells EA.hy926 under flow-induced mechanical stress in a microfluidic system

2020 ◽  
pp. 71-77
Author(s):  
А.А. Московцев ◽  
А.Н. Мыльникова ◽  
Д.В. Колесов ◽  
А.А. Микрюкова ◽  
Д.М. Зайченко ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Mechanical Stress ◽  
Microfluidic System ◽  
Hydrodynamic Characteristics ◽  
No Production ◽  
Hydrodynamic Conditions ◽  
Cellular Mechanisms ◽  
Vascular Walls

Эндотелиальные клетки, выстилающие стенки сосудов, преобразовывают деформацию собственных структур, вызванную током крови, в химические сигналы, одним из которых является важный регулятор просвета сосуда - оксид азота (NO). К настоящему моменту накоплен большой объём данных о клеточных механизмах активации продукции NO, однако сведений о динамике генерации оксида азота эндотелиальными клетками в зависимости от гидродинамических условий недостаточно. В этой связи разработка микрофлюидных систем in vitro, имитирующих кровеносное русло, и изучение в них эндотелия в сложных гидродинамических условиях является актуальной задачей. В данной работе для создания контролируемых гидродинамических условий для монослоя эндотелиоцитоподобных клеток EA.hy926 была спроектирована и разработана микрофлюидная система, имитирующая линейные участки микрососудистого русла. Методом непрямого определения содержания оксида азота (II) NO с использованием флуоресцентного зонда 4,5-диаминофлуоресцеина DAF-2 впервые получены данные об увеличении продукции NO клетками EA.hy926 при механическом стрессе, создаваемом потоком ростовой среды. Представлены расчетные гидродинамические характеристики микрофлюидной системы, а также методика измерения продукции NO. Возможность исследования функциональной активности эндотелия позволяет использовать разработанную микрофлюидную модельную систему как для изучения клеточно-автономных регуляторных свойств эндотелия при действии ряда вазоактивных фармакологических препаратов и других методов воздействия на эндотелий, так и при моделируемой дисфункции эндотелия. Endothelial cells lining vascular walls transform the flow-induced deformation of their own structures into chemical signals, one of which, nitric oxide (NO), is an important regulator of the vascular lumen diameter. By present, a large amount of data on cellular mechanisms for activation of NO production has been accumulated. However, there is insufficient information on changes in endothelial NO generation under different hydrodynamic conditions. Therefore, development of microfluidic systems that model blood vessels in vitro and using them to study the endothelium under complex hydrodynamic conditions are relevant tasks. In this study, a microfluidic system was developed to create controlled hydrodynamic conditions for a monolayer of endotheliocyte-like cells EAhy.926. This system simulates linear sections of the microvasculature. By indirect measurement of NO (II) content with a fluorescent 4,5-diaminofluorescein (DAF-2) probe, we showed an increase in the NO production by EAhy.926 cells under mechanical stress generated by the medium flow. The article presents the method for measuring NO production and the calculated hydrodynamic characteristics of the microfluidic system. The results showed that the developed microfluidic model system is promising for studying cell-autonomous regulatory properties of the endothelium both under the action of vasoactive agents and in simulated endothelial dysfunction.

scholarly journals Cellular tropism of SARS-CoV-2 in the respiratory tract of Syrian hamsters and B6.Cg-Tg(K18-ACE2)2Prlmn/J transgenic mice

2021 ◽  
pp. 030098582110430
Author(s):  
Hui-Ling Yen ◽  
Sophie Valkenburg ◽  
Sin Fun Sia ◽  
Ka Tim Choy ◽  
J. S. Malik Peiris ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Lung Parenchyma ◽  
Productive Infection ◽  
Type I ◽  
Syrian Hamsters ◽  
Transmission Electron ◽  
Type Ii Pneumocytes ◽  
Viral Particles ◽  
Cellular Tropism ◽  
Conducting Airways

Several animal models have been developed to study the pathophysiology of SARS-CoV-2 infection and to evaluate vaccines and therapeutic agents for this emerging disease. Similar to infection with SARS-CoV-1, infection of Syrian hamsters with SARS-CoV-2 results in moderate respiratory disease involving the airways and lung parenchyma but does not lead to increased mortality. Using a combination of immunohistochemistry and transmission electron microscopy, we showed that the epithelium of the conducting airways of hamsters was the primary target for viral infection within the first 5 days of infection, with little evidence of productive infection of pneumocytes. At 6 days postinfection, antigen was cleared but parenchymal damage persisted, and the major pathological changes resolved by day 14. These findings are similar to those previously reported for hamsters with SARS-CoV-1 infection. In contrast, infection of K18-hACE2 transgenic mice resulted in pneumocyte damage, with viral particles and replication complexes in both type I and type II pneumocytes together with the presence of convoluted or cubic membranes; however, there was no evidence of virus replication in the conducting airways. The Syrian hamster is a useful model for the study of SARS-CoV-2 transmission and vaccination strategies, whereas infection of the K18-hCE2 transgenic mouse results in lethal disease with fatal neuroinvasion but with sparing of conducting airways.

scholarly journals Cell type-specific biogenesis of novel vesicles containing viral products in human cytomegalovirus infection

2020 ◽  
Author(s):  
Samina Momtaz ◽  
Belen Molina ◽  
Luwanika Mlera ◽  
Felicia Goodrum ◽  
Jean M. Wilson
Keyword(s):  
Endothelial Cells ◽  
Human Cytomegalovirus ◽  
Biosynthetic Pathway ◽  
Cell Types ◽  
Endocytic Pathway ◽  
Specific Cell ◽  
Cell Type ◽  
Subcellular Compartment ◽  
Increased Risk ◽  
Cell Type Specific

AbstractHuman cytomegalovirus (HCMV), while highly restricted for the human species, infects an unlimited array of cell types in the host. Patterns of infection are dictated by the cell type infected, but cell type-specific factors and how they impact tropism for specific cell types is poorly understood. Previous studies in primary endothelial cells showed that HCMV infection induces large multivesicular-like bodies that incorporate viral products including dense bodies and virions. Here we define the nature of these large vesicles using a recombinant virus where UL32, encoding the pp150 tegument protein, is fused in frame with green fluorescent protein (GFP, TB40/E-UL32-GFP). Cells were fixed and labeled with antibodies against subcellular compartment markers and imaged using confocal and super-resolution microscopy. In fibroblasts, UL32-GFP-positive vesicles were marked with classical markers of MVBs, including CD63 and lysobisphosphatidic acid (LBPA), both classical MVB markers, as well as the clathrin and LAMP1. Unexpectedly, UL32-GFP-positive vesicles in endothelial cells were not labeled by CD63, and LBPA was completely lost from infected cells. We defined these UL32-positive vesicles in endothelial cells using markers for the cis-Golgi (GM130), lysosome (LAMP1), and autophagy (LC3B). These findings suggest that virus-containing MVBs in fibroblasts are derived from the canonical endocytic pathway and takeover classical exosomal release pathway. Virus containing MVBs in HMVECs are derived from the early biosynthetic pathway and exploit a less characterized early Golgi-LAMP1-associated non-canonical secretory autophagy pathway. These results reveal striking cell-type specific membrane trafficking differences in host pathways that are exploited by HCMV.ImportanceHuman cytomegalovirus (HCMV) is a herpesvirus that, like all herpesvirus, that establishes a life long infection. HCMV remains a significant cause of morbidity and mortality in the immunocompromised and HCMV seropositivity is associated with increased risk vascular disease. HCMV infects many cells in the human and the biology underlying the different patterns of infection in different cell types is poorly understood. Endothelial cells are important target of infection that contribute to hematogenous spread of the virus to tissues. Here we define striking differences in the biogenesis of large vesicles that incorporate virions in fibroblasts and endothelial cells. In fibroblasts, HCMV is incorporated into canonical MVBs derived from an endocytic pathway, whereas HCMV matures through vesicles derived from the biosynthetic pathway in endothelial cells. This work defines basic biological differences between these cell types that may impact the outcome of infection.

scholarly journals Molecular and cellular mechanisms underlying potyvirus infection

2014 ◽  
Vol 95 (7) ◽  
pp. 1415-1429 ◽  
Author(s):  
K. I. Ivanov ◽  
K. Eskelin ◽  
A. Lõhmus ◽  
K. Mäkinen
Keyword(s):  
Infected Cell ◽  
Molecular Basis ◽  
Plant Viruses ◽  
Host Factors ◽  
Considerable Progress ◽  
Productive Infection ◽  
Cellular Mechanisms ◽  
Viral Translation ◽  
Normal Functions ◽  
Cellular Organelles

Potyviruses represent one of the most economically important and widely distributed groups of plant viruses. Despite considerable progress towards understanding the cellular and molecular basis of their pathogenicity, many questions remain about the mechanisms by which potyviruses suppress host defences and create an optimal intracellular environment for viral translation, replication, assembly and spread. The review focuses on the multifunctional roles of potyviral proteins and their interplay with various host factors in different compartments of the infected cell. We place special emphasis on the recently discovered and currently putative mechanisms by which potyviruses subvert the normal functions of different cellular organelles in order to establish an efficient and productive infection.

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