Biology subjects

Rodenhuis-Zybert, I. A.

Publications and source records attributed to Rodenhuis-Zybert, I. A..

7 recordsLinked to original sources

bioRxiv2026

A TLR2-associated entry pathway promotes productive dengue virus serotype 2 infection and inflammatory responses in classical monocytes

Severe dengue is characterized by systemic inflammation and vascular leakage. Recent evidence indicates that Toll-like receptor 2 (TLR2) supports early steps of dengue virus (DENV) infection in monocytes, whereas intracellular viral RNA sensing drives much of the subsequent inflammatory response. However, whether this entry-associated function of TLR2 differs among human monocyte subsets remains unknown. Here, we investigated how the TLR2 axis regulates DENV serotype 2 (DENV2) infection and inflammatory output in purified primary human monocytes and sorted monocyte subsets. Blocking TLR2 at the cell surface or targeting its intracellular TIR domain reduced virus-cell association, productive infection, and the release of inflammatory and antiviral mediators. Among TLR2-associated receptors, CD14 and, to a lesser extent, CD36 contributed to efficient infection, whereas TLR1, TLR6, and TLR10 were dispensable under the conditions tested. Notably, TLR2 blockade reduced viral production and cytokine release in CD16- classical monocytes but not in pooled CD16+ monocytes, identifying a subset-specific dependence on the TLR2 axis. TLR3 contributed to inflammatory and antiviral responses without affecting viral replication, consistent with post-entry recognition of viral RNA. Finally, blocking the TLR2 axis during monocyte infection prevented the loss of endothelial barrier integrity induced by monocyte-derived soluble mediators. Together, these findings identify a TLR2-associated entry route that preferentially supports productive DENV2 infection in classical monocytes and thereby enables RNA-sensor-dependent inflammatory responses that compromise endothelial function.

microbiology↗

bioRxiv2022

TLR2 axis on peripheral blood mononuclear cells regulates inflammatory responses to circulating, non-infectious immature dengue virus particles

Severe dengue virus (DENV) infection is characterized by exacerbated inflammatory responses that lead to endothelial dysfunction and plasma leakage. We have recently demonstrated that Toll-like receptor 2 (TLR2) on blood monocytes senses DENV infection leading to endothelial activation. Here, we report that non-infectious immature DENV particles, which are released in large numbers by DENV-infected cells, drive endothelial activation via the TLR2 axis. We show that fully immature DENV particles induce a rapid, within 6 hours post-infection, inflammatory response in PBMCs. Furthermore, pharmacological blocking of TLR2/TLR6/CD14 and/or NF-kB prior to exposure of PBMCs to immature DENV reduces the initial production of inter alia TNF- and IL-1{beta} by monocytes and prevents endothelial activation. However, prolonged TLR2 block induces TNF- production and leads to exacerbated endothelial activation, indicating that TLR2-mediated responses play an important role not only in the initiation but also the resolution of inflammation. Altogether, these data indicate that the maturation status of the virus has the potential to influence the kinetics and extent of inflammatory responses during DENV infection. Author SummarySevere dengue virus (DENV) infection is characterized by endothelial dysfunction leading to vascular permeability and plasma leakage. This is thought to be mediated by the exacerbated production of inflammatory mediators from cells of the innate immune system. We have previously demonstrated that Toll-like receptor 2 (TLR2), a pattern recognition receptor present on the surface of human monocytes, can sense DENV infection leading to the activation of the endothelium. Importantly, a large proportion of DENV particles is immature and are not readily infectious. Here we aimed to elucidate if and how these non-infectious immature DENV particles contribute to systemic inflammation. We evaluated if monocytes sense immature DENV and found that sensing of immature DENV induced early inflammatory responses in PBMCs. Pharmacological inhibition of TLR2/TLR6/CD14 and/or NF-{kappa}B prior to exposure to immature DENV demonstrates that this is the pathway leading to the early production inflammatory mediators and endothelial activation. However, prolonged inhibition of TLR2 induced the production of TNF- and the subsequent activation of the endothelium, suggesting that TLR2-mediated responses play an important tole during both, initiation and resolution of inflammation. We propose that the maturation status of DENV in the human host can influence the extent and kinetics of the inflammatory responses during DENV infection.

immunology↗

bioRxiv2021

Moxidectin and ivermectin inhibit SARS-CoV-2 replication in Vero E6 cells but not in human primary airway epithelium cells

Antiviral therapies are urgently needed to treat and limit the development of severe COVID-19 disease. Ivermectin, a broad-spectrum anti-parasitic agent, has been shown to have anti-SARS-CoV-2 activity in Vero cells at a concentration of 5 {micro}M. These in vitro results triggered the investigation of ivermectin as a treatment option to alleviate COVID-19 disease. In April 2021, the World Health Organization stated, however, the following: "the current evidence on the use of ivermectin to treat COVID-19 patients is inconclusive". It is speculated that the in vivo concentration of ivermectin is too low to exert a strong antiviral effect. Here, we performed a head-to head comparison of the antiviral activity of ivermectin and a structurally related, but metabolically more stable, moxidectin in multiple in vitro models of SARS-CoV-2 infection, including physiologically relevant human respiratory epithelial cells. Both moxidectin and ivermectin exhibited antiviral activity in Vero E6 cells. Subsequent experiments revealed that the compounds predominantly act on a step after virus cell entry. Surprisingly, however, in human airway-derived cell models, moxidectin and ivermectin failed to inhibit SARS-CoV-2 infection, even at a concentration of 10 {micro}M. These disappointing results calls for a word of caution in the interpretation of anti-SARS-CoV-2 activity of drugs solely based on Vero cells. Altogether, these findings suggest that, even by using a high-dose regimen of ivermectin or switching to another drug in the same class are unlikely to be useful for treatment against SARS-CoV-2 in humans.

microbiology↗

bioRxiv2020

Resveratrol And Pterostilbene Potently Inhibit SARS-CoV-2 Infection In Vitro

The current COVID-19 pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has an enormous impact on human health and economy. In search for therapeutic options, researchers have proposed resveratrol, a food supplement with known antiviral, anti-inflammatory and anti-oxidant properties as an advantageous antiviral therapy for SARS-CoV-2 infection. Here, we provide evidence that both resveratrol and its metabolically more stable structural analog, pterostilbene, exhibit potent antiviral properties against SARS-CoV-2 in vitro. Resveratrol and pterostilbene showed antiviral activity in African green monkey kidney cells and in human primary bronchial epithelial cells cultured in an air-liquid interface system. Both compounds actively inhibit virus replication within infected cells as reduced virus progeny production was observed when the compound was added at post-inoculation conditions. Without replenishment of the compound, antiviral activity was observed up to roughly 5 rounds of replication, demonstrating the long-lasting effect of these compounds. Collectively, our data indicate that resveratrol and pterostilbene are promising antiviral compounds to treat SARS-CoV-2 infection. Because these results represent laboratory findings in cells, we advocate evaluation of these compounds in clinical trials before statements are made whether or not these drugs are advantageous for COVID-19 treatment.

microbiology↗

bioRxiv2020

Rewiring PBMC responses to prevent CHIKV infection-specific monocyte subset redistribution and cytokine responses

Infection with the mosquito-borne Chikungunya virus (CHIKV) causes acute or chronic arthritis in humans. Inflammatory responses mediated by monocytes, the primary target cells of CHIKV infection in the blood, are considered to play an important role in CHIKV pathogenesis. A recent study revealed that the acute phase of CHIKV infection is characterized by a monocyte-driven response, with an expansion of the intermediate monocyte (IM) subset. In this study, we adopted a previously established in vitro model of CHIKV infection in peripheral blood mononuclear cells, to elucidate the mechanism and relevance of IM expansion in CHIKV replication and associated inflammatory responses. Our data show that infectious but not replication-incompetent CHIKV increases the frequency of IM and to a lesser extent, non-classical (NM) monocytes while reducing the number of classical monocytes (CM). The increase of IM or NM frequency coincided with the activation of inflammatory response and occurred in the absence of lymphocytes implying that monocyte-derived cues are sufficient to drive this effect. Importantly, priming of monocytes with LPS prevented expansion of IM and NM but had no effect on viral replication. It did however alter CHIKV-induced cytokine signature. Taken together, our data delineate the role of IM in CHIKV infection-specific innate immune responses and provide insight for the development of therapeutic strategies that may focus on rewiring monocyte immune responses to prevent CHIKV-mediated arthralgia and arthritis.

immunology↗

bioRxiv2020

Chikungunya virus requires an intact microtubule network for efficient viral genome delivery

Chikungunya virus (CHIKV) is a re-emerging mosquito-borne virus, which has rapidly spread around the globe thereby causing millions of infections. CHIKV is an enveloped virus belonging to the Togaviridae family and enters its host cell primarily via clathrin-mediated endocytosis. Upon internalization, the endocytic vesicle containing the virus particle moves through the cell and delivers the virus to early endosomes where membrane fusion is observed. Thereafter, the nucleocapsid dissociates and the viral RNA is translated into proteins. In this study, we examined the importance of the microtubule network during the early steps of infection and dissected the intracellular trafficking behavior of CHIKV particles during cell entry. We observed two distinct CHIKV intracellular trafficking patterns prior to membrane hemifusion. Whereas half of the CHIKV virions remained static during cell entry and fused in the cell periphery, the other half showed fast-directed microtubule-dependent movement prior to delivery to Rab5-positive early endosomes and predominantly fused in the perinuclear region of the cell. Disruption of the microtubule network reduced the number of infected cells. At these conditions, membrane hemifusion activity was not affected yet fusion was restricted to the cell periphery. Furthermore, follow-up experiments revealed that disruption of the microtubule network impairs the delivery of the viral genome to the cell cytosol. We therefore hypothesize that microtubules may direct the particle to a cellular location that is beneficial for establishing infection or aids in nucleocapsid uncoating. Author SummaryChikungunya virus (CHIKV) is an alphavirus that is transmitted to humans by infected mosquitoes. Disease symptoms can include fever, rash, myalgia, and long-lasting debilitating joint pains. Unfortunately, there is currently no licensed vaccine or antiviral treatment available to combat CHIKV. Understanding the virus:host interactions during the replication cycle of the virus is crucial for the development of effective antiviral therapies. In this study we elucidated the trafficking behavior of CHIKV particles early in infection. During cell entry, CHIKV virions require an intact microtubule network for efficient delivery of the viral genome into the host cell thereby increasing the chance to productively infect a cell.

microbiology↗

bioRxiv2020

Serotonergic drugs inhibit CHIKV infection at different stages of the cell entry pathway

Chikungunya virus (CHIKV) is an important re-emerging human pathogen transmitted by mosquitoes. The virus causes an acute febrile illness, chikungunya fever, which is characterized by headache, rash and debilitating (poly)arthralgia that can reside for months to years after infection. Currently, effective antiviral therapies and vaccines are lacking. Due to the high morbidity and economic burden in the countries affected by CHIKV, there is a strong need for new strategies to inhibit CHIKV replication. The serotonergic drug, 5-nonyloxytryptamine (5-NT), was previously identified as a potential host-directed inhibitor for CHIKV infection. In this study, we determined the mechanism of action by which the serotonin receptor agonist 5-NT controls CHIKV infection. Using time-of-addition and entry bypass assays we found that 5-NT predominantly inhibits CHIKV in the early phases of the replication cycle; at a step prior to RNA translation and genome replication. Intriguingly, however, no effect was seen during virus-cell binding, internalization, membrane fusion and gRNA release into the cell cytosol. Additionally, we show that the serotonin receptor antagonist MM also has antiviral properties towards CHIKV and specifically interferes with the cell entry process and/or membrane fusion. Taken together, pharmacological targeting of 5-HT receptors may represent a potent way to limit viral spread and disease severity. ImportanceThe rapid spread of mosquito-borne viral diseases in humans puts a huge economic burden on developing countries. For many of these infections, including Chikungunya virus (CHIKV), there are no specific treatment possibilities to alleviate disease symptoms. Understanding the virus:host interactions that are involved in the viral replication cycle is imperative for the rational design of therapeutic strategies. In this study, we discovered an antiviral compound and elucidated the mechanism of action and propose serotonergic drugs as potential host-directed antivirals for CHIKV.

microbiology↗