Severe acute respiratory coronavirus 2 (SARS‐CoV‐2) causes neurological disease in the peripheral and central nervous system (PNS and CNS, respectively) of some patients. It is not clear whether ...SARS‐CoV‐2 infection or the subsequent immune response are the key factors that cause neurological disease. Here, we addressed this question by infecting human induced pluripotent stem cell‐derived CNS and PNS neurons with SARS‐CoV‐2. SARS‐CoV‐2 infected a low number of CNS neurons and did not elicit a robust innate immune response. On the contrary, SARS‐CoV‐2 infected a higher number of PNS neurons. This resulted in expression of interferon (IFN) λ1, several IFN‐stimulated genes and proinflammatory cytokines. The PNS neurons also displayed alterations characteristic of neuronal damage, as increased levels of sterile alpha and Toll/interleukin receptor motif‐containing protein 1, amyloid precursor protein and α‐synuclein, and lower levels of cytoskeletal proteins. Interestingly, blockade of the Janus kinase and signal transducer and activator of transcription pathway by Ruxolitinib did not increase SARS‐CoV‐2 infection, but reduced neuronal damage, suggesting that an exacerbated neuronal innate immune response contributes to pathogenesis in the PNS. Our results provide a basis to study coronavirus disease 2019 (COVID‐19) related neuronal pathology and to test future preventive or therapeutic strategies.
The SARS-CoV-2 coronavirus has led to a pandemic with millions of people affected. The present study finds that risk-factors for severe COVID-19 disease courses, i.e. male sex, older age and ...sedentary life style are associated with higher prostaglandin E2 (PGE2) serum levels in blood samples from unaffected subjects. In COVID-19 patients, PGE2 blood levels are markedly elevated and correlate positively with disease severity. SARS-CoV-2 induces PGE2 generation and secretion in infected lung epithelial cells by upregulating cyclo-oxygenase (COX)-2 and reducing the PG-degrading enzyme 15-hydroxyprostaglandin-dehydrogenase. Also living human precision cut lung slices (PCLS) infected with SARS-CoV-2 display upregulated COX-2. Regular exercise in aged individuals lowers PGE2 serum levels, which leads to increased Paired-Box-Protein-Pax-5 (PAX5) expression, a master regulator of B-cell survival, proliferation and differentiation also towards long lived memory B-cells, in human pre-B-cell lines. Moreover, PGE2 levels in serum of COVID-19 patients lowers the expression of PAX5 in human pre-B-cell lines. The PGE2 inhibitor Taxifolin reduces SARS-CoV-2-induced PGE2 production. In conclusion, SARS-CoV-2, male sex, old age, and sedentary life style increase PGE2 levels, which may reduce the early anti-viral defense as well as the development of immunity promoting severe disease courses and multiple infections. Regular exercise and Taxifolin treatment may reduce these risks and prevent severe disease courses.
An under-explored target for SARS-CoV-2 is the
-adenosyl methionine (SAM)-dependent methyltransferase Nsp14, which methylates the N7-guanosine of viral RNA at the 5'-end, allowing the virus to evade ...host immune response. We sought new Nsp14 inhibitors with three large library docking strategies. First, up to 1.1 billion lead-like molecules were docked against the enzyme's SAM site, leading to three inhibitors with IC
values from 6 to 50 μM. Second, docking a library of 16 million fragments revealed 9 new inhibitors with IC
values from 12 to 341 μM. Third, docking a library of 25 million electrophiles to covalently modify Cys387 revealed 7 inhibitors with IC
values from 3.5 to 39 μM. Overall, 32 inhibitors encompassing 11 chemotypes had IC
values < 50 μM and 5 inhibitors in 4 chemotypes had IC
values < 10 μM. These molecules are among the first non-SAM-like inhibitors of Nsp14, providing starting points for future optimization.
Hepatitis C virus (HCV) exploits the four entry factors CD81, scavenger receptor class B type I (SR-BI, also known as SCARB1), occludin, and claudin-1 as well as the co-factor epidermal growth factor ...receptor (EGFR) to infect human hepatocytes. Here, we report that the disintegrin and matrix metalloproteinase 10 (ADAM10) associates with CD81, SR-BI, and EGFR and acts as HCV host factor. Pharmacological inhibition, siRNA-mediated silencing and genetic ablation of ADAM10 reduced HCV infection. ADAM10 was dispensable for HCV replication but supported HCV entry and cell-to-cell spread. Substrates of the ADAM10 sheddase including epidermal growth factor (EGF) and E-cadherin, which activate EGFR family members, rescued HCV infection of ADAM10 knockout cells. ADAM10 did not influence infection with other enveloped RNA viruses such as alphaviruses and a common cold coronavirus. Collectively, our study reveals a critical role for the sheddase ADAM10 as a HCV host factor, contributing to EGFR family member transactivation and as a consequence to HCV uptake.
Chikungunya virus (CHIKV) is an arthritogenic reemerging virus replicating in plasma membrane-derived compartments termed "spherules." Here, we identify the human transmembrane protein CD81 as host ...factor required for CHIKV replication. Ablation of CD81 results in decreased CHIKV permissiveness, while overexpression enhances infection. CD81 is dispensable for virus uptake but critically required for viral genome replication. Likewise, murine CD81 is crucial for CHIKV permissiveness and is expressed in target cells such as dermal fibroblasts, muscle and liver cells. Whereas related alphaviruses, including Ross River virus (RRV), Semliki Forest virus (SFV), Sindbis virus (SINV) and Venezuelan equine encephalitis virus (VEEV), also depend on CD81 for infection, RNA viruses from other families, such as coronaviruses, replicate independently of CD81. Strikingly, the replication-enhancing function of CD81 is linked to cholesterol binding. These results define a mechanism exploited by alphaviruses to hijack the membrane microdomain-modeling protein CD81 for virus replication through interaction with cholesterol.
In this study, we discover the tetraspanin CD81 as a host factor for the globally emerging chikungunya virus and related alphaviruses. We show that CD81 promotes replication of viral genomes in human and mouse cells, while virus entry into cells is independent of CD81. This provides novel insights into how alphaviruses hijack host proteins to complete their life cycle. Alphaviruses replicate at distinct sites of the plasma membrane, which are enriched in cholesterol. We found that the cholesterol-binding ability of CD81 is important for its function as an alphavirus host factor. This discovery thus broadens our understanding of the alphavirus replication process and the use of host factors to reprogram cells into virus replication factories.
•Biaxial fatigue behaviour was characterised in initiation and propagation stages.•Crack growth rate at initiation showed oscillations due to microstructure.•Propagation stage has been studied with ...the help of five critical plane models.•Models’ performance was evaluated in terms of crack orientation and fatigue life.•The overall best predictions were obtained by FS model.
The crack initiation and propagation behaviour of St52-3N steel is investigated under biaxial conditions. Short crack behaviour is studied and pearlite bands are found to modify the crack growth rate. The behaviour under proportional and non-proportional (90°) loading is studied with different critical plane models; Wang–Brown, Fatemi–Socie, Smith–Watson–Topper, Liu I and Liu II. Smith–Watson–Topper and Liu I are found to best predict the crack orientation under proportional loading and Fatemi–Socie and Liu II under non-proportional loading. The best predictions in terms of fatigue life are obtained by Fatemi–Socie, both under proportional and non-proportional loads.
Inhibitors of bromodomain and extra-terminal proteins (iBETs), including JQ-1, have been suggested as potential prophylactics against SARS-CoV-2 infection. However, molecular mechanisms underlying ...JQ-1-mediated antiviral activity and its susceptibility to viral subversion remain incompletely understood. Pretreatment of cells with iBETs inhibited infection by SARS-CoV-2 variants and SARS-CoV, but not MERS-CoV. The antiviral activity manifested itself by reduced reporter expression of recombinant viruses, and reduced viral RNA quantities and infectious titers in the culture supernatant. While we confirmed JQ-1-mediated downregulation of expression of angiotensin-converting enzyme 2 (ACE2) and interferon-stimulated genes (ISGs), multi-omics analysis addressing the chromatin accessibility, transcriptome and proteome uncovered induction of an antiviral nuclear factor erythroid 2-related factor 2 (NRF-2)-mediated cytoprotective response as an additional mechanism through which JQ-1 inhibits SARS-CoV-2 replication. Pharmacological inhibition of NRF-2, and knockdown of NRF-2 and its target genes reduced JQ-1-mediated inhibition of SARS-CoV-2 replication. Serial passaging of SARS-CoV-2 in the presence of JQ-1 resulted in predominance of ORF6-deficient variant, which exhibited resistance to JQ-1 and increased sensitivity to exogenously administered type I interferon (IFN-I), suggesting a minimised need for SARS-CoV-2 ORF6-mediated repression of IFN signalling in the presence of JQ-1. Importantly, JQ-1 exhibited a transient antiviral activity when administered prophylactically in human airway bronchial epithelial cells (hBAECs), which was gradually subverted by SARS-CoV-2, and no antiviral activity when administered therapeutically following an established infection. We propose that JQ-1 exerts pleiotropic effects that collectively induce an antiviral state in the host, which is ultimately nullified by SARS-CoV-2 infection, raising questions about the clinical suitability of the iBETs in the context of COVID-19.
This work presents a novel methodology for characterising fatigue cracks under biaxial conditions on a low carbon steel. It allows both short crack and early propagation stages to be studied in ...tubular specimens. Short crack growth is studied with a long-distance microscope acquiring images of the bare metal surface. Results showed oscillations in crack growth rate due to microstructure. Early propagation stage is studied with high magnification Digital Image Correlation (DIC) technique for measuring displacement and strain crack-tip fields. By applying micro-speckle pattern on the metal surface it is possible to achieve high magnification for DIC technique. Ultra-fine black and white speckles were created by electro-spray technique. The validity of this novel technique is demonstrated by direct comparison with extensometer measurements, under combined tension–compression and torsion conditions. It was also possible to estimate satisfactorily the mixed-mode stress intensity factor.
•The strip yield models implemented in NASGRO software are used.•Constant constraint-loss model and variable constraint-loss model are compared.•A new crack growth rate relation (da/dN–ΔKeff) is ...proposed for Al-2024T351.•Comparisons between three different crack growth rate relations are made.•Good predictions of fatigue lives are obtained under random loadings.
This paper is concerned with the application of the strip yield models implemented in NASGRO to estimate fatigue crack growth under random loading. The two different strip yield model options (constant constraint-loss (CCL) and variable constraint-loss (VCL)) implemented in the software are considered and compared. In addition, three crack grow rate relations, obtained from constant amplitude data, the NASGRO material database and the literature, were also considered and compared. The capacity of the models to estimate the fatigue life and variability is analyzed by comparing simulated results with experimental data of fatigue crack growth under different stationary Gaussian random load processes on compact, C(T), specimens of aluminum alloy 2024-T351. The analysis performed showed that both model options provide very similar good fatigue life predictions. The estimated fatigue lives are within ±13% of the test lives with the best option. The variability of the results due to the randomness of the load is also analyzed. In this case, the variable constraint-loss option provides better estimation than the constant constraint-loss option.
Clinical data of patients suffering from COVID-19 indicates that statin therapy, used to treat hypercholesterolemia, is associated with a better disease outcome. Whether statins directly affect virus ...replication or influence the clinical outcome through modulation of immune responses is unknown. We therefore investigated the effect of statins on SARS-CoV-2 infection in human lung cells and found that only fluvastatin inhibited low and high pathogenic coronaviruses in vitro and ex vivo in a dose-dependent manner. Quantitative proteomics revealed that fluvastatin and other tested statins modulated the cholesterol synthesis pathway without altering innate antiviral immune responses in infected lung epithelial cells. However, fluvastatin treatment specifically downregulated proteins that modulate protein translation and viral replication. Collectively, these results support the notion that statin therapy poses no additional risk to individuals exposed to SARS-CoV-2 and that fluvastatin has a moderate beneficial effect on SARS-CoV-2 infection of human lung cells.
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•Fluvastatin inhibits low (HCoV-229E) and high (SARS-CoV-2) pathogenic coronaviruses•SARS-CoV-2 infection does not affect statin-induced cholesterol biosynthesis arrest•Statin treatment does not alter SARS-CoV-2 innate immune responses in vitro•Fluvastatin downregulates protein translation and viral replication pathways
Drugs; Virology
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