ABSTRACT
Macrophages are paracrine signalers that regulate tissular responses to injury through interactions with parenchymal cells. Connexin hemichannels have recently been shown to mediate efflux of ATP by macrophages, with resulting cytosolic calcium responses in adjacent cells. Here we report that lung macrophages with deletion of connexin 43 (MacΔCx43) had decreased ATP efflux into the extracellular space and induced a decreased cytosolic calcium response in co-cultured fibroblasts compared to WT macrophages. Furthermore, MacΔCx43 mice had decreased lung fibrosis after bleomycin-induced injury. Interrogating single cell data for human and mouse, we found that P2rx4 was the most highly expressed ATP receptor and calcium channel in lung fibroblasts and that its expression was increased in the setting of fibrosis. Fibroblast-specific deletion of P2rx4 in mice decreased lung fibrosis and collagen expression in lung fibroblasts in the bleomycin model. Taken together, these studies reveal a Cx43-dependent profibrotic effect of lung macrophages and support development of fibroblast P2rx4 as a therapeutic target for lung fibrosis.
Subject(s)
Connexin 43 , Idiopathic Pulmonary Fibrosis , Adenosine Triphosphate/metabolism , Animals , Bleomycin/pharmacology , Calcium/metabolism , Connexin 43/genetics , Connexin 43/metabolism , Fibroblasts/metabolism , Idiopathic Pulmonary Fibrosis/metabolism , Macrophages/metabolism , Mice , Mice, KnockoutABSTRACT
Immunoglobulin E (IgE) is the least abundant antibody isotype in mammalians, yet it plays a critical role in allergy and asthma. IgE-producing (IgE+) B cells are rare and difficult to detect, which have hindered progress to understand their generation and differentiation. Recently developed new fluorescent IgE reporter mice have enabled better understanding of the biology of IgE+ B cells. We here describe the usage of the Verigem IgE reporter mouse to study IgE+ B cells and plasma cells by flow cytometry and microscopy.
Subject(s)
Antibody Formation/immunology , B-Lymphocytes/immunology , B-Lymphocytes/metabolism , Immunoglobulin E/biosynthesis , Immunoglobulin E/immunology , Animals , Antibody Formation/genetics , Cell Culture Techniques , Flow Cytometry , Gene Expression , Genes, Reporter , Genetic Loci , Germinal Center/immunology , Germinal Center/metabolism , Immunization , Immunoglobulin E/genetics , Mice , Microscopy , Plasma Cells/immunology , Plasma Cells/metabolismABSTRACT
Cocaine abuse has been shown to have broad-ranging effects on human immunity. With regards to HIV infection, in vitro studies have shown that cocaine enhances infection of stimulated lymphocytes. Moreover, cohort studies in the pre- and post-HAART era have linked stimulant abuse with increased HIV pathogenesis. The latter data, however, have been undermined by a series of confounding factors underscoring the importance of controlled in vivo models to fully assess the impact of cocaine use and abuse on HIV infection and pathogenesis. Here, we have infected humanized mice with HIV-1 following acute cocaine exposure to assess the impact on infection. Stimulant exposure resulted in increased inflammatory cytokine expression, accelerated HIV infection, while blunting effector function of cytotoxic T lymphocytes. These data demonstrate cocaine's multifactorial impact on HIV infection that extends beyond high-risk behavior.
Subject(s)
CD8-Positive T-Lymphocytes/immunology , Cocaine/adverse effects , Cytokines/immunology , Gene Expression Regulation/drug effects , HIV Infections/immunology , HIV-1/immunology , Animals , CD8-Positive T-Lymphocytes/pathology , Cocaine/pharmacology , Disease Models, Animal , Gene Expression Regulation/immunology , HIV Infections/pathology , Humans , MiceABSTRACT
In vivo and in vitro exposure to stimulants has been associated with increased levels of HIV infection in PBMCs. Among these lymphocyte subsets, quiescent CD4(+) T cells make up the majority of circulating T cells in the blood. Others and we have demonstrated that HIV infects this population of cells inefficiently. However, minor changes in their cell state can render them permissive to infection, significantly impacting the viral reservoir. We have hypothesized that stimulants, such as cocaine, may perturb the activation state of quiescent cells enhancing permissiveness to infection. Quiescent T cells isolated from healthy human donors were exposed to cocaine and infected with HIV. Samples were harvested at different time-points to assess the impact of cocaine on their susceptibility to infection at various stages of the HIV life cycle. Our data show that a 3-day exposure to cocaine enhanced infection of quiescent cells, an effect that appears to be mediated by σ1R and D4R. Overall, our results indicate that cocaine-mediated effects on quiescent T cells may increase the pool of infection-susceptible T cells. The latter underscores the impact that stimulants have on HIV-seropositive individuals and the challenges posed for treatment.
