Distinct Chemokine Triggers and In Vivo Migratory Paths of Fluorescein Dye-Labeled T Lymphocytes in Acutely Simian Immunodeficiency Virus SIV(mac251)-Infected and Uninfected Macaques

To define the possible impact of T-lymphocyte trafficking parameters on simian immunodeficiency virus (SIV) pathogenesis, we examined migratory profiles of carboxyfluorescein diacetate succinimidyl ester (CFSE)-labeled T lymphocytes in acutely SIV(mac251)-infected and uninfected macaques within 48 h after autologous transfer. Despite significant upregulation of homeostatic chemokine CCL19/macrophage inflammatory protein 3β and proinflammatory chemokine CXCL9/monokine induced by gamma interferon in secondary lymphoid tissue in SIV infection, no differences in CFSE(+) T-lymphocyte frequencies or cell compartmentalization in lymph nodes were identified between animal groups. By contrast, a higher frequency of CFSE(+) T lymphocytes in the small intestine was detected in acute SIV infection. This result correlated with increased numbers of gut CD4 T lymphocytes expressing chemokine receptors CCR9, CCR7, and CXCR3 and high levels of their respective chemokine ligands in the small intestine. The changes in trafficking parameters in SIV-infected macaques occurred concomitantly with acute gut CD4 T-lymphocyte depletion. Here, we present the first in vivo T-lymphocyte trafficking study in SIV infection and a novel approach to delineate T-lymphocyte recruitment into tissues in the nonhuman primate animal model for AIDS. Such studies are likely to provide unique insights into T-lymphocyte sequestration in distinct tissue compartments and possible mechanisms of CD4 T-lymphocyte depletion and immune dysfunction in simian AIDS

Severe acute respiratory syndrome-coronavirus infection in aged nonhuman primates is associated with modulated pulmonary and systemic immune responses

BACKGROUND: Many respiratory viruses disproportionately impact the elderly. Likewise, advanced age correlated with more adverse disease outcomes following severe acute respiratory syndrome coronavirus (SARS-CoV) infection in humans. We used an aged African green monkey SARS-CoV infection model to better understand age-related mechanisms of increased susceptibility to viral respiratory infections. Nonhuman primates are critical translational models for such research given their similarities to humans in immune-ageing as well as lung structure. RESULTS: Significant age- and infection-dependent differences were observed in both systemic and mucosal immune compartments. Peripheral lymphocytes, specifically CD8 T and B cells were significantly lower in aged monkeys pre- and post- SARS-CoV infection, while neutrophil and monocyte numbers were not impacted by age or infection status. Serum proinflammatory cytokines were similar in both age groups, whereas significantly lower levels of IL-1beta, IL-18, IL-6, IL-12 and IL-15 were detected in the lungs of SARS-CoV-infected aged monkeys at either 5 or 10 days post infection. Total lung leukocyte numbers and relative frequency of CD8 T cells, B cells, macrophages and dendritic cells were greatly reduced in the aged host during SARS-CoV infection, despite high levels of chemoattractants for many of these cells in the aged lung. Dendritic cells and monocytes/macrophages showed age-dependent differences in activation and chemokine receptor profiles, while the CD8 T cell and B cell responses were significantly reduced in the aged host. In examination of viral titers, significantly higher levels of SARS-CoV were detected in the nasal swabs early, at day 1 post infection, in aged as compared to juvenile monkeys, but virus levels were only slightly higher in aged animals by day 3. Although there was a trend of higher titers in respiratory tissues at day 5 post infection, this did not reach statistical significance and virus was cleared from all animals by day 10, regardless of age. CONCLUSIONS: This study provides unique insight into how several parameters of the systemic and mucosal immune response to SARS-CoV infection are significantly modulated by age. These immune differences may contribute to deficient immune function and the observed trend of higher SARS-CoV replication in aged nonhuman primates

Discharged patients and prognoses of caregivers after two years - Part III of the Berlin deinstitutionalisation study

<p>Airway epithelial cells harvested from one-year-old juvenile rhesus monkeys with prior ozone and/or LPS exposure, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090401#pone-0090401-g001" target="_blank">Figure 1</a> were cultured under air-liquid interface conditions and subsequently treated with increasing doses of LPS <i>in vitro</i>. Cultures were evaluated for IL-8 mRNA (A, C) and protein (B, D) expression at 24 h post-treatment. Results show the average +/− SE. *p<0.05, **p<0.01 by two-way ANOVA comparing <i>in vivo</i> exposure and <i>in vitro</i> LPS concentration (n = 4 for each group except filtered air controls n = 5).</p

Effect of postnatal ozone exposure on miR-149, miR-202, miR-410 and miR-let7a expression in juvenile monkey airway epithelial cell cultures.

<p>Comparison of constitutive (0 µg/ml) and LPS-induced (1 µg/ml, 6 h post-treatment) microRNA expression in primary airway epithelial cell cultures derived from one-year-old juvenile rhesus monkeys with prior ozone and/or LPS exposure as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0090401#pone-0090401-g001" target="_blank">Figure 1</a>. (A) miRNA-149, (B) miRNA-202, (C) miRNA-410, (D) miRNA-let7a. Graphs show the average +/− SE of the 2?-(delta CT) where the delta CT =  (microRNA of interest – endogenous control microRNA (RNU-6b)). microRNAs were extracted from n = 3–5 animals per group. Results from Student's t-tests comparing microRNA expression at baseline and post- <i>in vitro</i> LPS treatment are shown with horizontal bars. *p<0.05, **<i>p</i><0.005, ***<i>p</i><0.0003.</p

Assessment of microRNA regulation of IL-6 in human airway epithelial cell cultures.

<p>To evaluate the ability of the putative IL-6-targeting microRNAs to regulate expression of IL-6, the BEAS-2B cell line was transfected with a negative control (NegCO) microRNA or mimics for miR-149, miR-202 or miR-410. (A) Expression of microRNAs in BEAS-2B 24 h post-transfection with a NegCO microRNA or mimics for miR-149, miR-202, or miR-410. Values are reported as the average +/− SE of the 2?-(delta CT) relative to the endogenous control microRNA RNU-6b. (B) IL-6 mRNA and (C) IL-6 protein expression was evaluated in transfected BEAS-2B cells after 24 h. Data are reported as the fold-change in IL-6 mRNA or IL-6 protein as compared to no mimic controls for each experiment. (D) Binding of microRNA mimics to IL-6 mRNA was tested in the HBE1 cell line with the IL-6 3′UTR cloned into a firefly/renilla luciferase plasmid reporter system. For transfection normalization across samples, the relative luciferase units are reported for firefly versus renilla luciferase. Data are from 4 separate experiments. *p<0.05, **p<0.005 by Student's t-test.</p

IL-6 immunofluorescence staining in juvenile monkey trachea following postnatal ozone and/or <i>in vivo</i> LPS.

<p>Trachea cryosections from a representative animal in each exposure group were stained with FITC-conjugated anti-human IL-6 mouse monoclonal antibody (B, D, F, and H). FITC-conjugated mouse IgG1 isotype controls are included for comparison (A, C, E, and G). Images were collected at 20× magnification. Scale bar =  100 µm. White arrows indicate IL-6 staining.</p