The late 2019 emergence of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, caused profound and unprecedented disruption to the global socio-economic structure, negatively affecting millions of lives worldwide. A typical hallmark of severe COVID-19 is hyper inflammation due to aberrant cytokine release (cytokine storm) by innate immune cells. Recent studies have revealed that SARS-CoV-2, through its spike (S) protein, can activate the body’s innate immune cells via Toll-Like Receptors (TLRs), particularly TLR4. In silico studies have demonstrated that the S protein binds with high affinity to TLR4, triggering downstream signaling processes that result in pro-inflammatory cytokine release. Compared to other TLRs, such as TLR2, TLR4 plays a more significant role in initiating and sustaining the inflammatory response associated with severe COVID-19. Furthermore, interactions between the virus and target cells can enhance the cellular expression of TLR4, making cells more susceptible to viral interactions and subsequent inflammation. This increased expression of TLR4 upon viral entry creates a feedback loop, where heightened TLR4 levels lead to amplified inflammatory responses, contributing to the severity of the disease. Additionally, TLR4’s potent activation of inflammatory pathways sets it apart from other TLRs, underscoring its pivotal role in the pathogenesis of COVID-19. In this review, we thoroughly explore the multitude of regulatory signaling pathways that SARS-CoV-2 employs to incite inflammation. We specifically focus on the critical impact of TLR4 activation compared to other TLRs, highlighting how TLR4’s interactions with the viral S protein can exacerbate the severity of COVID-19. By delving into the mechanisms of TLR4-mediated inflammation, we aim to shed light on potential therapeutic targets that could mitigate the inflammatory damage caused by severe COVID-19. Understanding the unique role of TLR4 in the context of SARS-CoV-2 infection could pave the way for novel treatment strategies that specifically inhibit this receptor’s activity, thereby reducing the overall disease burden and improving patient outcomes.

Problem: Although it is still uncertain whether Severe Acute Respiratory Coronavirus (SARS‐CoV‐2) placental infection and vertical transmission occur, inflammation during early pregnancy can have devastating consequences for gestation itself and the growing fetus. If and how SARS‐CoV‐2‐specific immune cells negatively affect placenta functionality is still unknown.Method of study: We stimulated peripheral blood mononuclear cells (PBMCs) from women of reproductive age with SARS‐CoV‐2 peptides and cocultured them with trophoblast spheroids (HTR‐8/SVneo and JEG‐3) to dissect if SARS‐CoV‐2‐activated immune cells can interfere with trophoblast functionality. The activation and cytokine profile of the PBMCs were determined using multicolor flow cytometry. The functionality of trophoblast spheroids was assessed using microscopy, enzyme‐linked immunosorbent assay (ELISA), and RT‐qPCR.Results: SARS‐CoV‐2 S and M peptides significantly activated PBMCs (monocytes, NK cells, and T cells with memory subsets) and induced the upregulation of proinflammatory cytokines, such as IFNγ. The activated PBMCs did not impact the viability, growth rate, and invasion capabilities of trophoblast spheroids. Furthermore, the hormonal production of hCG by JEG‐3 spheroids was not compromised upon coculture with the activated PBMCs. mRNA transcript levels of genes involved in trophoblast spheroid functional pathways were also not dysregulated after coculture.Conclusions: Together, the findings of our in vitro coculture model, although not fully representative of in vivo conditions, strongly support the claim that the interaction of SARS‐CoV‐2‐activated peripheral blood immune cells with trophoblast cells at the fetal–maternal interface does not negatively affect trophoblast functionality. This goes in hand with the recommendation of vaccinating pregnant women in their first trimester.

Per-and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants that are widely detected in human blood samples. Recent epidemiological studies suggest a negative correlation between PFAS blood concentrations and SARS-CoV-2 antibody responses following COVID-19 vaccination. Despite these observations, the direct impact of PFAS mixtures on the interaction between circulating T follicular helper (cTfh) and B cells, which is pivotal for the production of neutralizing antibodies in response to SARS-CoV-2 vaccination, remains unexplored. Using human peripheral blood mononuclear cells (PBMCs) isolated from six healthy female donors who had previously been vaccinated and subsequently boosted or recovered from COVID-19, we examined in vitro the effect of a PFAS mixture on cTfh and B-cell immune responses to SARS-CoV-2 spike peptide. The PBMCs were exposed to a PFAS mixture comprising perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA) at human-relevant blood concentrations (1X), and higher concentrations (10X, 100X, 1000X), followed by stimulation with SARS-CoV-2 spike peptides for five days. We showed that the PFAS mixtures did not induce cytotoxicity but caused immunomodulation. Exposure to the highest PFAS concentration tested (1000X human blood concentrations) led to a selective reduction in cTfh17 cells and upregulation of ICOS and IL-21 in cTfh1/17 cells. B cells exhibited unusual activation, with naïve B cells showing elevated CD80/CD86 expression and switched memory B cells upregulating CD69. Furthermore, despite unperturbed levels of IL-21 and CXCL13, important mediators involved in antibody production, we observed a significant reduction in plasmablasts and SARS-CoV-2 spike-specific IgG secretion at the highest PFAS concentrations. In conclusion, this study provides in vitro evidence that PFAS mixtures can modulate selected components of the cTfh-B-cell axis and may contribute to impaired vaccine-induced antibody responses under high-exposure conditions. These findings add an important piece of information as to why high PFAS-exposed individuals present reduced COVID-19 vaccine antibody titers.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental pollutants that have been associated with adverse pregnancy outcomes and immune dysfunction. However, the impact of PFAS on trophoblast-mediated immune interactions at the maternal-fetal interface remains unclear. In this study, we evaluated the effects of human-relevant PFAS on trophoblast function and immune crosstalk using a three-dimensional (3D) trophoblast spheroid model and a co-culture system with CD4+ T cells. JEG-3 spheroids were exposed to multiple PFAS, including perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA), individually and as a mixture at real-life concentrations to assess viability, invasion and trophoblast-derived β-hCG secretion. Using a placental–immune cell co-culture model, we examined bidirectional interactions and their respective effects on functional endpoints in both cellular compartments and further investigated how exposure to PFAS may interfere in the bidirectional communication. We report a significant decrease in trophoblast viability, after exposure to individual PFOS, PFOA and the PFAS mixture. PFAS mixture, PFOS, PFHxS, PFNA and PFDA exposure significantly increased invasiveness in JEG-3 spheroids, while PFOA decreased JEG-3 invasion significantly. Additionally, β-hCG production declined after 48 hours of PFOS and PFAS mixture exposure in JEG-3 spheroids. Moreover, following PFAS mixture exposure, we observed a downregulation of CD69 and IFNγ production in CD4+ T cells, particularly within Th2 cells, while CD154 expression was also reduced. Overall, these findings suggest that PFAS exposure may influence trophoblast-mediated immune signaling, with potential implications for pregnancy-related processes.