Zika virus infection suppresses CYP24A1 and CAMP expression in human monocytes.

Monocytes are the primary targets of Zika virus (ZIKV) and are associated with ZIKV pathogenesis. Currently, there is no effective treatment for ZIKV infection. It is known that 1,25-dihydroxy vitamin D3 (VitD3) has strong antiviral activity in dengue virus-infected macrophages, but it is unknown whether VitD3 inhibits ZIKV infection in monocytes. We investigated the relationship between ZIKV infection and the expression of genes of the VitD3 pathway, as well as the inflammatory response of infected monocytes in vitro. ZIKV replication was evaluated using a plaque assay, and VitD3 pathway gene expression was analyzed by RT-qPCR. Pro-inflammatory cytokines/chemokines were quantified using ELISA. We found that VitD3 did not suppress ZIKV replication. The results showed a significant decrease in the expression of vitamin D3 receptor (VDR), cytochrome P450 family 24 subfamily A member 1 (CYP24A1), and cathelicidin antimicrobial peptide (CAMP) genes upon ZIKV infection. Treatment with VitD3 was unable to down-modulate production of pro-inflammatory cytokines, except TNF-α, and chemokines. This suggests that ZIKV infection inhibits the expression of VitD3 pathway genes, thereby preventing VitD3-dependent inhibition of viral replication and the inflammatory response. This is the first study to examine the effects of VitD3 in the context of ZIKV infection, and it has important implications for the role of VitD3 in the control of viral replication and inflammatory responses during monocyte infection.

Modeling HIV-1 Infection in CNS via Infected Monocytes Using Immunocompetent Brain Organoids.

The development of 3D-organoid models has revolutionized the way diseases are studied. Recently, our brain organoid model has been shown to recapitulate in in vitro the human brain cytoarchitecture originally encountered in HIV-1 neuropathogenesis, allowing downstream applications. Infected monocytes, macrophages, and microglia are critically important immune cells for infection and dissemination of HIV-1 throughout brain during acute and chronic phase of the disease. Once in the brain parenchyma, long-lived infected monocytes/macrophages along with resident microglia contribute to the establishment of CNS latency in people with HIV (PWH). Hence, it is important to better understand how HIV-1 enters and establishes infection and latency in CNS to further develop cure strategies. Here we detailed an accessible protocol to incorporate monocytes (infected and/or labeled) as a model of transmigration of peripheral monocytes into brain organoids that can be applied to characterize HIV-1 neuroinvasion and virus dissemination.

Cytokine Response of Natural Killer Cells to Hepatitis B Virus Infection Depends on Monocyte Co-Stimulation.

Hepatitis B virus (HBV) is a major driver of chronic hepatic inflammation, which regularly leads to liver cirrhosis or hepatocellular carcinoma. Immediate innate immune cell response is crucial for the rapid clearance of the infection. Here, natural killer (NK) cells play a pivotal role in direct cytotoxicity and the secretion of antiviral cytokines as well as regulatory function. The aim of this study was to further elucidate NK cell responses triggered by an HBV infection. Therefore, we optimized HBV in vitro models that reliably stimulate NK cells using hepatocyte-like HepG2 cells expressing the Na+-taurocholate co-transporting polypeptide (NTCP) and HepaRG cells. Immune cells were acquired from healthy platelet donors. Initially, HepG2-NTCP cells demonstrated higher viral replication compared to HepaRG cells. Co-cultures with immune cells revealed increased production of interferon-γ and tumor necrosis factor-α by NK cells, which was no longer evident in isolated NK cells. Likewise, the depletion of monocytes and spatial separation from target cells led to the absence of the antiviral cytokine production of NK cells. Eventually, the combined co-culture of isolated NK cells and monocytes led to a sufficient cytokine response of NK cells, which was also apparent when communication between the two immune cell subpopulations was restricted to soluble factors. In summary, our study demonstrates antiviral cytokine production by NK cells in response to HBV+ HepG2-NTCP cells, which is dependent on monocyte bystander activation.

Enterovirus A71 crosses a human blood-brain barrier model through infected immune cells.

Enterovirus A71 (EV-A71) is associated with neurological conditions such as acute meningitis and encephalitis. The virus is detected in the bloodstream, and high blood viral loads are associated with central nervous system (CNS) manifestations. We used an in vitro blood-brain barrier (BBB) model made up of human brain-like endothelial cells (hBLECs) and brain pericytes grown in transwell systems to investigate whether three genetically distinct EV-A71 strains (subgenogroups C1, C1-like, and C4) can cross the human BBB. EV-A71 poorly replicated in hBLECs, which released moderate amounts of infectious viruses from their luminal side and trace amounts of infectious viruses from their basolateral side. The barrier properties of hBLECs were not impaired by EV-A71 infection. We investigated the passage through hBLECs of EV-A71-infected white blood cells. EV-A71 strains efficiently replicated in immune cells, including monocytes, neutrophils, and NK/T cells. Attachment to hBLECs of immune cells infected with the C1-like virus was higher than attachment of cells infected with C1-06. EV-A71 infection did not impair the transmigration of immune cells through hBLECs. Overall, EV-A71 targets different white blood cell populations that have the potential to be used as a Trojan horse to cross hBLECs more efficiently than cell-free EV-A71 particles.IMPORTANCEEnterovirus A71 (EV-A71) was first reported in the USA, and numerous outbreaks have since occurred in Asia and Europe. EV-A71 re-emerged as a new multirecombinant strain in 2015 in Europe and is now widespread. The virus causes hand-foot-and-mouth disease in young children and is involved in nervous system infections. How the virus spreads to the nervous system is unclear. We investigated whether white blood cells could be infected by EV-A71 and transmit it across human endothelial cells mimicking the blood-brain barrier protecting the brain from adverse effects. We found that endothelial cells provide a strong roadblock to prevent the passage of free virus particles but allow the migration of infected immune cells, including monocytes, neutrophils, and NK/T cells. Our data are consistent with the potential role of immune cells in the pathogenesis of EV-A71 infections by spreading the virus in the blood and across the human blood-brain barrier.

C-C motif chemokine receptor 2 and 7 synergistically control inflammatory monocyte recruitment but the infecting virus dictates monocyte function in the brain.

Inflammatory monocytes (iMO) are recruited from the bone marrow to the brain during viral encephalitis. C-C motif chemokine receptor (CCR) 2 deficiency substantially reduces iMO recruitment for most, but not all encephalitic viruses. Here we show CCR7 acts synergistically with CCR2 to control this process. Following Herpes simplex virus type-1 (HSV-1), or La Crosse virus (LACV) infection, we find iMO proportions are reduced by approximately half in either Ccr2 or Ccr7 knockout mice compared to control mice. However, Ccr2/Ccr7 double knockouts eliminate iMO recruitment following infection with either virus, indicating these receptors together control iMO recruitment. We also find that LACV induces a more robust iMO recruitment than HSV-1. However, unlike iMOs in HSV-1 infection, LACV-recruited iMOs do not influence neurological disease development. LACV-induced iMOs have higher expression of proinflammatory and proapoptotic but reduced mitotic, phagocytic and phagolysosomal transcripts compared to HSV-1-induced iMOs. Thus, virus-specific activation of iMOs affects their recruitment, activation, and function.

Monocyte to macrophage differentiation and changes in cellular redox homeostasis promote cell type-specific HIV latency reactivation.

HIV latency regulation in monocytes and macrophages can vary according to signals directing differentiation, polarization, and function. To investigate these processes, we generated an HIV latency model in THP-1 monocytes and showed differential levels of HIV reactivation among clonal populations. Monocyte-to-macrophage differentiation of HIV-infected primary human CD14+ and THP-1 cells induced HIV reactivation and showed that virus production increased concomitant with macrophage differentiation. We applied the HIV-infected THP-1 monocyte-to-macrophage (MLat) model to assess the biological mechanisms regulating HIV latency dynamics during monocyte-to-macrophage differentiation. We pinpointed protein kinase C signaling pathway activation and Cyclin T1 upregulation as inherent differentiation mechanisms that regulate HIV latency reactivation. Macrophage polarization regulated latency, revealing proinflammatory M1 macrophages suppressed HIV reactivation while anti-inflammatory M2 macrophages promoted HIV reactivation. Because macrophages rely on reactive-oxygen species (ROS) to exert numerous cellular functions, we disrupted redox pathways and found that inhibitors of the thioredoxin (Trx) system acted as latency-promoting agents in T-cells and monocytes, but opposingly acted as latency-reversing agents in macrophages. We explored this mechanism with Auranofin, a clinical candidate for reducing HIV reservoirs, and demonstrated Trx reductase inhibition led to ROS induced NF-κB activity, which promoted HIV reactivation in macrophages, but not in T-cells and monocytes. Collectively, cell type-specific differences in HIV latency regulation could pose a barrier to HIV eradication strategies.

Journey of monocytes and macrophages upon influenza A virus infection.

Influenza A virus (IAV) infections pose a global health challenge that necessitates a comprehensive understanding of the host immune response to devise effective therapeutic interventions. As monocytes and macrophages play crucial roles in host defence, inflammation, and repair, this review explores the intricate journey of these cells during and after IAV infection. First, we highlight the dynamics and functions of lung-resident macrophage populations post-IAV. Second, we review the current knowledge of recruited monocytes and monocyte-derived cells, emphasising their roles in viral clearance, inflammation, immunomodulation, and tissue repair. Third, we shed light on the consequences of IAV-induced macrophage alterations on long-term lung immunity. We conclude by underscoring current knowledge gaps and exciting prospects for future research in unravelling the complexities of macrophage responses to respiratory viral infections.

Virion-incorporated CD14 enables HIV-1 to bind LPS and initiate TLR4 signaling in immune cells.

HIV-1 has a broad range of nuanced interactions with the immune system, and the incorporation of cellular proteins by nascent virions continues to redefine our understanding of the virus-host relationship. Proteins located at the sites of viral egress can be selectively incorporated into the HIV-1 envelope, imparting new functions and phenotypes onto virions, and impacting viral spread and disease. Using virion capture assays and western blot, we show that HIV-1 can incorporate the myeloid antigen CD14 into its viral envelope. Virion-incorporated CD14 remained biologically active and able to bind its natural ligand, bacterial lipopolysaccharide (LPS), as demonstrated by flow virometry and immunoprecipitation assays. Using a Toll-like receptor 4 (TLR4) reporter cell line, we also demonstrated that virions with bound LPS can trigger TLR4 signaling to activate transcription factors that regulate inflammatory gene expression. Complementary assays with THP-1 monocytes demonstrated enhanced secretion of inflammatory cytokines like tumor necrosis factor alpha (TNF-α) and the C-C chemokine ligand 5 (CCL5), when exposed to LPS-loaded virus. These data highlight a new type of interplay between HIV-1 and the myeloid cell compartment, a previously well-established cellular contributor to HIV-1 pathogenesis and inflammation. Persistent gut inflammation is a hallmark of chronic HIV-1 infection, and contributing to this effect is the translocation of microbes across the gut epithelium. Our data herein provide proof of principle that virion-incorporated CD14 could be a novel mechanism through which HIV-1 can drive chronic inflammation, facilitated by HIV-1 particles binding bacterial LPS and initiating inflammatory signaling in TLR4-expressing cells.IMPORTANCEHIV-1 establishes a lifelong infection accompanied by numerous immunological changes. Inflammation of the gut epithelia, exacerbated by the loss of mucosal T cells and cytokine dysregulation, persists during HIV-1 infection. Feeding back into this loop of inflammation is the translocation of intestinal microbes across the gut epithelia, resulting in the systemic dissemination of bacterial antigens, like lipopolysaccharide (LPS). Our group previously demonstrated that the LPS receptor, CD14, can be readily incorporated by HIV-1 particles, supporting previous clinical observations of viruses derived from patient plasma. We now show that CD14 can be incorporated by several primary HIV-1 isolates and that this virion-incorporated CD14 can remain functional, enabling HIV-1 to bind to LPS. This subsequently allowed CD14+ virions to transfer LPS to monocytic cells, eliciting pro-inflammatory signaling and cytokine secretion. We posit here that virion-incorporated CD14 is a potential contributor to the dysregulated immune responses present in the setting of HIV-1 infection.

SARS-CoV-2 superinfection in CD14+ monocytes with latent human cytomegalovirus (HCMV) promotes inflammatory cascade.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiologic agent of coronavirus disease 2019 (COVID-19), has posed significant challenges to global health. While much attention has been directed towards understanding the primary mechanisms of SARS-CoV-2 infection, emerging evidence suggests co-infections or superinfections with other viruses may contribute to increased morbidity and mortality, particularly in severe cases of COVID-19. Among viruses that have been reported in patients with SARS-CoV-2, seropositivity for Human cytomegalovirus (HCMV) is associated with increased COVID-19 risk and hospitalization. HCMV is a ubiquitous beta-herpesvirus with a seroprevalence of 60-90 % worldwide and one of the leading causes of mortality in immunocompromised individuals. The primary sites of latency for HCMV include CD14+ monocytes and CD34+ hematopoietic cells. In this study, we sought to investigate SARS-CoV-2 infection of CD14+ monocytes latently infected with HCMV. We demonstrate that CD14+ cells are susceptible and permissive to SARS-CoV-2 infection and detect subgenomic transcripts indicative of replication. To further investigate the molecular changes triggered by SARS-CoV-2 infection in HCMV-latent CD14+ monocytes, we conducted RNA sequencing coupled with bioinformatic differential gene analysis. The results revealed significant differences in cytokine-cytokine receptor interactions and inflammatory pathways in cells superinfected with replication-competent SARS-CoV-2 compared to the heat-inactivated and mock controls. Notably, there was a significant upregulation in transcripts associated with pro-inflammatory response factors and a decrease in anti-inflammatory factors. Taken together, these findings provide a basis for the heightened inflammatory response, offering potential avenues for targeted therapeutic interventions among HCMV-infected severe cases of COVID-19. SUMMARY: COVID-19 patients infected with secondary viruses have been associated with a higher prevalence of severe symptoms. Individuals seropositive for human cytomegalovirus (HCMV) infection are at an increased risk for severe COVID-19 disease and hospitalization. HCMV reactivation has been reported in severe COVID-19 cases with respiratory failure and could be the result of co-infection with SARS-CoV-2 and HCMV. In a cell culture model of superinfection, HCMV has previously been shown to increase infection of SARS-CoV-2 of epithelial cells by upregulating the human angiotensin-converting enzyme-2 (ACE2) receptor. In this study, we utilize CD14+ monocytes, a major cell type that harbors latent HCMV, to investigate co-infection of SARS-CoV-2 and HCMV. This study is a first step toward understanding the mechanism that may facilitate increased COVID-19 disease severity in patients infected with SARS-CoV-2 and HCMV.

Human cytomegalovirus modulates mTORC1 to redirect mRNA translation within quiescently infected monocytes.

Human cytomegalovirus (HCMV) utilizes peripheral blood monocytes as a means to systemically disseminate throughout the host. Following viral entry, HCMV stimulates non-canonical Akt signaling leading to the activation of mTORC1 and the subsequent translation of select antiapoptotic proteins within infected monocytes. However, the full extent to which the HCMV-initiated Akt/mTORC1 signaling axis reshapes the monocyte translatome is unclear. We found HCMV entry alone was able to stimulate widescale changes to mRNA translation levels and that inhibition of mTOR, a component of mTORC1, dramatically attenuated HCMV-induced protein synthesis. Although monocytes treated with normal myeloid growth factors also exhibited increased levels of translation, mTOR inhibition had no effect, suggesting HCMV activation of mTOR stimulates the acquisition of a unique translatome within infected monocytes. Indeed, polyribosomal profiling of HCMV-infected monocytes identified distinct prosurvival transcripts that were preferentially loaded with ribosomes when compared to growth factor-treated cells. Sirtuin 1 (SIRT1), a deacetylase that exerts prosurvival effects through regulation of the PI3K/Akt pathway, was found to be highly enriched following HCMV infection in an mTOR-dependent manner. Importantly, SIRT1 inhibition led to the death of HCMV-infected monocytes while having minimal effect on uninfected cells. SIRT1 also supported a positive feedback loop to sustain Akt/mTORC1 signaling following viral entry. Taken together, HCMV profoundly reshapes mRNA translation in an mTOR-dependent manner to enhance the synthesis of select factors necessary for the survival of infected monocytes.IMPORTANCEHuman cytomegalovirus (HCMV) infection is a significant cause of morbidity and mortality among the immunonaïve and immunocompromised. Peripheral blood monocytes are a major cell type responsible for disseminating the virus from the initial site of infection. In order for monocytes to mediate viral spread within the host, HCMV must subvert the naturally short lifespan of these cells. In this study, we performed polysomal profiling analysis, which demonstrated HCMV to globally redirect mRNA translation toward the synthesis of cellular prosurvival factors within infected monocytes. Specifically, HCMV entry into monocytes induced the translation of cellular SIRT1 to generate an antiapoptotic state. Defining the precise mechanisms through which HCMV stimulates survival will provide insight into novel anti-HCMV drugs able to target infected monocytes.

Human Cytomegalovirus Manipulates Syntaxin 6 for Successful Trafficking and Subsequent Infection of Monocytes.

Human cytomegalovirus (HCMV) exhibits a complex host-pathogen interaction with peripheral blood monocytes. We have identified a unique, cell-type specific retrograde-like intracellular trafficking pattern that HCMV utilizes to gain access to the monocyte nucleus and for productive infection. We show that infection of primary human monocytes, epithelial cells, and fibroblasts leads to an increase in the amount of the trafficking protein Syntaxin 6 (Stx6). However, only knockdown (KD) of Stx6 in monocytes inhibited viral trafficking to the trans-Golgi network (TGN), a requisite step for nuclear translocation in monocytes. Conversely, KD of Stx6 in epithelial cells and fibroblasts did not change the kinetics of nuclear translocation and productive infection. Stx6 predominantly functions at the level of the TGN where it facilitates retrograde transport, a trafficking pathway used by only a few cellular proteins and seldom by pathogens. We also newly identify that in monocytes, Stx6 exhibits an irregular vesicular localization rather than being concentrated at the TGN as seen in other cell-types. Lastly, we implicate that viral particles that associate with both Stx6 and EEA1 early in infection are the viral population that successfully traffics to the TGN at later time points and undergo nuclear translocation. Additionally, we show for the first time that HCMV enters the TGN, and that lack of Stx6 prevents viral trafficking to this organelle. We argue that we have identified an essential cell-type specific regulator that controls early steps in efficient productive infection of a cell-type required for viral persistence and disease. IMPORTANCE Human cytomegalovirus (HCMV) infection causes severe and often fatal disease in the immunocompromised. It is one of the leading infectious causes of birth defects and causes severe complications in transplant recipients. By uncovering the unique pathways used by the virus to infect key cells, such as monocytes, responsible for dissemination and persistence, we provide new potential targets for therapeutic intervention.

Regulation of Siglec-7-mediated varicella-zoster virus infection of primary monocytes by cis-ligands.

Varicella-zoster virus (VZV) first infects hematopoietic cells, with the infected cells then acting to distribute the virus throughout the body. Sialic acid-binding immunoglobulin-like lectin (Siglec) family molecules recognize sialic acid-containing molecules on the same cell surface, called cis-ligands, or molecules on other cells or soluble agents, called trans-ligands. Among the Siglec family molecules, Siglec-4 and Siglec-7 mediate VZV infection through association with glycoprotein B (gB). As Siglec-7, but not Siglec-4, is expressed on hematopoietic cells such as monocytes, the regulatory mechanism by which Siglec-7 associates with gB is important to our understanding of VZV infection of blood cells. Here, we found that Siglec-7 is required for VZV to infect human primary monocytes. Furthermore, treatment of primary monocytes with sialidase enhanced both VZV gB binding to monocytes and VZV infectivity. Calcium influx in primary monocytes decreased the expression of Siglec-7 cis-ligands and increased VZV infectivity. These results demonstrate that the Siglec-7 cis-ligands present on primary monocytes play an important role in VZV infection through regulation of the interaction between gB and Siglec-7.

Vitamin D modulates the expression of Toll-like receptors and pro-inflammatory cytokines without affecting Chikungunya virus replication, in monocytes and macrophages.

Chikungunya virus (CHIKV) is a zoonotic arthropod-borne virus that causes Chikungunya fever (CHIKF), a self-limiting disease characterized by myalgia and acute or chronic arthralgia. CHIKF pathogenesis has an important immunological component since higher levels of pro-inflammatory factors, including cytokines and chemokines, are detected in CHIKV-infected patients. In vitro studies, using monocytes and macrophages have shown that CHIKV infection promotes elevated production of pro-inflammatory cytokines and antiviral response factors. Vitamin D3 (VD3) has been described as an important modulator of immune response and as an antiviral factor for several viruses. Here, we aimed to study the effects of VD3 treatment on viral replication and pro-inflammatory response in CHIKV-infected human monocytes (VD3-Mon) and monocyte-derived macrophages differentiated in the absence (MDMs) or the presence of VD3 (VD3-MDMs). We found that VD3 treatment did not suppress CHIKV replication in either VD3-Mon or VD3-MDMs. However, the expression of VDR, CAMP and CYP24A1 mRNAs was altered by CHIKV infection. Furthermore, VD3 treatment alters TLRs mRNA expression and production of pro-inflammatory cytokines, including TNFα and CXCL8/IL8, but not IL1ß and IL6, in response to CHIKV infection in both VD3-Mon and VD3-MDMs. While a significant decrease in CXCL8/IL8 production was observed in CHIKV-infected VD3-Mon, significantly higher production of CXCL8/IL8 was observed in CHIKV-infected VD3-MDM at 24 hpi. Altogether, our results suggest that vitamin D3 may play an important role in ameliorating pro-inflammatory response during CHIKV infection in human Mon, but not in MDMs. Although further studies are needed to evaluate the efficacy of VD3; nevertheless, this study provides novel insights into its benefits in modulating the inflammatory response elicited by CHIKV infection in humans.

Highly restricted SARS-CoV-2 receptor expression and resistance to infection by primary human monocytes and monocyte-derived macrophages.

Severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV2), which causes the disease COVID-19, has caused an unprecedented global pandemic. Angiotensin-converting enzyme 2 (ACE2) is the major cellular receptor for SARS-CoV2 entry, which is facilitated by viral Spike priming by cellular TMPRSS2. Macrophages play an important role in innate viral defense and are also involved in aberrant immune activation that occurs in COVID-19, and thus direct macrophage infection might contribute to severity of SARS-CoV2 infection. Here, we demonstrate that monocytes and monocyte-derived macrophages (MDM) under in vitro conditions express low-to-undetectable levels of ACE2 and TMPRSS2 and minimal coexpression. Expression of these receptors remained low in MDM induced to different subtypes such as unpolarized, M1 and M2 polarized. Untreated, unpolarized, M1 polarized, and M2 polarized MDM were all resistant to infection with SARS-CoV2 pseudotyped virions. These findings suggest that direct infection of myeloid cells is unlikely to be a major mechanism of SARS-CoV2 pathogenesis. Summary sentence: Monocytes and macrophages express minimal ACE2 and TMPRSS2 and resist SARS-CoV-2 Spike-mediated infection, suggesting direct myeloid cell infection is unlikely a major contributor to pathogenesis.

FcγR-mediated SARS-CoV-2 infection of monocytes activates inflammation.

SARS-CoV-2 can cause acute respiratory distress and death in some patients1. Although severe COVID-19 is linked to substantial inflammation, how SARS-CoV-2 triggers inflammation is not clear2. Monocytes and macrophages are sentinel cells that sense invasive infection to form inflammasomes that activate caspase-1 and gasdermin D, leading to inflammatory death (pyroptosis) and the release of potent inflammatory mediators3. Here we show that about 6% of blood monocytes of patients with COVID-19 are infected with SARS-CoV-2. Monocyte infection depends on the uptake of antibody-opsonized virus by Fcγ receptors. The plasma of vaccine recipients does not promote antibody-dependent monocyte infection. SARS-CoV-2 begins to replicate in monocytes, but infection is aborted, and infectious virus is not detected in the supernatants of cultures of infected monocytes. Instead, infected cells undergo pyroptosis mediated by activation of NLRP3 and AIM2 inflammasomes, caspase-1 and gasdermin D. Moreover, tissue-resident macrophages, but not infected epithelial and endothelial cells, from lung autopsies from patients with COVID-19 have activated inflammasomes. Taken together, these findings suggest that antibody-mediated SARS-CoV-2 uptake by monocytes and macrophages triggers inflammatory cell death that aborts the production of infectious virus but causes systemic inflammation that contributes to COVID-19 pathogenesis.

Activation of Complement Components on Circulating Blood Monocytes From COVID-19 Patients.

The coronavirus disease-2019 (COVID-19) caused by the SARS-CoV-2 virus may vary from asymptomatic to severe infection with multi-organ failure and death. Increased levels of circulating complement biomarkers have been implicated in COVID-19-related hyperinflammation and coagulopathy. We characterized systemic complement activation at a cellular level in 49-patients with COVID-19. We found increases of the classical complement sentinel C1q and the downstream C3 component on circulating blood monocytes from COVID-19 patients when compared to healthy controls (HCs). Interestingly, the cell surface-bound complement inhibitor CD55 was also upregulated in COVID-19 patient monocytes in comparison with HC cells. Monocyte membrane-bound C1q, C3 and CD55 levels were associated with plasma inflammatory markers such as CRP and serum amyloid A during acute infection. Membrane-bounds C1q and C3 remained elevated even after a short recovery period. These results highlight systemic monocyte-associated complement activation over a broad range of COVID-19 disease severities, with a compensatory upregulation of CD55. Further evaluation of complement and its interaction with myeloid cells at the membrane level could improve understanding of its role in COVID-19 pathogenesis.

A Viral Long Non-Coding RNA Protects against Cell Death during Human Cytomegalovirus Infection of CD14+ Monocytes.

Long non-coding RNA ß2.7 is the most highly transcribed viral gene during latent human cytomegalovirus (HCMV) infection. However, as yet, no function has ever been ascribed to ß2.7 during HCMV latency. Here we show that ß2.7 protects against apoptosis induced by high levels of reactive oxygen species (ROS) in infected monocytes, which routinely support latent HCMV infection. Monocytes infected with a wild-type (WT) virus, but not virus deleted for the ß2.7 gene (Δß2.7), are protected against mitochondrial stress and subsequent apoptosis. Protected monocytes display lower levels of ROS and additionally, stress-induced death in the absence of ß2.7 can be reversed by an antioxidant which reduces ROS levels. Furthermore, we show that infection with WT but not Δß2.7 virus results in strong upregulation of a cellular antioxidant enzyme, superoxide dismutase 2 (SOD2) in CD14+ monocytes. These observations identify a role for the ß2.7 viral transcript, the most abundantly expressed viral RNA during latency but for which no latency-associated function has ever been ascribed, and demonstrate a novel way in which HCMV protects infected monocytes from pro-death signals to optimise latent carriage.

Monocytes in HIV and SIV Infection and Aging: Implications for Inflamm-Aging and Accelerated Aging.

Before the antiretroviral therapy (ART) era, people living with HIV (PLWH) experienced complications due to AIDS more so than aging. With ART and the extended lifespan of PLWH, HIV comorbidities also include aging-most likely due to accelerated aging-as well as a cardiovascular, neurocognitive disorders, lung and kidney disease, and malignancies. The broad evidence suggests that HIV with ART is associated with accentuated aging, and that the age-related comorbidities occur earlier, due in part to chronic immune activation, co-infections, and possibly the effects of ART alone. Normally the immune system undergoes alterations of lymphocyte and monocyte populations with aging, that include diminished naïve T- and B-lymphocyte numbers, a reliance on memory lymphocytes, and a skewed production of myeloid cells leading to age-related inflammation, termed "inflamm-aging". Specifically, absolute numbers and relative proportions of monocytes and monocyte subpopulations are skewed with age along with myeloid mitochondrial dysfunction, resulting in increased accumulation of reactive oxygen species (ROS). Additionally, an increase in biomarkers of myeloid activation (IL-6, sCD14, and sCD163) occurs with chronic HIV infection and with age, and may contribute to immunosenescence. Chronic HIV infection accelerates aging; meanwhile, ART treatment may slow age-related acceleration, but is not sufficient to stop aging or age-related comorbidities. Overall, a better understanding of the mechanisms behind accentuated aging with HIV and the effects of myeloid activation and turnover is needed for future therapies.

Pathological effects of SARS-CoV-2 on hematological and immunological cells: Alterations in count, morphology, and function.

The novel Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), the causative agent of COVID-19 outbreak, spread rapidly and infected more than 140 million people with more than three million victims worldwide. The SARS-CoV-2 causes destructive changes in the immunological and hematological system of the host. These alterations appear to play a critical role in disease pathology and the emerging of clinical manifestations. In this review, we aimed to discuss the effect of COVID-19 on the count, function and morphology of immune and blood cells and the role of these changes in the pathophysiology of the disease. Knowledge of these changes may help with better management and treatment of COVID-19 patients.

Integrating single-cell sequencing data with GWAS summary statistics reveals CD16+monocytes and memory CD8+T cells involved in severe COVID-19.

BACKGROUND: Understanding the host genetic architecture and viral immunity contributes to the development of effective vaccines and therapeutics for controlling the COVID-19 pandemic. Alterations of immune responses in peripheral blood mononuclear cells play a crucial role in the detrimental progression of COVID-19. However, the effects of host genetic factors on immune responses for severe COVID-19 remain largely unknown. METHODS: We constructed a computational framework to characterize the host genetics that influence immune cell subpopulations for severe COVID-19 by integrating GWAS summary statistics (N = 969,689 samples) with four independent scRNA-seq datasets containing healthy controls and patients with mild, moderate, and severe symptom (N = 606,534 cells). We collected 10 predefined gene sets including inflammatory and cytokine genes to calculate cell state score for evaluating the immunological features of individual immune cells. RESULTS: We found that 34 risk genes were significantly associated with severe COVID-19, and the number of highly expressed genes increased with the severity of COVID-19. Three cell subtypes that are CD16+monocytes, megakaryocytes, and memory CD8+T cells were significantly enriched by COVID-19-related genetic association signals. Notably, three causal risk genes of CCR1, CXCR6, and ABO were highly expressed in these three cell types, respectively. CCR1+CD16+monocytes and ABO+ megakaryocytes with significantly up-regulated genes, including S100A12, S100A8, S100A9, and IFITM1, confer higher risk to the dysregulated immune response among severe patients. CXCR6+ memory CD8+ T cells exhibit a notable polyfunctionality including elevation of proliferation, migration, and chemotaxis. Moreover, we observed an increase in cell-cell interactions of both CCR1+ CD16+monocytes and CXCR6+ memory CD8+T cells in severe patients compared to normal controls among both PBMCs and lung tissues. The enhanced interactions of CXCR6+ memory CD8+T cells with epithelial cells facilitate the recruitment of this specific population of T cells to airways, promoting CD8+T cell-mediated immunity against COVID-19 infection. CONCLUSIONS: We uncover a major genetics-modulated immunological shift between mild and severe infection, including an elevated expression of genetics-risk genes, increase in inflammatory cytokines, and of functional immune cell subsets aggravating disease severity, which provides novel insights into parsing the host genetic determinants that influence peripheral immune cells in severe COVID-19.