Antibody and T-cell responses to coronavirus disease 2019 vaccination in common variable immunodeficiency and specific antibody deficiency

BACKGROUND: Clinical trials of the mRNA coronavirus disease 2019 (COVID-19) vaccines excluded individuals with primary antibody deficiencies. OBJECTIVE: To evaluate whether antibody and T-cell responses to mRNA COVID-19 vaccination in patients with common variable immunodeficiency (CVID) and specific antibody deficiency (SAD) were comparable to those in healthy controls. METHODS: We measured antibody responses against the spike glycoprotein and the receptor-binding domain (RBD) in addition to severe acute respiratory syndrome coronavirus 2 specific T-cell responses using peripheral blood mononuclear cells 2 to 8 weeks after the subjects completed the primary 2-dose vaccine series. RESULTS: The study comprised 12 patients with CVID, 7 patients with SAD, and 10 controls. Individuals with CVID had lower immunoglobulin (Ig) G and Ig A levels against spike glycoprotein than did both individuals with SAD (P = .27 and P = .01, respectively) and controls (P = .01 and P = .004, respectively). The CVID group developed lower IgG titers against the RBD epitope than did the control group (P = .01). Participants with CVID had lower neutralizing titers than did the control group (P = .002). All participants with SAD developed neutralizing titers. All 3 groups (SAD, CVID, and control) developed antigen-specific CD4 and CD8 T-cell responses after vaccination. CONCLUSION: Our results suggest that patients with CVID may have impaired antibody responses to COVID-19 vaccination but intact T-cell responses, whereas patients with SAD would be expected to have both intact antibody and T-cell responses to vaccination

Dengue Virus Directly Stimulates Polyclonal B Cell Activation

<div><p>Dengue infection is associated to vigorous inflammatory response, to a high frequency of activated B cells, and to increased levels of circulating cross-reactive antibodies. We investigated whether direct infection of B cells would promote activation by culturing primary human B lymphocytes from healthy donors with DENV <i>in vitro</i>. B cells were susceptible, but poorly permissive to infection. Even though, primary B cells cultured with DENV induced substantial IgM secretion, which is a hallmark of polyclonal B cell activation. Notably, DENV induced the activation of B cells obtained from either DENV immune or DENV naïve donors, suggesting that it was not dependent on DENV-specific secondary/memory response. B cell stimulation was dependent on activation of MAPK and CD81. B cells cultured with DENV also secreted IL-6 and presented increased expression of CD86 and HLA-DR, which might contribute to B lymphocyte co-stimulatory function. Indeed, PBMCs, but not isolated B cells, secreted high amounts of IgG upon DENV culture, suggesting that interaction with other cell types <i>in vivo</i> might promote Ig isotype switching and IgG secretion from different B cell clones. These findings suggest that activation signaling pathways triggered by DENV interaction with non-specific receptors on B cells might contribute to the exacerbated response observed in dengue patients.</p></div

Infection of Endothelial Cells by Dengue Virus Induces ROS Production by Different Sources Affecting Virus Replication, Cellular Activation, Death and Vascular Permeability

Exacerbated inflammatory response and altered vascular function are hallmarks of dengue disease. Reactive oxygen species (ROS) production has been associated to endothelial barrier disturbance and microvascular alteration in distinct pathological conditions. Increased ROS has been reported in models of dengue virus (DENV) infection, but its impact for endothelial cell physiology had not been fully investigated. Our group had previously demonstrated that infection of human brain microvascular endothelial cells (HBMEC) with DENV results in the activation of RNA sensors and production of proinflammatory cytokines, which culminate in cell death and endothelial permeability. Here, we evaluated the role of mitochondrial function and NADPH oxidase (NOX) activation for ROS generation in HBMEC infected by DENV and investigated whether altered cellular physiology could be a consequence of virus-induced oxidative stress. DENV-infected HBMECs showed a decrease in the maximal respiratory capacity and altered membrane potential, indicating functional mitochondrial alteration, what might be related to mtROS production. Indeed, mtROS was detected at later time points after infection. Specific inhibition of mtROS diminished virus replication, cell death, and endothelial permeability, but did not affect cytokine production. On the other hand, inhibition of NOX-associated ROS production decreased virus replication and cell death, as well as the secretion of inflammatory cytokines, including IL-6, IL-8, and CCL5. These results demonstrated that DENV replication in endothelial cells induces ROS production by different pathways, which impacts biological functions that might be relevant for dengue pathogenesis. Those data also indicate oxidative stress events as relevant therapeutical targets to avoid vascular permeability, inflammation, and neuroinvasion during DENV infection

Purified B cells cultured with dengue virus showed increased expression of costimulatory molecules.

<p>B lymphocytes were mock-treated or cultured with DENV2 (MOI = 1) for the indicated time points and the expression of CD86 (A) or HLA-DR (B) in CD19+ cells were evaluated by flow cytometry. Each point indicate an individual donor. Statistical analysis were performed and p values are indicated in the figures.</p

DENV-induced B cell activation depends on CD81 activation.

<p>B lymphocytes were mock-treated or cultured with DENV2 (MOI = 1) in the presence or absence of anti-CD81 neutralizing antibody. <b>A)</b> At 12 days post infection, the supernatants were harvested and IgM levels were measured by ELISA. Data are representative of four independent experiments. Statistical analysis were performed and p values are indicated in the figures. <b>B)</b> The cells were harvested after different time points and DENV RNA levels were evaluated by qRT-PCR. Data are representative of three independent experiments. <b>C)</b> After 48h, the cells were harvested and the expression of ERK, p38 and JNK MAPK, phosphorylated (phospho) or not (unphospho) were analyzed in the cell lysates by western blotting, as indicated. The bars indicate the ratio between the analyzed phosphorylated protein and the corresponding unphosphorylated one; dots represent individual data.</p

DENV-induced IgM secretion does not depend on TLR4 activation.

<p>B lymphocytes were mock-treated or cultured with DENV2 (MOI = 1) in the presence or absence of anti-TLR4 neutralizing antibody. LPS (10μg/ml; Sigma Aldrich) was used as positive control. At 12 days post infection, the supernatants were harvested and IgM levels were measured by ELISA. B cell cultures which showed increased IgM secretion in response to LPS (LPS responders) were graphed separately from the ones that didn’t show (LPS nonresponders). Data are representative of four independent experiments. Statistical analysis were performed and p values are indicated in the figures.</p

Bradykinin enhances Sindbis virus infection in human brain microvascular endothelial cells

Submitted by Nuzia Santos ([email protected]) on 2014-12-04T15:11:49Z No. of bitstreams: 1 Bradykinin enhances Sindbis virus infection in human brain microvascular endothelial cells.pdf: 1158448 bytes, checksum: 5c208b59273623f263acf5a38adfebf2 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2014-12-04T15:16:21Z (GMT) No. of bitstreams: 1 Bradykinin enhances Sindbis virus infection in human brain microvascular endothelial cells.pdf: 1158448 bytes, checksum: 5c208b59273623f263acf5a38adfebf2 (MD5)Made available in DSpace on 2014-12-04T15:16:21Z (GMT). No. of bitstreams: 1 Bradykinin enhances Sindbis virus infection in human brain microvascular endothelial cells.pdf: 1158448 bytes, checksum: 5c208b59273623f263acf5a38adfebf2 (MD5) Previous issue date: 2011Universidade Federal do Rio de Janeiro. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Virologia. Rio de Janeiro, RJ. BrasilUniversidade Federal do Rio de Janeiro. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Virologia. Rio de Janeiro, RJ. BrasilUniversidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Rio de Janeiro, RJ. BrasilFundação Oswaldo Cruz. Centro de Pesquisas Aggeu Magalhães. Laboratório de Virologia e Terapia Experimental. Recife, PE, BrasilFundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Laboratório de Imunologia Celular e Molecular. Belo Horizonte, MG, BrasilFundação Oswaldo Cruz. Centro de Pesquisas Aggeu Magalhães. Laboratório de Virologia e Terapia Experimental. Recife, PE, BrasilUniversidade Federal do Rio de Janeiro. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Imunologia Rio de Janeiro, RJ, BrasilUniversidade Federal do Rio de Janeiro. Instituto de Biofísica Carlos Chagas Filho. Rio de Janeiro, RJ. BrasilUniversidade Federal do Rio de Janeiro. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Virologia. Rio de Janeiro, RJ. BrasilSindbis virus (SINV) induces inflammatory and vasoactive responses that are associated with rash and arthritis in human infections. The mechanisms underlying infection-associated microvasculopathy are still unknown. We investigated whether endothelial cells infected by SINV are differentially responsive to bradykinin (BK), a potent inducer of inflammatory edema in a broad range of infectious diseases. Human endothelial cells (HBMECs) infected with SINV presented an upregulation of bradykinin B2 receptors (BK2R) expression. Also, BK reduced SINV-induced apoptosis and enhanced virus replication in HBMECs in a way dependent on BK2R, PI3 kinase and ERK signaling. Strikingly, intracerebral infection of mice in the presence of a BK2R antagonist reduced the local viral load. Our data suggest that SINV infection renders human endothelial cells hypersensitive to BK, which increases host cell survival and viral replication. Ongoing studies may clarify if the deregulation of the kinin pathway contributes to infection-associated vasculopathies in life-threatening arbovirus infection

DENV infection of PBMC promotes IgG secretion.

<p><b>A)</b> PBMCs or purified B cells were mock-treated or cultured with DENV2. After 12 days p.i., the supernatants were harvested and IgG secretion was evaluated by ELISA. Squares (mock) and triangles (DENV) indicate individual donors. <b>B)</b> PBMCs obtained from DENV immune (serum IgG+) and DENV naïve (serum IgG-) donors were mock-treated or cultured with DENV2, and IgM secretion was evaluated after 12 days. Stimulation index was calculated as the ratio between IgM concentration obtained from DENV and mock treated cultures. Dots indicate individual donors. <b>C-D)</b> PBMCs, CD3-CD27- cells (naïve Bcells), or CD3-CD27- and CD19+ cells (coculture between naïve B cells and non B cells), from the same donor were mock treated or DENV-infected. After 12 days p.i., the supernatants were harvested and IgM <b>(C)</b> or IgG <b>(D)</b> secretion were evaluated by ELISA. Bars indicate the average and SD of, at least, two independent experiments.</p

B cell infection by DENV promotes MAPK phosphorylation.

<p>B lymphocytes were mock-treated or cultured with DENV2 (MOI = 1). The cells were harvested after 2h, 24h or 48h p.i., and the expression of ERK, p38 and JNK MAPK were analyzed in the cell lysates by western blotting, using the indicated antibodies. The bars indicate the ratio between the analyzed phosphorylated protein and the corresponding non phosphorylated one; β actin staining were performed as a loading control and is shown in the bottom of the figure. Data are representative of three independent experiments.</p

Activation of MAPK is essential for DENV-induced B cell activation.

<p>Mock or DENV-treated cells were cultured in the presence or absence of ERK (PD98059), p38 (SB203580) and JNK (SP600125) inhibitors. <b>A)</b> After 12 days culture, supernatants were harvested and IgM levels were measured by ELISA. <b>B)</b> After 48h, culture supernatants were harvested and IL-6 levels were measured by ELISA. Data are representative of at least four independent experiments. Statistical analysis were performed and p values are indicated in the figures.</p