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

Attrition of T-cell functions and simultaneous upregulation of inhibitory markers correspond with the waning of BCG-induced protection against tuberculosis in mice.

Mycobacterium bovis bacille Calmette-Guérin (BCG) is the most widely used live attenuated vaccine. However, the correlates of protection and waning of its immunity against tuberculosis is poorly understood. In this study, we correlated the longitudinal changes in the magnitude and functional quality of CD4(+) and CD8(+) T-cell response over a period of two years after mucosal or parenteral BCG vaccination with the strength of protection against Mycobacterium tuberculosis in mice. The BCG vaccination-induced CD4(+) and CD8(+) T cells exhibited comparable response kinetics but distinct functional attributes in-terms of IFN-γ, IL-2 and TNF-α co-production and CD62L memory marker expression. Despite a near life-long BCG persistence and the induction of enduring CD4(+) T-cell responses characterized by IFN-γ and/or TNF-α production with comparable protection, the protective efficacy waned regardless of the route of vaccination. The progressive decline in the multifactorial functional abilities of CD4(+) and CD8(+) T cells in-terms of type-1 cytokine production, proliferation and cytolytic potential corresponded with the waning of protection against M. tuberculosis infection. In addition, simultaneous increase in the dysfunctional and terminally-differentiated T cells expressing CTLA-4, KLRG-1 and IL-10 during the contraction phase of BCG-induced response coincided with the loss of protection. Our results question the empirical development of BCG-booster vaccines and emphasize the pursuit of strategies that maintain superior T-cell functional capacity. Furthermore, our results underscore the importance of understanding the comprehensive functional dynamics of antigen-specific T-cell responses in addition to cytokine polyfunctionality in BCG-vaccinated hosts while optimizing novel vaccination strategies against tuberculosis

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

BCG vaccination induces polyfunctional CD4⁺ but monofunctional CD8⁺ T-cell response.

<p>(A–C) The lung and spleen cells of i.n. and s.c. BCG-vaccinated mice (<i>n</i> = 4/time point/group) were stimulated with WCL, and the magnitudes and polyfunctionality of CD3⁺CD4⁺ and CD3⁺CD8⁺ T cells in-terms of IFN-γ, IL-2 and TNF-α production were determined using polychromatic flow cytometry. (A) The longitudinal changes in the magnitudes of WCL-specific individual IFN-γ, IL-2 or TNF-α-producing cells among CD4⁺ or CD8⁺ T cells in the lung and spleen are plotted. (B) Representative dot plots show the frequency of WCL-specific IFN-γ and IL-2 or IFN-γ and TNF-α-producing cells among splenic CD4⁺ and CD8⁺ T cells from one mouse per vaccinated group at week 32 in comparison with age-matched naïve control, and the distributions of single, double and triple-cytokine-producers are illustrated. (C) The magnitudes of 7 possible combinations of WCL-specific cytokine-producing subsets constituting total cytokine⁺ CD4⁺ or CD8⁺ T cells in the lung, spleen, cervical lymph node (CLN) and inguinal lymph node (ILN) are depicted. The data (A, C) are mean ± s.e.m. responses of 2 (at week 3, 78 and 104), 3 (at week 32) or 4 (at week 6 and 12) independent experiments.</p

BCG-induced T cells predominantly exhibit effector or effector memory phenotype and their proliferative potential wanes with time.

<p>(A–B) The proliferation of CD4⁺ and CD8⁺ T cells from the lung and spleen of the BCG-vaccinated mice was investigated based on CFSE-dilution at 72 h following WCL or no antigen stimulation. (A) Representative plots illustrate the proliferation of pooled lung CD4⁺ and CD8⁺ T cells of vaccinated mice (<i>n</i> = 4) from one experiment at the expansion and contraction phase. (B) The longitudinal changes in the proliferative potential of the lung and spleen CD4⁺ or CD8⁺ T cells from the two vaccinated groups at 4 different time points were summarized and are presented as the fold increase in proliferation compared to unstimulated controls (*<i>P</i><0.05, **<i>P</i><0.01 using 1-way ANOVA with Tukey's post-test compared to corresponding week 12). (C) The CD44 and CD62L expression on the pooled splenic CD4⁺ and CD8⁺ T cells of vaccinated mice (<i>n</i> = 4) from one representative experiment at the expansion and contraction phase are shown. The green dots represent WCL-specific IFN-γ-producing T cells. (D) The magnitudes of WCL-specific IFN-γ-producing CD44⁺CD62L⁺ and CD44⁺CD62L⁻ CD4⁺ and CD8⁺ T cells from the lung and spleen of two groups at week 12 and 52. The data in (B) and (D) are mean ± s.e.m. responses of 2 independent experiments using pooled organ cells (<i>n</i> = 4 mice/time point/group).</p