The abortive SARS-CoV-2 infection of osteoclast precursors promotes their differentiation into osteoclasts.

The Coronavirus Disease 2019 (COVID-19) pandemic has resulted in the loss of millions of lives, although a majority of those infected have managed to survive. Consequently, a set of outcomes, identified as long COVID, is now emerging. While the primary target of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the respiratory system, the impact of COVID-19 extends to various body parts, including the bone. This study aims to investigate the effects of acute SARS-CoV-2 infection on osteoclastogenesis, utilizing both ancestral and Omicron viral strains. Monocyte-derived macrophages, which serve as precursors to osteoclasts, were exposed to both viral variants. However, the infection proved abortive, even though ACE2 receptor expression increased postinfection, with no significant impact on cellular viability and redox balance. Both SARS-CoV-2 strains heightened osteoclast formation in a dose-dependent manner, as well as CD51/61 expression and bone resorptive ability. Notably, SARS-CoV-2 induced early pro-inflammatory M1 macrophage polarization, shifting toward an M2-like profile. Osteoclastogenesis-related genes (RANK, NFATc1, DC-STAMP, MMP9) were upregulated, and surprisingly, SARS-CoV-2 variants promoted RANKL-independent osteoclast formation. This thorough investigation illuminates the intricate interplay between SARS-CoV-2 and osteoclast precursors, suggesting potential implications for bone homeostasis and opening new avenues for therapeutic exploration in COVID-19.

Long-term recruitment of peripheral immune cells to brain scars after a neonatal insult.

Although brain scars in adults have been extensively studied, there is less data available regarding scar formation during the neonatal period, and the involvement of peripheral immune cells in this process remains unexplored in neonates. Using a murine model of neonatal hypoxic-ischemic encephalopathy (HIE) and confocal microscopy, we characterized the scarring process and examined the recruitment of peripheral immune cells to cortical and hippocampal scars for up to 1 year post-insult. Regional differences in scar formation were observed, including the presence of reticular fibrotic networks in the cortex and perivascular fibrosis in the hippocampus. We identified chemokines with chronically elevated levels in both regions and demonstrated, through a parabiosis-based strategy, the recruitment of lymphocytes, neutrophils, and monocyte-derived macrophages to the scars several weeks after the neonatal insult. After 1 year, however, neutrophils and lymphocytes were absent from the scars. Our data indicate that peripheral immune cells are transient components of HIE-induced brain scars, opening up new possibilities for late therapeutic interventions.

Polyacrylic acid-coated iron oxide nanoparticles could be a useful tool for tracking inflammatory monocytes.

AIM: To establish the effect of poly(acrylic acid)-coated iron oxide nanoparticles (PAC-IONs) and later exposure to a magnetic field on the differentiation of mononuclear phagocytes into macrophages. METHODS: By flow cytometry, cell death was evaluated with DIOC6 and PI, Poly (ADP-ribose) Polymerases (PARP) fragmentation, H2AX phosphorylation and TUNEL assay. Cytokines by Cytokine bead array and the intracellular amount of iron by atomic absorption spectrometry. RESULTS: PAC-IONs did not induce apoptosis, modify the cell membrane integrity or alter the mitochondrial membrane potential. They did not affect the cell morphology, the pattern of cytokine accumulation or the activating role of differentiation of mononuclear phagocytes into macrophages on the proliferation of autologous T cells. CONCLUSION: This evidence indicates that the PAC-IONs are safe and biocompatible. Moreover, the selectivity of the PAC-IONs for mononuclear phagocytes, as well as their increased uptake by non-classical monocytes, warrant future research with a view to their use as a contrast agent, a useful tool for in vivo tracking of tissue-infiltrating mononuclear phagocytes.

Use of Human Monocyte-Derived Macrophages to Study Neisseria gonorrhoeae Infection.

Macrophages are critical cells in the innate immune response to microorganisms sensed in the tissues. During infections, the interaction between pathogens and macrophages leads to a macrophage response that includes cytokine production, antigen processing and presentation in the context of MHC molecules, expression of T cell costimulatory molecules and recruitment of innate defense effectors, which results in clearance of infection. However, Neisseria gonorrhoeae can suppress the protective immune response at this level, avoiding its detection and elimination. Studies addressed to develop the interactions between macrophages and Neisseria gonorrhoeae allow us to find potential targets to be exploited with vaccines and therapeutic drugs. In this chapter, we describe protocols to generate human monocyte-derived macrophages and assess their response to infection with Neisseria gonorrhoeae.

Interaction Between Macrophage Migration Inhibitory Factor and CD74 in Human Immunodeficiency Virus Type I Infected Primary Monocyte-Derived Macrophages Triggers the Production of Proinflammatory Mediators and Enhances Infection of Unactivated CD4+ T Cells.

Understanding the mechanisms of human immunodeficiency virus type I (HIV-1) pathogenesis would facilitate the identification of new therapeutic targets to control the infection in face of current antiretroviral therapy limitations. CD74 membrane expression is upregulated in HIV-1-infected cells and the magnitude of its modulation correlates with immune hyperactivation in HIV-infected individuals. In addition, plasma level of the CD74 activating ligand macrophage migration inhibitory factor (MIF) is increased in infected subjects. However, the role played by MIF/CD74 interaction in HIV pathogenesis remains unexplored. Here, we studied the effect of MIF/CD74 interaction on primary HIV-infected monocyte-derived macrophages (MDMs) and its implications for HIV immunopathogenesis. Confocal immunofluorescence analysis of CD74 and CD44 (the MIF signal transduction co-receptor) expression indicated that both molecules colocalized at the plasma membrane specifically in wild-type HIV-infected MDMs. Treatment of infected MDMs with MIF resulted in an MIF-dependent increase in TLR4 expression. Similarly, there was a dose-dependent increase in the production of IL-6, IL-8, TNFα, IL-1ß, and sICAM compared to the no-MIF condition, specifically from infected MDMs. Importantly, the effect observed on IL-6, IL-8, TNFα, and IL-1ß was abrogated by impeding MIF interaction with CD74. Moreover, the use of a neutralizing αMIF antibody or an MIF antagonist reverted these effects, supporting the specificity of the results. Treatment of unactivated CD4+ T-cells with MIF-treated HIV-infected MDM-derived culture supernatants led to enhanced permissiveness to HIV-1 infection. This effect was lost when CD4+ T-cells were treated with supernatants derived from infected MDMs in which CD74/MIF interaction had been blocked. Moreover, the enhanced permissiveness of unactivated CD4+ T-cells was recapitulated by exogenous addition of IL-6, IL-8, IL-1ß, and TNFα, or abrogated by neutralizing its biological activity using specific antibodies. Results obtained with BAL and NL4-3 HIV laboratory strains were reproduced using transmitted/founder primary isolates. This evidence indicated that MIF/CD74 interaction resulted in a higher production of proinflammatory cytokines from HIV-infected MDMs. This caused the generation of an inflammatory microenvironment which predisposed unactivated CD4+ T-cells to HIV-1 infection, which might contribute to viral spreading and reservoir seeding. Overall, these results support a novel role of the MIF/CD74 axis in HIV pathogenesis that deserves further investigation.

Functional evaluation of gene silencing on macrophages derived from U937 cells using interference RNA (shRNA) in a model of macrophages infected with Leishmania (Viannia) braziliensis.

Leishmaniasis development is multifactorial; nonetheless, the establishment of the infection, which occurs by the survival and replication of the parasite inside its main host cell, the macrophage, is mandatory. Thus, the importance of studying the molecular mechanisms involved in the Leishmania-macrophage interaction is highlighted. The aim of this study was to characterize a cellular model of macrophages derived from U937 cells that would allow for the identification of infection phenotypes induced by genetic silencing with interference RNA in the context of macrophages infected with Leishmania (Viannia) braziliensis. The model was standardized by silencing an exogenous gene (gfp), an endogenous gene (lmna) and a differentially expressed gene between infected and non-infected macrophages (gro-ß). The silencing process was successful for the three genes studied, obtaining reductions of 88·9% in the GFP levels, 87·5% in LMNA levels and 74·4% for Gro-ß with respect to the corresponding control cell lines. The cell model revealed changes in the infection phenotype of the macrophages in terms of number of amastigotes per infected macrophage, number of amastigotes per sampled macrophage and percentage of infected macrophages as a result of gene silencing. Thus, this cell model constitutes a research platform for the study of parasite-host interactions and for the identification of potentially therapeutic targets.

HIV gp120 sequence variability associated with HAND in Hispanic Women.

OBJECTIVE: HIV-1 variants with different tropisms are associated with various neuropathologies. This study intends to determine if this correlation is determined by unique viral env sequences. We hypothesize that HIV-1 envelope gene sequence changes are associated with cognition status. METHODS: Viral RNA was extracted from peripheral blood mononuclear cells (PBMCs) co-cultures derived from HIV-1 infected Hispanic women that had been characterized for HIV associated neurocognitive disorders (HAND). RESULTS: Analyses of the C2V4 region of HIV gp120 demonstrated that increased sequence diversity correlates with cognition status as sequences derived from subjects with normal cognition exhibited less diversity than sequences derived from subjects with cognitive impairment. In addition, differences in V3 and V4 loop charges were also noted as well as differences in the N-glycosylation of the V4 region. CONCLUSIONS: Our data suggest that the genetic signature within the C2V4 region may contribute to the pathogenesis of HAND. HIV env sequence characteristics for the isolates grouped in milder forms of HAND can provide insightful information of prognostic value to assess neurocognitive status in HIV+ subjects, particularly during the era of highly prevalent milder forms of HAND.