GM-CSF DNA: An adjuvant for higher avidity IgG, rectal IgA, and increased protection against the acute phase of a SHIV-89.6P challenge by a DNA/MVA immunodeficiency virus vaccine

AbstractSingle intradermal or intramuscular inoculations of GM-CSF DNA with the DNA prime for a simian–human immunodeficiency virus (SHIV)-89.6 vaccine, which consists of DNA priming followed by modified vaccinia Ankara (MVA) boosting, increased protection of both the blood and intestines against the acute phase of an intrarectal SHIV-89.6P challenge. GM-CSF appeared to contribute to protection by enhancing two antibody responses: the avidity maturation of anti-Env IgG in blood (p=<0.01) and the presence of long lasting anti-viral IgA in rectal secretions (p<0.01). The avidity of anti-Env IgG showed strong correlations with protection both pre and post challenge. Animals with the highest avidity anti-Env Ab had 1000-fold reductions in peak viremia over those with the lowest avidity anti-Env Ab. The enhanced IgA response was associated with the best protection, but did not achieve significance

Oral microbiome in HIV-associated periodontitis

M.N-J, J.R, Y.G. and R.P acknowledge funds from the Gala contra la SIDA 2013 and 2014 editions, the Nit per la Recerca a la Catalunya Central 2015 edition and the Red de investigación en SIDA.Supplemental Digital Content is available in the text HIV-associated periodontal diseases (PD) could serve as a source of chronic inflammation. Here, we sought to characterize the oral microbial signatures of HIV+ and HIV- individuals at different levels of PD severity. This cross-sectional study included both HIV+ and HIV- patients with varying degrees of PD. Two tooth, 2 cheek, and 1 saliva samples were obtained for microbiome analysis. Mothur/SILVADB were used to classify sequences. R/Bioconductor (Vegan, PhyloSeq, and DESeq2) was employed to assess overall microbiome structure differences and differential abundance of bacterial genera between groups. Polychromatic flow cytometry was used to assess immune activation in CD4 and CD8 cell populations. Around 250 cheek, tooth, and saliva samples from 50 participants (40 HIV+ and 10 HIV-) were included. Severity of PD was classified clinically as None/Mild (N), Moderate (M), and Severe (S) with 18 (36%), 16 (32%), and 16 (32%) participants in each category, respectively. Globally, ordination analysis demonstrated clustering by anatomic site (R 2 = 0.25, P < 0.001). HIV status and PD severity showed a statistically significant impact on microbiome composition but only accounted for a combined 2% of variation. HIV+ samples were enriched in genera Abiotrophia, Neisseria, Kingella, and unclassified Neisseriaceae and depleted in Leptotrichia and Selenomonas. The Neisseria genus was consistently enriched in HIV+ participants regardless of sampling site and PD level. Immune markers were altered in HIV+ participants but did not show association with the oral microbiome. HIV-associated changes in oral microbiome result in subtle microbial signatures along different stages of PD that are common in independent oral anatomic sites

Expansion and Exhaustion of T-Cell Responses during Mutational Escape from Long-Term Viral Control in Two DNA/Modified Vaccinia Virus Ankara-Vaccinated and Simian-Human Immunodeficiency Virus SHIV-89.6P-Challenged Macaques▿

In this study, we monitored the temporal breadths, frequencies, and functions of antiviral CD4 and CD8 T cells in 2 of 22 DNA/modified vaccinia virus Ankara-vaccinated macaques that lost control of a simian-human immunodeficiency virus 89.6P challenge by 196 weeks postchallenge. Our results show that both mutation and exhaustion contributed to escape. With the reappearance of viremia, responding CD8 and CD4 T cells underwent an initial increase and then loss of breadth and frequency. Antiviral gamma interferon (IFN-γ)- and interleukin 2-coproducing cells were lost before IFN-γ-producing cells and CD4 cells before CD8 cells. At euthanasia, all CD8, but no CD4, Gag epitopes detected during long-term control contained mutations

Presenilin1 regulates Th1 and Th17 effector responses but is not required for experimental autoimmune encephalomyelitis.

Multiple Sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) where pathology is thought to be regulated by autoreactive T cells of the Th1 and Th17 phenotype. In this study we sought to understand the functions of Presenilin 1 (PSEN1) in regulating T cell effector responses in the experimental autoimmune encephalomyelitis (EAE) murine model of MS. PSEN1 is the catalytic subunit of γ-secretase a multimolecular protease that mediates intramembranous proteolysis. γ-secretase is known to regulate several pathways of immune importance. Here we examine the effects of disrupting PSEN1 functions on EAE and T effector differentiation using small molecule inhibitors of γ-secretase (GSI) and T cell-specific conditional knockout mice (PSEN1 cKO). Surprisingly, blocking PSEN1 function by GSI treatment or PSEN1 cKO had little effect on the development or course of MOG35-55-induced EAE. In vivo GSI administration reduced the number of myelin antigen-specific T cells and suppressed Th1 and Th17 differentiation following immunization. In vitro, GSI treatment inhibited Th1 differentiation in neutral but not IL-12 polarizing conditions. Th17 differentiation was also suppressed by the presence of GSI in all conditions and GSI-treated Th17 T cells failed to induce EAE following adoptive transfer. PSEN cKO T cells showed reduced Th1 and Th17 differentiation. We conclude that γ-secretase and PSEN1-dependent signals are involved in T effector responses in vivo and potently regulate T effector differentiation in vitro, however, they are dispensable for EAE

BCG persists and induces a durable cell-mediated immunity in mice.

<p>(A–B) BALB/c mice were vaccinated with 1×10⁶ CFU of BCG Copenhagen by intranasal (i.n.) or subcutaneous (s.c.) route. At 7 different time points after vaccination at the same time, mice were euthanized and their lungs and spleen were isolated. (A) BCG load in the two organs of individual mice was determined. Detection limit of the assay as shown by dotted line was 30 CFU/organ, and data shown are mean CFU ± s.e.m. in the lung (left) and spleen (right) from two independent experiments using 5 individually analyzed mice per group at each time point. The experiment was not performed at the104-week time point. (*<i>P</i><0.05, ** <i>P</i><0.01, *** <i>P</i><0.001, **** <i>P</i><0.0001 using 1-way analysis of variance (ANOVA) with Tukey's post-test). (B) In parallel, pooled cells from the lung (left) or spleen (right) of BCG-vaccinated and naïve mice (<i>n</i> = 4/time point/group) were stimulated with WCL in a cultured ELISPOT assay and IFN-γ, IL-4 and IL-17A SFU were enumerated. Data are means ± s.e.m. of individual cytokine SFU constituting the total cytokine responses of 2 (at week 3, 78 and 104), 3 (at week 32) or 4 (at week 6 and 12) independent experiments evaluated in triplicate. Responses are significant (<i>P</i><0.001) in vaccinated compared to naïve mice at 6 time points after week 3 using 1-way ANOVA with Tukey's post-test.</p

Waning of BCG-induced protection against <i>Mtb</i> coincides with decreased T-cell functional capacity.

<p>(A–C) BCG-vaccinated or age-matched naïve mice were infected with <i>Mtb</i> Erdman, and 6 weeks later the spleen and lungs were isolated. (A) Differences in <i>Mtb</i> load between vaccinated and control mice are shown. The data are mean ± s.e.m. <i>Mtb</i> CFU in each organ at eight different time points involving two to four independent experiments each using five individually analyzed mice per time point. (*<i>P</i><0.05, ** <i>P</i><0.01, *** <i>P</i><0.001, compared to the corresponding age-matched naïve control by Kruskal-Wallis test followed by Dunn's post-test). (B) The longitudinal changes in the frequency of rESAT-6+rCFP-10-specific total CD4⁺ cytokine⁺ (of IFN-γ, IL-2 and TNF-α) T cells from the lung (upper panel) and spleen (lower panel) of vaccinated and naïve mice 6 weeks after challenge (*<i>P</i><0.05, ** <i>P</i><0.01 compared to the corresponding week 32 BCG-vaccinated mice by 1-way ANOVA and Tukey's post-test). At the 32-week time point, rESAT-6+rCFP-10-specific responses in naïve mice are statistically higher compared to the corresponding vaccinated groups (<i>P</i><0.05). (C) The magnitudes of WCL-specific total CD4⁺ cytokine⁺ T cells before and 6 weeks after <i>Mtb</i> challenge. The data are mean ± s.e.m. responses measured by intracellular cytokine staining (ICS) of four mice per time point. (*<i>P</i><0.05 comparing pre- and post-challenge WCL-specific responses using 1-way ANOVA and Tukey's post-test) (D) The schematic summary of longitudinal changes in BCG load in the lung and spleen of i.n. and s.c. BCG-vaccinated mice over a period of 2 years and its correlation with the magnitude and functional capacity of T cells and levels of protection against <i>Mtb</i> infection.</p

T-cell dysfunction markers are up-regulated during waning of BCG responses.

<p>(A) The expression of KLRG-1 on the pooled spleen T cells of age-matched naïve, i.n. or s.c. BCG-vaccinated mice (<i>n</i> = 4) at week 78. (B) Longitudinal changes in the magnitude of KLRG-1⁺ CD4⁺ and CD8⁺ T cells from the lung and spleen. (C) CTLA-4 and PD-1 expression on the pooled splenic CD4⁺ T cells of vaccinated or age-matched naive mice (<i>n</i> = 4) at week 78. (D) Summary of magnitude of CTLA-4⁺ CD4⁺ and CD8⁺ T cells in the lung and spleen. (E) The WCL-specific IL-10 production by the pooled splenic CD4⁺ T cells of vaccinated or age-matched naïve mice. (F) Summary of magnitude of WCL-specific IL-10⁺ CD4⁺ and CD8⁺ T cells in the lung and spleen. (G–H) The spleen and lung cells isolated from i.n. BCG-vaccinated mice at week 78 were treated with rIL-2 to evaluate potential to reinvigorate WCL-specific CD4⁺ and CD8⁺ T-cell responses. (G) The WCL-specific IFN-γ, IL-4 and IL-17 SFU after treatment with or without rIL-2. (H) The magnitude of WCL-specific cytokine-producing 7 subsets of CD4⁺ and CD8⁺ T cells by flow cytometry. The data in figure B, D, F are mean ± s.e.m. responses of 2 independent experiments using pooled organ cells (<i>n</i> = 4 mice/time point/group). * <i>P</i><0.05, ** <i>P</i><0.01 compared to the corresponding 12-week time point by 1-way ANOVA with Tukey's post-test. 5G, H are mean ± s.e.m responses of pooled cells evaluated in triplicates. *Significant using 1-way ANOVA with Tukey's post-test.</p