Early Stages Of Viral Infection Research Articles

Effect of COVID-19 infection on the gastrointestinal tract considering preventive methods during endoscopic procedures.

This study aimed to prevent the infection risk of environmental contamination by feces during endoscopic procedures. We evaluated the effect of coronavirus disease 2019 (COVID-19) on the gastrointestinal tract using fecal polymerase chain reaction (PCR) and examined risk factors affecting endoscopic procedures, to develop infection prevention strategies. This single-center prospective observational study enrolled 32 patients diagnosed with COVID-19 at Tokyo Medical University Hospital between January and December 2022. We performed reverse transcriptase-PCR to detect severe acute respiratory syndrome coronavirus 2 in human stool specimens and evaluated the COVID-19 positivity rate in stool, the effect of vaccination on infection detection, and differences in positivity rates considering different patient backgrounds. Among the 32 nasal PCR-positive patients who underwent fecal PCR testing, the fecal PCR positivity rate was 21.8%. Compared to the negative cases, 71.4% vs. 32% were older than 65 years (p < 0.016), 71.4% vs. 0.8% (p < 0.001) had malignant tumors, the rate during BA.5 variant outbreaks was significantly higher (100% vs. 60% [p = 0.044]), and the rate of diarrheal symptoms was also higher (42.9% vs. 24%). The median collection period for fecal PCR-positive cases was 2 days after sampling. The severe acute respiratory syndrome coronavirus 2 affects not only the upper respiratory tract but also the gastrointestinal tract. These findings may indicate the risk of digestive fluid infection in older patients with gastrointestinal symptoms and immunocompromised patients with malignant tumor comorbidities, especially during the early stages of viral infection. Therefore, it is advisable to establish a system to prevent infection by using personal protective equipment, including eye guards, in future endoscopic procedures.

The Expression of P35 Plays a Key Role in the Difference in Apoptosis Induced by AcMNPV Infection in Different Spodoptera exigua Cell Lines.

Baculovirus infection induces apoptosis in host cells, and apoptosis significantly affects virus production. Autographa californica multiple nucleopolyhedrovirus (AcMNPV) can regulate apoptosis, but the regulatory mechanism is unclear. Here, we found that AcMNPV infection induced different apoptosis responses in different Spodoptera exigua cell lines. In the early stages of viral infection (1-6 h), Se-1 cells underwent severe apoptosis, while Se-3 cells underwent very slight apoptosis. In the late stages of viral infection (12-72 h), Se-1 cells continued to undergo apoptosis and formed a large number of apoptotic bodies, while the apoptosis of Se-3 cells was inhibited and no apoptotic bodies were formed. To determine the reasons for the apoptosis differences in the two cell lines, we measured the expression of the six S. exigua cysteine-dependent aspartate specific protease genes (SeCaspase-1 to -6) and the three AcMNPV antiapoptotic protein genes (iap1, iap2 and p35) during viral infection. We found that SeCaspase-1 to -6 were all activated in Se-1 cells and inhibited in Se-3 cells, whereas iap1, iap2 and p35 were all inhibited in Se-1 cells and normally expressed in Se-3 cells. And p35 was expressed earlier than iap1 and iap2 in Se-3 cells. Otherwise, Se-1 and Se-3 cells would all be apoptotic when infected with the recombinant p35 knockout AcMNPV, whereas only Se-1 cells were apoptotic, but Se-3 cells were not apoptotic when infected with the recombinant p35 repair AcMNPV. Combined with the fact that the expression of P35 protein is inhibited in Se-1 cells but normally expressed in Se-3 cells during the infection of recombinant p35 repair AcMNPV, we proposed that the different expression of P35 is an important reason for the apoptosis differences between the two cell lines. We also found that some genes associated with apoptosis can probably regulate the expression of P35. However, the major upstream regulators of P35 and their mechanisms are still unclear and will be studied in the future.

Catalases mediate tobacco resistance to virus infection through crosstalk between salicylic acid and auxin signaling pathways.

Catalases (CATs) play important roles in plant growth, development and defense responses. Previous studies have shown that CATs exhibit different or even opposite effects on plant immunity in different plant-pathogen interactions, but little is known about the mechanisms. In this study, Nicotiana tabacum plants with overexpression or knockout of CAT genes, tobacco mosaic virus (TMV) and cucumber mosaic virus (CMV) were employed to investigate the role of CAT in compatible plant-virus interactions. The results showed that there were dynamic changes in the effect of CAT on N. tabacum defense responses. Overexpression of catalase 1 (CAT1) and catalase 3 (CAT3) improved N. tabacum resistance in the early stage of virus infection but depressed it during the late stages of pathogenesis, especially in CAT3 overexpressing plants. The lower level of electrolyte leakage, lower contents of malonaldehyde and hydrogen peroxide (H2 O2 ), higher activities of antioxidant enzymes and improved functions of photosystem II corresponded to the milder symptoms and higher resistance of infected tobacco plants. In addition, the infection of TMV and CMV resulted in expression changes of CATs in tobacco plants, and pretreatment with H2 O2 facilitated TMV and CMV infection. Further experiments showed that the content of salicylic acid (SA) and the expression of genes related to SA signaling pathway were positively correlated with plant resistance, whereas auxin and its related signaling pathway were related to the viral susceptibility of plants. Taken together, our results demonstrated that CAT1 and CAT3 mediated tobacco resistance to virus infection through crosstalk between SA and auxin signaling pathways.

Liang-Ge-San inhibits dengue virus serotype 2 infection by reducing caveolin1-induced cytoplasmic heat shock protein 70 translocation into the plasma membrane

Dengue virus (DENV) is a major public health threat. However, there are no specific therapeutic drugs for DENV. Many Chinese heat-cleaning formulas, such as Liang-Ge-San (LGS), have been frequently used in the virus-induced diseases. The antiviral effect of LGS has not been reported yet. In this study, the effect of LGS on the inhibition of dengue virus serotype 2 (DENV-2) was investigated and the relevant mechanism was explored. High-performance liquid chromatography was applied to analyze the chemical characterization of LGS. The in vitro antiviral activities of LGS against DENV-2 were evaluated by time-of-drug-addition assay. The binding of heat shock protein 70 (Hsp70) and envelope (E) protein or caveolin1 (Cav1) were analyzed by immunofluorescence and immunoprecipitation assays. Then the role of Cav1 in the anti-DENV-2 effects of LGS was further examined. DENV-2 infected Institute of Cancer Research suckling mice (n=10) and AG129 mice (n=8) were used to examine the protective effects of LGS. It was found that geniposide, liquiritin, forsythenside A, forsythin, baicalin, baicalein, rhein, and emodin maybe the characteristic components of LGS. LGS inhibited the early stage of DENV-2 infection, decreased the expression levels of viral E and non-structural protein 1 (NS1) proteins. LGS also reduced E protein and Hsp70 binding and attenuated the translocation of Hsp70 from cytoplasm to the cell membrane. Moreover, LGS decreased the binding of Hsp70 to Cav1. Further study showed that the overexpression of Cav1 reversed LGS-mediated E protein and Hsp70 inhibition in the plasma membrane. In the in vivo study, LGS was highly effective in prolonging the survival time, reducing viral loads. This work demonstrates for the first time that LGS exerts anti-DENV-2 activity in vitro and in vivo. LGS decreases DENV-2-stimulated cytoplasmic Hsp70 translocation into the plasma membrane by Cav1 inhibition, thereby inhibiting the early stage of virus infection. These findings indicate that LGS may be a candidate for the treatment of DENV.

Identification of serum immunoglobulin M (IgM) of crucian carp (Carassius auratus gibelio) and immune response to cyprinid herpesvirus 2 infection

Crucian carp (Carassius auratus gibelio), an extensively cultivated freshwater fish, was one of the model species for the study of fish immunology. Polyclonal antibodies were advantageous molecular tools for studying teleost immune system. Specifically, polyclonal antibodies reacting with immunoglobulins (Ig) were used successfully in studies of the teleost fishes. In the present study, we produced polyclonal antibody against CH2 domains of crucian carp IgM, and measured the in vivo dynamics of IgM mRNA caused by CyHV-2 infection. The recombinant protein IgM with relative molecular weight about 53 KD was correctly expressed in prokaryotic cells. The specificity of the polyclonal antibody was evaluated by Western blotting and results revealed that the antibody not only specifically recognized crucian carp serum but also cross-reacted with grass carp serum. Quantitative RT-PCR analysis demonstrated the expression of IgM mRNA changed significantly after CyHV-2 infection. The expression of IgM in the kidney increased and reached a maximum at 6 h post-infection (hpi), while dropped to a low level at 5 days post-infection (dpi). In conclusion, the expression of IgM was significantly upregulated in the kidney of crucian carp infected with CyHV-2, indicating that IgM played a potential role in systemic immunity against viral infection. Polyclonal antibody against crucian carp IgM had certain clinical relevance, which might provide insight into the early stage of virus infection and prevention of the disease.

HCPro affects heterologous virus infection through salicylic acid and auxin pathways

AbstractMixed infection by plant viruses is common in nature, but how a key viral protein of one virus affects the infection by heterologous viruses is not yet fully understood. The helper component proteinase (HCPro) is a widely studied RNA silencing suppressor encoded by viruses of the family Potyviridae. Here, we investigated the defence response of Nicotiana tabacum plants overexpressing HCPro of chilli veinal mottle virus (HCPro‐OX) to tobacco mosaic virus and cucumber mosaic virus infection. We monitored the physiological and molecular changes of HCPro‐OX plants in response to virus infection. The results showed that HCPro‐OX plants under virus infection exhibited higher susceptibility at the early stage but stronger tolerance at the later stage compared to wild‐type plants. The tolerance to heterologous virus infection of HCPro‐OX plants corresponded to a lower level of reactive oxygen species accumulation and higher activities of several antioxidant enzymes. Reverse transcription‐quantitative PCR assays showed that the expression of genes related to salicylic acid (SA) pathways was significantly upregulated, but the expression of genes related to the auxin pathways was downregulated at the late stage of virus infection in HCPro‐OX plants compared to wild‐type plants. By contrast, the situation in the early stage of virus infection was reversed. In addition, pretreatment with SA, the auxin naphthylacetic acid (NAA) and their respective inhibitors 1‐aminobenzotriazole (ABT) and naphthalam (NPA) further confirmed the antagonistic effects of SA and NAA in the response of HCPro‐OX plants to heterologous virus infection. Thus, our results demonstrate that HCPro affects heterologous virus infection through SA and auxin pathways.

PcTrim prevents early infection with white spot syndrome virus by inhibiting AP1-induced endocytosis

Viruses have evolved various strategies to achieve early infection by initiating transcription of their own early genes via host transcription factors, such as NF-κb, STAT, and AP1. How the host copes with this immune escape has been a topic of interest. Tripartite motif (TRIM) family proteins with RING-type domains have E3 ubiquitin ligase activity and are known as host restriction factors. Trim has been reported to be associated with phagocytosis and is also believed to be involved in the activation of autophagy. Preventing the virus from entering the host cell may be the most economical way for the host to resist virus infection. The role of TRIM in the early stage of virus infection in host cells remains to be further interpreted. In the current study, a crayfish TRIM with a RING-type domain, designated as PcTrim, was significantly upregulated under white spot syndrome virus (WSSV) infection in the red swamp crayfish (Procambarus clarkii). Recombinant PcTrim significantly inhibited WSSV replication in crayfish. RNAi targeting PcTrim or blocking PcTrim with an antibody promoted WSSV replication in crayfish. Pulldown and co-IP assays showed that PcTrim can interact with the virus protein VP26. PcTrim restricts the expression level of dynamin, which is involved in the regulation of phagocytosis, by inhibiting AP1 entry into the nucleus. AP1-RNAi effectively reduced the expression levels of dynamin and inhibited host cell endocytosis of WSSV in vivo. Our study demonstrated that PcTrim might reduce early WSSV infection by binding to VP26 and then inhibiting AP1 activation, resulting in reduced endocytosis of WSSV in crayfish hemocytes.BjibKXcktbHxsdfgurhU3PVideo

Anisomycin inhibits Coxsackievirus B replication by promoting the lysosomal degradation of eEF1A1

Group B Coxsackieviruses (CVB) are non-enveloped small RNA viruses in the genus Enterovirus, family Picornaviridae. CVB infection causes diverse conditions from common cold to myocarditis, encephalitis, and pancreatitis. No specific antiviral is available for the treatment of CVB infection. Anisomycin, a pyrrolidine-containing antibiotic and translation inhibitor, was reported to inhibit the replication of some picornaviruses. However, it is unknown if anisomycin can act as an antiviral against CVB infection. Here we observed that anisomycin showed potent inhibition on CVB type 3 (CVB3) infection with negligible cytotoxicity when applied at the early stage of virus infection. Mice infected with CVB3 showed markedly alleviated myocarditis with reduced viral replication. We found that CVB3 infection significantly increased the transcription of eukaryotic translation elongation factor 1 alpha 1 (eEF1A1). CVB3 replication was suppressed by EEF1A1 knockdown, while elevated by EEF1A1 overexpression. Similar to the effect of CVB3 infection, EEF1A1 transcription was increased in response to anisomycin treatment. However, eEF1A1 protein level was decreased with anisomycin treatment in a dose-dependent manner in CVB3-infected cells. Moreover, anisomycin promoted eEF1A1 degradation, which was inhibited by the treatment of chloroquine but not MG132. We demonstrated that eEF1A1 interacted with the heat shock cognate protein 70 (HSP70), and eEF1A1 degradation was inhibited by LAMP2A knockdown, implicating that eEF1A1 is degraded through chaperone-mediated autophagy. Taken together, we demonstrated that anisomycin, which inhibits CVB replication through promoting the lysosomal degradation of eEF1A1, could be a potential antiviral candidate for the treatment of CVB infection.

Dextran sulfate from Leuconostoc mesenteroides B512F exerts potent antiviral activity against SARS-CoV-2 in vitro and in vivo.

The emergent human coronavirus SARS-CoV-2 and its resistance to current drugs makes the need for new potent treatments for COVID-19 patients strongly necessary. Dextran sulfate (DS) polysaccharides have long demonstrated antiviral activity against different enveloped viruses in vitro. However, their poor bioavailability has led to their abandonment as antiviral candidates. Here, we report for the first time the broad-spectrum antiviral activity of a DS-based extrapolymeric substance produced by the lactic acid bacterium Leuconostoc mesenteroides B512F. Time of addition assays with SARS-CoV-2 pseudoviruses in in vitro models confirm the inhibitory activity of DSs in the early stages of viral infection (viral entry). In addition, this exopolysaccharide substance also reports broad-spectrum antiviral activity against several enveloped viruses such as SARS-CoV-2, HCoV229E, HSV-1, in in vitro models and in human lung tissue. The toxicity and antiviral capacity of DS from L. mesenteroides was tested in vivo in mouse models which are susceptible to SARS-CoV-2 infection. The described DS, administered by inhalation, a new route of administration for these types of polymers, shows strong inhibition of SARS-CoV-2 infection in vivo, significantly reducing animal mortality and morbidity at non-toxic doses. Therefore, we suggest that it may be considered as a potential candidate for antiviral therapy against SARS-CoV-2.

Modulation of virus-induced neuroinflammation by the autophagy receptor SHISA9 in mice.

Microglia and astrocytes are subgroups of brain glia cells that support and protect neurons within the central nervous system (CNS). At early stages of viral infection in the CNS, they are predominant responding cells and lead to recruitment of peripheral immune cells for viral clearance. Inhibitor of nuclear factor κB kinase subunit epsilon (IKKi) is critical for type I interferon signalling and inflammation, which modulate heterogenic immune responses during CNS infection. Balanced autophagy is vital to maintain brain integrity, yet regulation of autophagy and immune activity within brain glia cells is poorly understood. Here we identify SHISA9 as an autophagy cargo receptor that mediates the autophagy-dependent degradation of IKKi during herpes simplex virus type 1 infection. IKKi is recognized by SHISA9 through unanchored K48-linked poly-ubiquitin chains and bridged to autophagosome membrane components GABARAPL1. Single-cell RNA sequencing analysis shows that SHISA9 has temporal characteristics while modulating both antiviral and inflammatory responses in microglia and astrocytes at different stages during viral infection. We found that Shisa9-/- mice are highly susceptible to herpes simplex virus encephalitis, have pathogenic astrocytes and display more severe neuroinflammation compared with wild-type mice. Taken together, our study unravels a critical role of selective autophagy by orchestrating immune heterogeneity of different CNS resident cells through the SHISA9-IKKi axis.

Antiviral Effect of Ginsenosides rk1 against Influenza a Virus Infection by Targeting the Hemagglutinin 1-Mediated Virus Attachment

Influenza A virus (IAV) infections have been a serious hazard to public health everywhere. With the growing concern of drug-resistant IAV strains, there is an urgent need for novel anti-IAV medications, especially those with alternative mechanisms of action. Hemagglutinin (HA), an IAV glycoprotein, plays critical roles in the early stage of virus infection, including receptor binding and membrane fusion, making it a good target for developing anti-IAV drugs. Panax ginseng is a widely used herb in traditional medicine with extensive biological effects in various disease models, and its extract was reported to show protection in IAV-infected mice. However, the main effective anti-IAV constituents in panax ginseng remain unclear. Here, we report that ginsenoside rk1 (G-rk1) and G-rg5, out of the 23 screened ginsenosides, exhibit significant antiviral effects against 3 different IAV subtypes (H1N1, H5N1, and H3N2) in vitro. Mechanistically, G-rk1 blocked IAV binding to sialic acid in a hemagglutination inhibition (HAI) assay and an indirect ELISA assay; more importantly, we showed that G-rk1 interacted with HA1 in a dose-dependent manner in a surface plasmon resonance (SPR) analysis. Furthermore, G-rk1 treatment by intranasal inoculation effectively reduced the weight loss and mortality of mice challenged with a lethal dose of influenza virus A/Puerto Rico/8/34 (PR8). In conclusion, our findings reveal for the first time that G-rk1 possesses potent anti-IAV effects in vitro and in vivo. We have also identified and characterized with a direct binding assay a novel ginseng-derived IAV HA1 inhibitor for the first time, which could present potential approaches to prevent and treat IAV infections.

On the Regulation of NF-κB Pathway by HPV Oncoproteins: Are Pathway Inhibitors a Good Alternative for the Treatment of Cervical Cancer?

Cervical cancer (CC) is one of the most prevalent cancer-related pathologies in the female population. It is considered the second leading cause of cancer-related deaths in developing countries. The most important etiological factor for the development of CC is the persistent infection with high-risk human papillomavirus. HPV-oncoproteins have evolved to modulate cellular mechanisms to permit viral replication and the generation of new infectious viral particles. When the viral infection persists, there is an uncontrolled viral protein expression essential to commence and maintain the transformation of infected cells. Different cell pathways are affected during the transformation stage, including the NF-κB signaling pathway. NF-κB controls different cellular mechanisms, and its role is critical for various processes, such as immunity, inflammation, cell differentiation, growth, and survival. NF-κB plays a double role in the development of CC. Evidence suggests that in the early stages of viral infection, the NF-κB activity impairs viral transcription and is beneficial for avoiding cellular immortalization. However, in the advanced stages of cervical carcinogenesis, the activation of the NF-κB correlates with a poor prognosis. Here, we discuss some aspects of NF-κB activity during the development of CC and the use of NF-κB inhibitors to treat this pathology.

Transcriptomes of Zebrafish in Early Stages of Multiple Viral Invasions Reveal the Role of Sterols in Innate Immune Switch-On

Although it is widely accepted that in the early stages of virus infection, fish pattern recognition receptors are the first to identify viruses and initiate innate immune responses, this process has never been thoroughly investigated. In this study, we infected larval zebrafish with four different viruses and analyzed whole-fish expression profiles from five groups of fish, including controls, at 10 h after infection. At this early stage of virus infection, 60.28% of the differentially expressed genes displayed the same expression pattern across all viruses, with the majority of immune-related genes downregulated and genes associated with protein synthesis and sterol synthesis upregulated. Furthermore, these protein synthesis- and sterol synthesis-related genes were strongly positively correlated in the expression pattern of the rare key upregulated immune genes, IRF3 and IRF7, which were not positively correlated with any known pattern recognition receptor gene. We hypothesize that viral infection triggered a large amount of protein synthesis that stressed the endoplasmic reticulum and the organism responded to this stress by suppressing the body's immune system while also mediating an increase in steroids. The increase in sterols then participates the activation of IRF3 and IRF7 and triggers the fish's innate immunological response to the virus infection.

COVID-19 and Support Products: The Effects of Nutritional Immunity

There is an ongoing interaction between nutrition, immune system, and infectious diseases. Inadequate and unbalanced nutrition may reduce the defense efficiency of the immune system and increase the risk of contracting infectious diseases and the severity of disease symptoms. COVID-19 (coronavirus disease-2019) outbreak, which emerged in December 2019 and was declared a pandemic by the World Health Organization (WHO) in a short time, has become the most important public health problem worldwide in the last few years. COVID-19 is asymptomatic or causes with mild symptoms in most people, however, it progresses to severe illness accompanied by serious symptoms in some people, and it is known that this difference is mainly related to the effective immune responses of individuals. There is currently any easily accessible antiviral drug with proven therapeutic efficacy for the prevention and treatment of COVID-19 disease. Therefore, there has been an increasing effort to explore immune-supportive additional therapeutic options to strengthen the immune system and reduce disease-related complaints. The innate immune response, which is responsible for preventing transmission and invasion in the early stages of viral infections, is the first line of defense to eliminate the virus, and it is critical in the effective and balanced activation of the adaptive immune response. Therefore, the use of antimicrobial and antioxidant minerals, vitamins, medicinal plants, and probiotics that will strengthen the immune response is one of the accessible, easily applicable, and effective preventive measures. In this review, the effectiveness of therapeutic and supportive nutritional supplements against COVID-19 disease, which has become a major problem worldwide, has been discussed.

Scaffold identification and drug repurposing for finding potential Dengue envelope inhibitors through ligand-based pharmacophore model

Most of the existing DENV entry inhibitors were discovered through structure-based, high-throughput screening techniques and optimization approaches by aiming β-OG pocket. However, the class of precise chemical scaffolds with superior antiviral activity targeting the early stages of virus infection that is considered to be beneficial in therapeutics and is still in process. In this study, ligand-based pharmacophore modeling using existing DENV entry inhibitors provided two best models, AADRR-2 and AAADR-2 (A- accepter, D- donor, R-ring) to screen public and DrugBank datasets. Further, approximately 36000 molecules were filtered using Zinc13 by employing the ideal validated models. Additionally, using β-OG binding pocket as target site, molecular docking experiments including induced-fit studies were conducted that provided further structurally divergent ligands. Moreover, the refined list of preferential hits were filtered out based on the best fitness score, binding energy and interaction paradigm, among them fused pyrimidine, hydrazone and biphenyl core comprising scaffolds were identified possessing profound interaction profile with key amino acid residues, ALA-50, GLN-200, PHE-193 and PHE-279 in 100 ns MD simulations. Additionally, the search for similar chemical fingerprints from DrugBank library was also carried out and Eltrombopag (Promacta/Revolade® prescribed in thrombocytopenia) was identified as a preferential β-OG pocket binder. The identified pyrazole-based hydrazone class of drug, Eltrombopag is in phase II clinical trials employed to treat dengue-mediated thrombocytopenia. Communicated by Ramaswamy H. Sarma

Metabolism of NK cells during viral infections.

Cellular metabolism is essential for the correct function of immune system cells, including Natural Killer cells (NK). These cells depend on energy to carry out their effector functions, especially in the early stages of viral infection. NK cells participate in the innate immune response against viruses and tumors. Their main functions are cytotoxicity and cytokine production. Metabolic changes can impact intracellular signals, molecule production, secretion, and cell activation which is essential as the first line of immune defense. Metabolic variations in different immune cells in response to a tumor or pathogen infection have been described; however, little is known about NK cell metabolism in the context of viral infection. This review summarizes the activation-specific metabolic changes in NK cells, the immunometabolism of NK cells during early, late, and chronic antiviral responses, and the metabolic alterations in NK cells in SARS-CoV2 infection. The modulation points of these metabolic routes are also discussed to explore potential new immunotherapies against viral infections.

Fluorogenic reporter enables identification of compounds that inhibit SARS-CoV-2.

The coronavirus SARS-CoV-2 causes the severe disease COVID-19. SARS-CoV-2 infection is initiated by interaction of the viral spike protein and host receptor angiotensin-converting enzyme 2 (ACE2). We report an improved bright and reversible fluorogenic reporter, named SURF (split UnaG-based reversible and fluorogenic protein-protein interaction reporter), that we apply to monitor real-time interactions between spike and ACE2 in living cells. SURF has a large dynamic range with a dark-to-bright fluorescence signal that requires no exogenous cofactors. Utilizing this reporter, we carried out a high-throughput screening of small-molecule libraries. We identified three natural compounds that block replication of SARS-CoV-2 in both Vero cells and human primary nasal and bronchial epithelial cells. Cell biological and biochemical experiments validated all three compounds and showed that they block the early stages of viral infection. Two of the inhibitors, bruceine A and gamabufotalin, were also found to block replication of the Delta and Omicron variants of SARS-CoV-2. Both bruceine A and gamabufotalin exhibited potent antiviral activity in K18-hACE2 and wild-type C57BL6/J mice, as evidenced by reduced viral titres in the lung and brain, and protection from alveolar and peribronchial inflammation in the lung, thereby limiting disease progression. We propose that our fluorescent assay can be applied to identify antiviral compounds with potential as therapeutic treatment for COVID-19 and other respiratory diseases.

Approaches to Evaluating Necroptosis in Virus-Infected Cells.

The immune system functions to protect the host from pathogens. To counter host defense mechanisms, pathogens have developed unique strategies to evade detection or restrict host immune responses. Programmed cell death is a major contributor to the multiple host responses that help to eliminate infected cells for obligate intracellular pathogens like viruses. Initiation of programmed cell death pathways during the early stages of viral infections is critical for organismal survival as it restricts the virus from replicating and serves to drive antiviral inflammation immune recruitment through the release of damage-associated molecular patterns (DAMPs) from the dying cell. Necroptosis has been implicated as a critical programmed cell death pathway in a diverse set of diseases and pathological conditions including acute viral infections. This cell death pathway occurs when certain host sensors are triggered leading to the downstream induction of mixed-lineage kinase domain-like protein (MLKL). MLKL induction leads to cytoplasmic membrane disruption and subsequent cellular destruction with the release of DAMPs. As the role of this cell death pathway in human disease becomes apparent, methods identifying necroptosis patterns and outcomes will need to be further developed. Here, we discuss advances in our understanding of how viruses counteract necroptosis, methods to quantify the pathway, its effects on viral pathogenesis, and its impact on cellular signaling.

Targeting SARS-CoV-2 by synthetic dual-acting thiol compounds that inhibit Spike/ACE2 interaction and viral protein production.

The SARS-CoV-2 life cycle is strictly dependent on the environmental redox state that influences both virus entry and replication. A reducing environment impairs the binding of the spike protein (S) to the angiotensin-converting enzyme 2 receptor (ACE2), while a highly oxidizing environment is thought to favor S interaction with ACE2. Moreover, SARS-CoV-2 interferes with redox homeostasis in infected cells to promote the oxidative folding of its own proteins. Here we demonstrate that synthetic low molecular weight (LMW) monothiol and dithiol compounds induce a redox switch in the S protein receptor binding domain (RBD) toward a more reduced state. Reactive cysteine residue profiling revealed that all the disulfides present in RBD are targets of the thiol compounds. The reduction of disulfides in RBD decreases the binding to ACE2 in a cell-free system as demonstrated by enzyme-linked immunosorbent and surface plasmon resonance (SPR) assays. Moreover, LMW thiols interfere with protein oxidative folding and the production of newly synthesized polypeptides in HEK293 cells expressing the S1 and RBD domain, respectively. Based on these results, we hypothesize that these thiol compounds impair both the binding of S protein to its cellular receptor during the early stage of viral infection, as well as viral protein folding/maturation and thus the formation of new viral mature particles. Indeed, all the tested molecules, although at different concentrations, efficiently inhibit both SARS-CoV-2 entry and replication in Vero E6 cells. LMW thiols may represent innovative anti-SARS-CoV-2 therapeutics acting directly on viral targets and indirectly by inhibiting cellular functions mandatory for viral replication.

Analysis of interactions between human immune system and a pathogenic virus

"In this paper we present a mathematical model for studying the interactions between human immune system and a pathogenic virus, such as Covid-19. A mathematical analysis based on dynamical systems theory is performed. More exactly, we model the interactions between the immune system and the virus by a modified predator-prey method. Several conclusions emerge from this study, and the main two of them are the followings: 1) a deficiency in the concentration of a single type of white blood cells in the early stages of virus proliferation may lead to the virus victory, and 2) if the number of at least one type of white blood cells can be increased beyond the normal threshold by medical interventions in the early stages of virus infection, then the immune system has a better chance to win against the virus."