Zika virus spreads through infection of lymph node-resident macrophages

To disseminate through the body, Zika virus (ZIKV) is thought to exploit the mobility of myeloid cells, in particular monocytes and dendritic cells. However, the timing and mechanisms underlying shuttling of the virus by immune cells remains unclear. To understand the early steps in ZIKV transit from the skin, at different time points, we spatially mapped ZIKV infection in lymph nodes (LNs), an intermediary site en route to the blood. Contrary to prevailing hypotheses, migratory immune cells are not required for the virus to reach the LNs or blood. Instead, ZIKV rapidly infects a subset of sessile CD16

An agonistic anti-CD137 antibody disrupts lymphoid follicle structure and T-cell-dependent antibody responses

CD137 is a costimulatory receptor expressed on natural killer cells, T cells, and subsets of dendritic cells. An agonistic monoclonal antibody (mAb) against CD137 has been used to reduce tumor burden or reverse autoimmunity in animal models and clinical trials. Here, we show that mice treated with an agonistic anti-CD137 mAb have reduced numbers of germinal center (GC) B cells and follicular dendritic cells (FDCs) in lymphoid tissues, which impair antibody responses to multiple T-cell-dependent antigens, including infectious virus, viral proteins, and conjugated haptens. These effects are not due to enhanced apoptosis or impaired proliferation of B cells but instead correlate with changes in lymphoid follicle structure and GC B cell dispersal and are mediated by CD137 signaling in CD

An agonistic anti-CD137 antibody disrupts lymphoid follicle structure and T-cell-dependent antibody responses

CD137 is a costimulatory receptor expressed on natural killer cells, T cells, and subsets of dendritic cells. An agonistic monoclonal antibody (mAb) against CD137 has been used to reduce tumor burden or reverse autoimmunity in animal models and clinical trials. Here, we show that mice treated with an agonistic anti-CD137 mAb have reduced numbers of germinal center (GC) B cells and follicular dendritic cells (FDCs) in lymphoid tissues, which impair antibody responses to multiple T-cell-dependent antigens, including infectious virus, viral proteins, and conjugated haptens. These effects are not due to enhanced apoptosis or impaired proliferation of B cells but instead correlate with changes in lymphoid follicle structure and GC B cell dispersal and are mediated by CD137 signaling in CD

Chemokines control naive CD8+ T cell selection of optimal lymph node antigen presenting cells

CCR5-binding chemokines produced in the draining lymph node after vaccinia virus infection guide naive CD8+ T cells toward DCs and away from the macrophage-rich zone, thereby facilitating optimal CD8+ T cell activation and cytokine production

Locally Produced IL-10 Limits Cutaneous Vaccinia Virus Spread.

Skin infection with the poxvirus vaccinia (VV) elicits a powerful, inflammatory cellular response that clears virus infection in a coordinated, spatially organized manner. Given the high concentration of pro-inflammatory effectors at areas of viral infection, it is unclear how tissue pathology is limited while virus-infected cells are being eliminated. To better understand the spatial dynamics of the anti-inflammatory response to a cutaneous viral infection, we first screened cytokine mRNA expression levels after epicutaneous (ec.) VV infection and found a large increase the anti-inflammatory cytokine IL-10. Ex vivo analyses revealed that T cells in the skin were the primary IL-10-producing cells. To understand the distribution of IL-10-producing T cells in vivo, we performed multiphoton intravital microscopy (MPM) of VV-infected mice, assessing the location and dynamic behavior of IL-10 producing cells. Although virus-specific T cells were distributed throughout areas of the inflamed skin lacking overt virus-infection, IL-10+ cells closely associated with large keratinocytic foci of virus replication where they exhibited similar motility patterns to bulk antigen-specific CD8+ T cells. Paradoxically, neutralizing secreted IL-10 in vivo with an anti-IL-10 antibody increased viral lesion size and viral replication. Additional analyses demonstrated that IL-10 antibody administration decreased recruitment of CCR2+ inflammatory monocytes, which were important for reducing viral burden in the infected skin. Based upon these findings, we conclude that spatially concentrated IL-10 production limits cutaneous viral replication and dissemination, likely through modulation of the innate immune repertoire at the site of viral growth

CXCR3 Chemokine Receptor Enables Local CD8+ T Cell Migration for the Destruction of Virus-Infected Cells

SummaryCD8+ T cells play a critical role in limiting peripheral virus replication, yet how they locate virus-infected cells within tissues is unknown. Here, we have examined the environmental signals that CD8+ T cells use to localize and eliminate virus-infected skin cells. Epicutaneous vaccinia virus (VV) infection, mimicking human smallpox vaccination, greatly increased expression of the CXCR3 chemokine receptor ligands CXCL9 and CXCL10 in VV-infected skin. Despite normal T cell numbers in the skin, Cxcr3−/− mice exhibited dramatically impaired CD8+-T-cell-dependent virus clearance. Intravital microscopy revealed that Cxcr3−/− T cells were markedly deficient in locating, engaging, and killing virus-infected cells. Further, transfer of wild-type CD8+ T cells restored viral clearance in Cxcr3−/− animals. These findings demonstrate a function for CXCR3 in enhancing the ability of tissue-localized CD8+ T cells to locate virus-infected cells and thereby exert anti-viral effector functions

MyD88-dependent influx of monocytes and neutrophils impairs lymph node B cell responses to chikungunya virus infection via Irf5, Nos2 and Nox2

Author summary Elucidating mechanisms by which viruses subvert B cell immunity and establish persistent infection is essential for the development of new therapeutic strategies against chronic viral infections. The humoral immune response initiates in the lymph node draining the site of viral infection. However, how persistent viruses evade B cell responses is poorly understood. In this study, we find that infection with pathogenic, persistent chikungunya virus triggers rapid recruitment of neutrophils and monocytes to the draining lymph node, which impair structural organization, lymphocyte accumulation, and downstream virus-specific B cell responses that are important for control of infection. This work enhances our understanding of the pathogenesis of acute and chronic CHIKV disease and highlights how local innate immune responses in draining lymphoid tissue dictate the effectiveness of downstream adaptive immunity. Humoral immune responses initiate in the lymph node draining the site of viral infection (dLN). Some viruses subvert LN B cell activation; however, our knowledge of viral hindrance of B cell responses of important human pathogens is lacking. Here, we define mechanisms whereby chikungunya virus (CHIKV), a mosquito-transmitted RNA virus that causes outbreaks of acute and chronic arthritis in humans, hinders dLN antiviral B cell responses. Infection of WT mice with pathogenic, but not acutely cleared CHIKV, induced MyD88-dependent recruitment of monocytes and neutrophils to the dLN. Blocking this influx improved lymphocyte accumulation, dLN organization, and CHIKV-specific B cell responses. Both inducible nitric oxide synthase (iNOS) and the phagocyte NADPH oxidase (Nox2) contributed to impaired dLN organization and function. Infiltrating monocytes expressed iNOS through a local IRF5- and IFNAR1-dependent pathway that was partially TLR7-dependent. Together, our data suggest that pathogenic CHIKV triggers the influx and activation of monocytes and neutrophils in the dLN that impairs virus-specific B cell responses

IL-10gfp⁺ CD8⁺ T cells closely localize to areas of virus infection.

<p>Maximum intensity projections (MIP) of MPM image of ears from IL-10gfp reporter mice taken on day 6 pi. Collagen = blue, rVV-infected cells = blue pseudocolored magenta, IL-10gfp⁺ cells = green, OT-I CD8⁺ T cells = red. <b>A</b>) MIP MPM images showing distribution of —10gfp+ T cells (green) versus non-IL-10gfp-transgenic OT-I CD8⁺ T cells (red). Left panel = merge, middle panel = only green and “virus” channels, right panel = only red and “virus” channels. <b>B</b>) As in A) but showing an outlying area without rVV-infected cells. Circles indicate IL-10gfp⁺ cells. <b>C</b>) Lines showing the distance of each cell to the border of the area of rVV-infection. Green lines show distance of IL-10gfp⁺ cells; red line OT-I CD8⁺ T cells. <b>D</b>) Calculated distance of each cell type to rVV lesion border. Line = mean. Statistics = student’s two-tailed t-test. <b>E</b>) Number of cells at each distance to the lesion border for each class of T cells. <b>F</b>) Percentage of each T cell class at indicated distance to lesion border. Scale bars = microns.</p

Kinetics of anti-VV IL-10⁺ CD8⁺ T cell responses.

<p><b>A)</b> Viral titers after epicutanous infection of the ear as determined by plaque assay at indicated day p.i. N = 3 <b>B)</b> Time course of total CD4⁺ (green lines) and CD8⁺ T cells (blue lines) in the ear skin. N = 4. <b>C)</b> IL-10⁺CD4⁺ (green) and IL-10⁺CD8⁺ (blue) T cells. N = 4. <b>D)</b> Percentage of IL-10gfp⁺ CD8⁺ T cells of total CD8⁺ T cells in the ear skin (circle), draining lymph node (dLN, square), or spleen (triangle). <b>E)</b> Percentage of IL-10gfp⁺ CD4⁺ T cells of the total CD4 T cells as in (D). Error bars = SEM. **** = P <0.0001.</p

IL-10 neutralization alters tissue monocyte numbers.

<p><b>A</b>) Percent (top, <i>p = 0</i>.<i>009</i>) and number (bottom, <i>p = 0</i>.<i>03</i>) of CCR2⁺ monocytes per ear on day 7 p.i. in either control mice or in mice given IL-10 neutralizing Ab (IL-10 NAb, black bars) on days 5 and 6 p.i. <b>B-C</b>) MPM images of CCR2rfp⁺/^- mice (with red monocytes) either uninfected (left panels) or 7 d.p.i. (right panels) with VV-NP-S-eGFP (green). For clarity, red channel only is shown in bottom panels. <b>D</b>) GFP⁺ (green) infected CCR2rfp⁺ monocytes. For clarity, single channels are shown. <b>E</b>) Timecourse MPM images showing movement of an infected (green) monocyte (red). Dashed circles follow one such infected cell. Time = min. <b>F</b>) Viral titers in CCR2rfp⁺/^- or control mice 7 d.p.i. (in genomes/μg of ear DNA). N = 3, <i>p = 0</i>.<i>0004</i>. Scale bars = microns.</p