Seminal plasma exposures strengthen vaccine responses in the female reproductive tract mucosae
ABSTRACT: HIV-1 sexual transmission occurs mainly via mucosal semen exposures. In the female reproductive tract (FRT), seminal plasma (SP) induces physiological modifications, including inflammation. An effective HIV-1 vaccine should elicit mucosal immunity. However, modifications of vaccine responses by the local environment remain to be characterized. Using a modified vaccinia virus Ankara (MVA) as a vaccine model, we characterized the impact of HIV-1+ SP intravaginal exposure on the local immune responses of nonhuman primates. Multiple HIV-1+ SP exposures did not impact the anti-MVA antibody responses. However, SP exposures revealed an anti-MVA responses mediated by CD4+ T cells, which was not observed in the control group. Multi-parameter approaches clearly identified the cervix as the most impacted compartment in the FRT. The frequency and the quality of specific anti-MVA CD8+ T cell responses increased in the FRT exposed to SP. Furthermore SP exposures induced a local cell recruitment of antigen presenting cells, especially CD11c+ cells and CD8+ T cell recruitment in the FRT draining lymph nodes. CD11c+ cell recruitment was associated with upregulation of inflammation-related gene expression after SP exposures in the cervix. We thus highlight the fact that physiological conditions, such as SP exposures, should be taken into consideration to test and to improve vaccine efficacy against HIV-1 and other sexually transmitted infections.
Project description:The female reproductive tract is one of the major mucosal invasion site of HIV-1. This site has been neglected in previous HIV-1 vaccine studies. Immune responses in the female reproductive tract after systemic vaccination remain to be characterized. Using a modified vaccinia virus Ankara (MVA) as a vaccine model, we characterized specific immune responses in all compartments of the female reproductive tract (FRT) of non-human primates after systemic vaccination. Memory T cells were preferentially found in the lower tract (vagina and cervix), whereas antigen-presenting cells and innate lymphoid cells were mainly located in the upper tract (uterus and fallopian tubes). This compartmentalisation of immune cells in the FRT was supported by transcriptomic analyses and correlation network. Polyfunctional MVA-specific CD8+ T cells were detected in the blood, lymph nodes, vagina, cervix, uterus and fallopian tubes. Anti-MVA IgG and IgA were detected in cervicovaginal fluid after a second vaccine dose. Systemic vaccination with an MVA vector thus elicits cellular and antibody responses in the female reproductive tract.
Project description:Modified vaccinia virus Ankara (MVA) has been explored as a vaccine vector for use against infectious diseases and cancer. MVA is an immunogenic, attenuated poxvirus capable of eliciting robust cellular and humoral responses in pre-clinical animal models and in patients. However, upon infection with MVA, cells undergo rapid apoptosis leading to faster clearance of recombinant antigens. The fragmentation of the anti-apoptotic gene B13R in MVA could contribute to this effect. Here, we replaced the fragmented B13R with a functional version and observed that MVA-B13R infected HeLa cells and muscle cell lines delayed caspase 3 activation compared to MVA indicating slower progression of apoptosis. For immunogenicity studies, mice were intramuscularly immunized with recombinant MVA or MVA-B13R expressing SIV Gag, Pol and HIV Env (SHIV). We observed higher Env-specific humoral responses from MVA-B13R SHIV compared to MVA SHIV mice. To determine differences in the innate immune response that may have contributed to the augmented humoral response, we performed RNA-Seq analysis on draining lymph node cells after immunization. Gene set enrichment analysis from day 1 after immunization showed that MVA-B13R SHIV immunizations were associated with a negative enrichment for type I and II interferon responses compared to MVA SHIV mice indicating MVA-B13R SHIV induces a delayed anti-viral interferon response that may lead to the enhanced humoral response observed. Taken together, these results demonstrate that restoring B13R functionality in MVA significantly delays MVA-induced apoptosis, augments Env-specific antibody responses, and is associated with reduced interferon-alpha and interferon-gamma responses induced after vaccination. Overall design: Female BALB/c mice were intramuscularly immunized at 10^7 pfu/dose with MVA expressing SHIV antigens or MVA-B13R expressing SHIV. At days 1, 2, and 6 post immunization, draining inguinal lymph node cells were harvested for RNA extraction. 5 animals per group per time point including 5 naive mice as a control group.
Project description:A better understanding of innate responses induced by vaccination is critical for designing optimal vaccines. Here, we studied the diversity and dynamics of the NK cell compartment after prime-boost immunization with the modified vaccinia virus Ankara using cynomolgus macaques as a model. Mass cytometry was used to deeply characterize blood NK cells. The NK cell subphenotype composition was modified by the prime. Certain phenotypic changes induced by the prime were maintained over time and, as a result, the NK cell composition prior to boost differed from that before prime. The key phenotypic signature that distinguished NK cells responding to the boost from those responding to the prime included stronger expression of several cytotoxic, homing, and adhesion molecules, suggesting that NK cells at recall were functionally distinct. Our data reveal potential priming or imprinting of NK cells after the first vaccine injection. This study provides novel insights into prime-boost vaccination protocols that could be used to optimize future vaccines.
Project description:Successful development of HIV-vaccination strategies will also depend on the ability to use novel approaches to analyse and integrate immunogenicity data generated in vaccine trials. The ANRS VAC 18 trial evaluated the immunogenicity of HIV-LIPO-5 vaccine (5 HIV peptides coupled to a palmytoil tail) administered at W0, 4, 12 and 24 in healthy volunteers. 62-69% of vaccinees developed HIV-specific ELISpot responses by W26. Here we present extensive immunogenicity assessments in a subset of vaccinees using ELISpot, lymphoproliferation, intracellular cytokine staining (ICS), cytokine multiplex and transcriptomic analyses. Peripheral blood mononuclear cells from volunteers collected before and following vaccinations were stimulated with HIV LIPO 5 vaccine, Gag peptides contained or not in the vaccine as controls. Different time points and stimulation conditions were compared, using false discovery rate to control for test multiplicity. 74% and 30% of vaccinees had cultured ELISpot and lymphoproliferation responses at W14, respectively. Ex-vivo ICS showed mainly single IL-2 producing cells. Secretion of IFN-γ, TNF-α, IL-5, and IL-13 increased significantly in response to Gag stimulation after culture at W14 compared to W0. An induction of metallothionein genes was consistently detected after HIV-LIPO-5 stimulation at W0 and W14 related to the adjuvant effect of the lipid tail. After vaccination (W14), significant probes increased substantially (>1200 probes) including IFN-γ, CXCL9, IL2RA, TNFAIP6, CCL3L1 and IL-6 W14 (fold change > 100%). In conclusion, HIV LIPO-5 vaccination elicited memory precursor responses with a Th1 and Th2 profile. The signature profile before vaccination provides information about the adjuvant effect of the lipid tail. Consistently with cytokine responses, vaccination is associated with a modulation in gene expression. This combined approach allowed to identify new signatures of HIV vaccine response and indicates that HIV-LIPO-5 could be further developed as a prime component of heterologous prime boost strategies. PBMC mRNA of 12 healthy volunteers, stimulate in four different conditions (HIV-LIPO-5, Gag+, Gag-, NS) during 6 and 24 hours before and after vaccination (week 0 and week 14)
Project description:Results from the Step trial in humans and studies in non-human primates highlighted a role for heightened activated CD4 T cell response in promoting HIV/SIV acquisition. However, the contribution of vaccine-specific CD4 T cell response in influencing protection is not known. Here, using the macaque model, we show that vaccine-induced Th1-biased CCR5+ CD4 T cell response in blood and mucosal tissue above a certain thresh¬old is detrimental for vaccine-mediated protection against pathogenic mucosal SIV infections. Overall design: 10 vaccinated animals that acquired SIV infection within the first two challenges (early SIV acquirers) and 12 vaccinated animals that acquired SIV infection after 6 challenges (late SIV acquirers). We performed microarray analysis on whole blood samples for both set of animals at multiple days after the 1st DNA vaccine (Wk0), 1st MVA boost (Wk16), 2nd MVA boost (Wk32) and 1st rectal challenge.
Project description:New vaccine design approaches would be greatly facilitated by a better understanding of the early systemic changes, and those that occur at the site of injection, responsible for the installation of a durable and oriented protective response. We performed a detailed characterization of very early infection and host response events following the intradermal administration of the modified vaccinia virus Ankara attenuated vaccine in non-human primates. Integrated analysis of the data obtained from in vivo imaging, histology, flow cytometry, multiplex cytokine, and transcriptomic analysis using tools derived from systems biology, such as correlation networks, showed a strong early local and systemic inflammatory response that peaked at 24 h, which was then progressively replaced by an adaptive response during the installation of the host response to the vaccine. Such comprehensive approaches should improve our understanding of how to effectively orientate the immune response, and could contribute to rational vaccine development.
Project description:Following exposure to vaccines, antigen-specific CD8+ T-cell responses develop as long-term memory pools. Novel vaccine strategies based on adenoviral vectors, e.g. those developed for HCV, are able to induce and sustain substantial CD8+ T-cell populations. How such populations evolve following vaccination remains to be defined at a transcriptional level. We addressed the transcriptional regulation of divergent CD8+ T-cell memory pools induced by an adenoviral vector encoding a model antigen (beta-galactosidase). We observe transcriptional profiles that mimic those following infection with persistent pathogens, murine and human cytomegalovirus (CMV). Key transcriptional hallmarks include up-regulation of homing receptors, and anti-apoptotic pathways, driven by conserved networks of transcription factors, including T-bet (TBX21). In humans, a novel adenovirus vaccine induced similar CMV-like phenotypes and underlying transcription factor regulation. These data clarify the core features of CD8+ T-cell memory following vaccination with adenovirus vectors and indicate a conserved pathway for memory development shared with persistent herpesviruses. Total RNA was extracted from sorted memory CD8 T cells induce by CMV and adenoviral vectors, and naïve CD8 cells
Project description:Aims. Serial invasive endomyocardial biopsies (EMB) remain the gold standard for acute cellular rejection (ACR) diagnosis. However histological grading has several limitations. We aimed to explore the value of myocardial Gene Expression Profiling (GEP) for diagnosing and identifying predictive biomarkers of ACR. Methods. A case-control study nested within a retrospective heart transplant patients cohort included 126 patients with median (IQR) age 50 (41-57) years and 111 (88%) males. Among 1157 EMB performed, 467 were eligible (i.e, corresponding to either ISHLT grade 0 or ≥3A), among which 36 were selected for GEP according to the grading: 0 (CISHLT, n= 13); rejection ≥3A (RISHLT, n=13); 0 one month before ACR (BRISHLT, n=10). Results. We found 294 genes differentially expressed between CISHLT and RISHLT, mainly involved in immune activation, and inflammation. Hierarchical clustering showed a clear segregation of CISHLT and RISHLT groups and heterogeneity of GEP within RISHLT. All EMB presented immune activation, but some RISHLT EMB were strongly subject to inflammation, whereas others, closer to CISHLT, were characterized by structural modifications with lower inflammation level. We identified 15 probes significantly different between BRISHLT and CISHLT, including the gene of the muscular protein TTN. This result suggests that structural alterations precede inflammation in ACR. Linear Discriminant Analysis based on these 15 probes was able to identify the histological status of every 36 samples. Conclusion. Myocardial GEP is a helpful method to accurately diagnose ACR, and predicts rejection one month before its histological occurrence. These results should be considered in cardiac allograft recipients’ care.
Project description:Background: Early prognostication is a major challenge after out-of-hospital cardiac arrest (OHCA). Aims: We hypothesized that a genome-wide analysis of blood gene expression could offer new prognostic tools and lines of research. Methods: Sixty-nine patients were enrolled from an ancillary study of the clinical trial NCT00999583 that tested the effect of erythropoietin (EPO) after OHCA. Blood samples were collected in comatose survivors of OHCA at hospital admission and 1 and 3 days after resuscitation. Gene expression profiles were analyzed (Illumina HumanHT-12 V4 BeadChip; >34,000 genes). Patients were classified into two categories representing neurologically favorable outcome (Cerebral Performance Category [CPC] = 1-2) vs unfavorable outcome (CPC>2) at Day 60 after OHCA. Differential and functional enrichment analyses were performed to compare transcriptomic profiles between these two categories. Results: Among the 69 enrolled patients, 33 and 36 patients were treated or not by EPO, respectively. Among them, 42% had a favorable neurological outcome in both groups. EPO did not affect the transcriptomic response at Day-0 and 1 after OHCA. In contrast, 76 transcripts differed at Day-0 between patients with unfavorable vs favorable neurological outcome. This signature persisted at Day-1 after OHCA. Functional enrichment analysis revealed a down-regulation of adaptive immunity with concomitant up-regulation of innate immunity and inflammation in patients with unfavorable vs favorable neurological outcome. The transcription of many genes of the HLA family was decreased in patients with unfavorable vs favorable neurological outcome. Concomitantly, neutrophil activation and inflammation were observed. Up-stream regulators analysis showed the implication of numerous factors involved in cell cycle and damages. A logistic regression including a set of genes allowed a reliable prediction of the clinical outcomes (Specificity=88%; Hit Rate=83%). Conclusions: A transcriptomic signature involving a counterbalance between adaptive and innate immune responses is able to predict neurological outcome very early after hospital admission after OHCA. This deserves confirmation in a larger population.
Project description:This SuperSeries is composed of the following subset Series: GSE22768: Systems analysis of the Merck Ad5/HIV vaccine reveals robust induction of a core innate immune gene network: in vivo analysis GSE22769: Systems analysis of the Merck Ad5/HIV vaccine reveals robust induction of a core innate immune gene network: in vitro analysis To better understand how innate immune responses to vaccination can lead to lasting protective immunity, we used a systems approach to define immune signatures in humans over 1 wk following MRKAd5/HIV vaccination that predicted subsequent HIV-specific T-cell responses. Within 24 h, striking increases in peripheral blood mononuclear cell gene expression associated with inflammation, IFN response, and myeloid cell trafficking occurred, and lymphocyte-specific transcripts decreased. These alterations were corroborated by marked serum inflammatory cytokine elevations and egress of circulating lymphocytes. Responses of vaccinees with preexisting adenovirus serotype 5 (Ad5) neutralizing antibodies were strongly attenuated, suggesting that enhanced HIV acquisition in Ad5-seropositive subgroups in the Step Study may relate to the lack of appropriate innate activation rather than to increased systemic immune activation. Importantly, patterns of chemoattractant cytokine responses at 24 h and alterations in 209 peripheral blood mononuclear cell transcripts at 72 h were predictive of subsequent induction and magnitude of HIV-specific CD8(+) T-cell responses. This systems approach provides a framework to compare innate responses induced by vectors, as shown here by contrasting the more rapid, robust response to MRKAd5/HIV with that to yellow fever vaccine. When applied iteratively, the findings may permit selection of HIV vaccine candidates eliciting innate immune response profiles more likely to drive HIV protective immunity. Refer to individual Series