IL-7 induces sCD127 release and mCD127 downregulation in human CD8+ T cells by distinct yet overlapping mechanisms, both of which are impaired in HIV infection.

The IL-7 receptor specific α chain, CD127, can be expressed both as a membrane-associated (mCD127) and a soluble form (sCD127), however, the mechanisms involved in their regulation remain to be defined. We first demonstrated in primary human CD8+ T cells that IL-7-induced downregulation of mCD127 expression is dependent on JAK and PI3K signaling, whereas IL-7-induced sCD127 release is also mediated by STAT5. Following stimulation with IL-7, expression of alternatively spliced variants of the CD127 gene, sCD127 mRNA, is reduced, but to a lesser degree than the full-length gene. Evaluation of the role of proteases revealed that MMP-9 was involved in sCD127 release, without affecting the expression of mCD127, suggesting it does not induce direct shedding from the cell surface. Since defects in the IL-7/CD127 pathway occur in various diseases, including HIV, we evaluated CD8+ T cells derived from HAART-treated HIV-infected individuals and found that IL-7-induced (1) downregulation of mCD127, (2) release of sCD127, and (3) expression of the sCD127 mRNA were all impaired. Expression of mCD127 and sCD127 is, therefore, regulated by distinct, but overlapping, mechanisms and their impairment in HIV infection contributes to our understanding of the CD8+ T cell dysfunction that persists despite effective antiretroviral therapy.

A Balanced Proinflammatory and Regulatory Cytokine Signature in Young African Children Is Associated With Lower Risk of Clinical Malaria.

BACKGROUND: The effect of timing of exposure to first Plasmodium falciparum infections during early childhood on the induction of innate and adaptive cytokine responses and their contribution to the development of clinical malaria immunity is not well established. METHODS: As part of a double-blind, randomized, placebo-controlled trial in Mozambique using monthly chemoprophylaxis with sulfadoxine-pyrimethamine plus artesunate to selectively control timing of malaria exposure during infancy, peripheral blood mononuclear cells collected from participants at age 2.5, 5.5, 10.5, 15, and 24 months were stimulated ex vivo with parasite schizont and erythrocyte lysates. Cytokine messenger RNA expressed in cell pellets and proteins secreted in supernatants were quantified by reverse-transcription quantitative polymerase chain reaction and multiplex flow cytometry, respectively. Children were followed up for clinical malaria from birth until 4 years of age. RESULTS: Higher proinflammatory (interleukin [IL] 1, IL-6, tumor necrosis factor) and regulatory (IL-10) cytokine concentrations during the second year of life were associated with reduced incidence of clinical malaria up to 4 years of age, adjusting by chemoprophylaxis and prior malaria exposure. Significantly lower concentrations of antigen-specific T-helper 1 (IL-2, IL-12, interferon-γ) and T-helper 2 (IL-4, IL-5) cytokines by 2 years of age were measured in children undergoing chemoprophylaxis compared to children receiving placebo (P < .03). CONCLUSIONS: Selective chemoprophylaxis altering early natural exposure to malaria blood stage antigens during infancy had a significant effect on T-helper lymphocyte cytokine production >1 year later. Importantly, a balanced proinflammatory and anti-inflammatory cytokine signature, probably by innate cells, around age 2 years was associated with protective clinical immunity during childhood. CLINICAL TRIALS REGISTRATION: NCT00231452.

High production of pro-inflammatory cytokines by maternal blood mononuclear cells is associated with reduced maternal malaria but increased cord blood infection.

BACKGROUND: Increased susceptibility to malaria during pregnancy is not completely understood. Cellular immune responses mediate both pathology and immunity but the effector responses involved in these processes have not been fully characterized. Maternal and fetal cytokine and chemokine responses to malaria at delivery, and their association with pregnancy and childhood outcomes, were investigated in 174 samples from a mother and child cohort from Mozambique. Peripheral and cord mononuclear cells were stimulated with Plasmodium falciparum lysate and secretion of IL-12p70, IFN-γ, IL-2, IL-10, IL-8, IL-6, IL-4, IL-5, IL-1ß, TNF, TNF-ß was quantified in culture supernatants by multiplex flow cytometry while cellular mRNA expression of IFN-γ, TNF, IL-2, IL-4, IL-6, IL-10 and IL-13 was measured by quantitative PCR. RESULTS: Higher concentrations of IL-6 and IL-1ß were associated with a reduced risk of P. falciparum infection in pregnant women (p < 0.049). Pro-inflammatory cytokines IL-6, IL-1ß and TNF strongly correlated among themselves (ρ > 0.5, p < 0.001). Higher production of IL-1ß was significantly associated with congenital malaria (p < 0.046) and excessive TNF was associated with peripheral infection and placental lesions (p < 0.044). CONCLUSIONS: Complex network of immuno-pathological cytokine mechanisms in the placental and utero environments showed a potential trade-off between positive and negative effects on mother and newborn susceptibility to infection.

Coadministration of seasonal influenza vaccine and MVA-NP+M1 simultaneously achieves potent humoral and cell-mediated responses.

Current seasonal influenza vaccines have reduced immunogenicity and are of suboptimal efficacy in older adults. We have previously shown that the novel candidate vaccine MVA-NP+M1 is able to boost memory T cell responses in adults aged 50-85 years. Preclinical studies have demonstrated that viral vectored vaccines can act as adjuvants when coadministered with protein-based vaccines. We have conducted a phase I clinical trial to compare the coadministration of seasonal influenza vaccine and MVA-NP+M1 with seasonal influenza vaccine alone in adults aged 50 years and above. This combination of vaccines was safe and well tolerated. T cell responses to internal influenza proteins were boosted to significantly higher levels in the group receiving MVA-NP+M1 compared with the group receiving seasonal influenza vaccine alone. Rates of seroprotection and seroconversion against the three vaccine strains were similar in both groups; however, there was a significant increase in the geometric mean titer ratio for the H3N2 component of seasonal influenza vaccine in the coadministration group. While some vaccine combinations result in immune interference, the coadministration of MVA-NP+M1 alongside seasonal influenza vaccine is shown here to increase some influenza strain-specific antibody responses and boost memory T cells capable of recognizing a range of influenza A subtypes.

Clinical assessment of a novel recombinant simian adenovirus ChAdOx1 as a vectored vaccine expressing conserved Influenza A antigens.

Adenoviruses are potent vectors for inducing and boosting cellular immunity to encoded recombinant antigens. However, the widespread seroprevalence of neutralizing antibodies to common human adenovirus serotypes limits their use. Simian adenoviruses do not suffer from the same drawbacks. We have constructed a replication-deficient chimpanzee adenovirus-vectored vaccine expressing the conserved influenza antigens, nucleoprotein (NP), and matrix protein 1 (M1). Here, we report safety and T-cell immunogenicity following vaccination with this novel recombinant simian adenovirus, ChAdOx1 NP+M1, in a first in human dose-escalation study using a 3+3 study design, followed by boosting with modified vaccinia virus Ankara expressing the same antigens in some volunteers. We demonstrate ChAdOx1 NP+M1 to be safe and immunogenic. ChAdOx1 is a promising vaccine vector that could be used to deliver vaccine antigens where strong cellular immune responses are required for protection.

Comparative analysis of human gut- and blood-derived mononuclear cells: contrasts in function and phenotype.

Introduction: Alterations in the gut immune system have been implicated in various diseases.The challenge of obtaining gut tissues from healthy individuals, commonly performed via surgical explants, has limited the number of studies describing the phenotype and function of gut-derived immune cells in health. Methods: Here, by means of recto-sigmoid colon biopsies obtained during routine care (colon cancer screening in healthy adults), the phenotype and function of immune cells present in the gut were described and compared to those found in blood. Results: The proportion of CD4+, CD8+, MAIT, γδ+ T, and NK cells phenotype, expression of integrins, and ability to produce cytokine in response to stimulation with PMA and ionomycin. T cells in the gut were found to predominantly have a memory phenotype as compared to T cells in blood where a naïve phenotype predominates. Recto-sigmoid mononuclear cells also had higher PD-1 and Ki67 expression. Furthermore, integrin expression and cytokine production varied by cell type and location in blood vs. gut. Discussion: These findings demonstrate the differences in functionality of these cells when compared to their blood counterparts and validate previous studies on phenotype within gut-derived immune cells in humans (where cells have been obtained through surgical means). This study suggests that recto-sigmoid biopsies collected during colonoscopy can be a reliable yet more accessible sampling method for follow up of alterations of gut derived immune cells in clinical settings.

Interleukin-10 (IL-10) polymorphisms are associated with IL-10 production and clinical malaria in young children.

The role of interleukin-10 (IL-10) in malaria remains poorly characterized. The aims of this study were to investigate (i) whether genetic variants of the IL-10 gene influence IL-10 production and (ii) whether IL-10 production as well as the genotypes and haplotypes of the IL-10 gene in young children and their mothers are associated with the incidence of clinical malaria in young children. We genotyped three IL-10 single nucleotide polymorphisms in 240 children and their mothers from a longitudinal prospective cohort and assessed the IL-10 production by maternal peripheral blood mononuclear cells (PBMCs) and cord blood mononuclear cells (CBMCs). Clinical episodes of Plasmodium falciparum malaria in the children were documented until the second year of life. The polymorphism IL-10 A-1082G (GCC haplotype of three SNPs in IL-10) in children was associated with IL-10 production levels by CBMC cultured with P. falciparum-infected erythrocytes (P = 0.043), with the G allele linked to low IL-10 production capacity. The G allele in children was also significantly associated with a decreased risk for clinical malaria infection in their second year of life (P = 0.016). Furthermore, IL-10 levels measured in maternal PBMCs cultured with infected erythrocytes were associated with increased risk of malaria infection in young children (P < 0.001). In conclusion, IL-10 polymorphisms and IL-10 production capacity were associated with clinical malaria infections in young children. High IL-10 production capacity inherited from parents may diminish immunological protection against P. falciparum infection, thereby being a risk for increased malaria morbidity.

Preliminary assessment of the efficacy of a T-cell-based influenza vaccine, MVA-NP+M1, in humans.

BACKGROUND: The novel influenza vaccine MVA-NP+M1 is designed to boost cross-reactive T-cell responses to internal antigens of the influenza A virus that are conserved across all subtypes, providing protection against both influenza disease and virus shedding against all influenza A viruses. Following a phase 1 clinical study that demonstrated vaccine safety and immunogenicity, a phase 2a vaccination and influenza challenge study has been conducted in healthy adult volunteers. METHODS: Volunteers with no measurable serum antibodies to influenza A/Wisconsin/67/2005 received either a single vaccination with MVA-NP+M1 or no vaccination. T-cell responses to the vaccine antigens were measured at enrollment and again prior to virus challenge. All volunteers underwent intranasal administration of influenza A/Wisconsin/67/2005 while in a quarantine unit and were monitored for symptoms of influenza disease and virus shedding. RESULTS: Volunteers had a significantly increased T-cell response to the vaccine antigens following a single dose of the vaccine, with an increase in cytolytic effector molecules. Intranasal influenza challenge was undertaken without safety issues. Two of 11 vaccinees and 5 of 11 control subjects developed laboratory-confirmed influenza (symptoms plus virus shedding). Symptoms of influenza were less pronounced in the vaccinees and there was a significant reduction in the number of days of virus shedding in those vaccinees who developed influenza (mean, 1.09 days in controls, 0.45 days in vaccinees, P = .036). CONCLUSIONS: This study provides the first demonstration of clinical efficacy of a T-cell-based influenza vaccine and indicates that further clinical development should be undertaken. CLINICAL TRIALS REGISTRATION: NCT00993083.

Potent CD8+ T-cell immunogenicity in humans of a novel heterosubtypic influenza A vaccine, MVA-NP+M1.

BACKGROUND: Influenza A viruses cause occasional pandemics and frequent epidemics. Licensed influenza vaccines that induce high antibody titers to the highly polymorphic viral surface antigen hemagglutinin must be re-formulated and readministered annually. A vaccine providing protective immunity to the highly conserved internal antigens could provide longer-lasting protection against multiple influenza subtypes. METHODS: We prepared a Modified Vaccinia virus Ankara (MVA) vector encoding nucleoprotein and matrix protein 1 (MVA-NP+M1) and conducted a phase I clinical trial in healthy adults. RESULTS: The vaccine was generally safe and well tolerated, with significantly fewer local side effects after intramuscular rather than intradermal administration. Systemic side effects increased at the higher dose in both frequency and severity, with 5 out of 8 volunteers experiencing severe nausea/vomiting, malaise, or rigors. Ex vivo T-cell responses to NP and M1 measured by IFN-γ ELISPOT assay were significantly increased after vaccination (prevaccination median of 123 spot-forming units/million peripheral blood mononuclear cells, postvaccination peak response median 339, 443, and 1443 in low-dose intradermal, low-dose intramuscular, and high-dose intramuscular groups, respectively), and the majority of the antigen-specific T cells were CD8(+). CONCLUSIONS: We conclude that the vaccine was both safe and remarkably immunogenic, leading to frequencies of responding T cells that appear to be much higher than those induced by any other influenza vaccination approach. Further studies will be required to find the optimum dose and to assess whether the increased T-cell response to conserved influenza proteins results in protection from influenza disease.

Comparison of commercial kits to measure cytokine responses to Plasmodium falciparum by multiplex microsphere suspension array technology.

BACKGROUND: Multiplex cytokine profiling systems are useful tools for investigating correlates of protective immunity. Several Luminex and flow cytometry methods are commercially available but there is limited information on the relative performance of different kits. A series of comparison experiments were carried out to determine the most appropriate method for our subsequent studies. METHODS: Two Luminex methods were compared, the Bio-Rad human 17-plex panel and the Invitrogen (formerly BioSource) human cytokine 10-plex kit, and two flow cytometry methods, the Becton Dickinson Human Th1/Th2 Cytokine Kit (CBA) and the Bender MedSystems Human Th1/Th2 11plex FlowCytomix Multiplex Kit. All kits were tested for the measurement of cytokines in supernatants collected from human leukocytes stimulated with viable Plasmodium falciparum infected red blood cells (iRBC) or P. falciparum schizont lysates. RESULTS: Data indicated that the kits differed in sensitivity and reproducibility depending on the cytokine, and detected different quantities of some cytokines. The Bio-Rad 17-plex kit was able to detect more positive responses than the Invitrogen 10-plex kit. However, only when detecting IL-1, IL-6 or TNF did the two Luminex based methods correlate with one another. In this study, the flow cytometry based techniques were less variable and correlated better with one another. The two flow cytometry based kits showed significant correlation when detecting IFN-γ, IL-2, TNF, IL-10 and IL-6, but overall the BD kit detected more positive responses than the Bender MedSystems kit. CONCLUSIONS: The microsphere suspension array technologies tested differed in reproducibility and the absolute quantity of cytokine detected. Sample volume, the number of cytokines measured, and the time and cost of the assays also differed. These data provide an accurate assessment of the four techniques, which will allow individual researchers to select the tool most suited for their study population.

Soluble IL-7Rα/sCD127 in Health, Disease, and Its Potential Role as a Therapeutic Agent.

Soluble cytokine receptors can influence immune responses by modulating the biological functions of their respective ligands. These effects can be either agonistic or antagonistic and a number of soluble cytokine receptors have been shown to play critical roles in both maintenance of health and disease pathogenesis. Soluble IL-7Ra (sCD127) is one such example. With its impact on the IL-7/CD127 pathway, which is fundamental for the development and homeostasis of T cells, the role of sCD127 in health and disease has been extensively studied in recent years. Within this review, the role of sCD127 in maintaining host immune function is presented. Next, by addressing genetic factors affecting sCD127 expression and the associated levels of sCD127 production, the roles of sCD127 in autoimmune disease, infections and cancer are described. Finally, advances in the field of soluble cytokine therapy and the potential for sCD127 as a biomarker and therapeutic agent are discussed.

An enveloped virus-like particle vaccine expressing a stabilized prefusion form of the SARS-CoV-2 spike protein elicits highly potent immunity.

We evaluated enveloped virus-like particles (eVLPs) expressing various forms of the Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) spike protein and several adjuvants in an effort to identify a highly potent Coronavirus disease 2019 (COVID-19) vaccine candidate. eVLPs expressing a modified prefusion form of SARS-CoV-2 spike protein were selected as they induced high antibody binding titers and neutralizing activity after a single injection in mice. Formulation of SARS-CoV-2 S eVLPs with aluminum phosphate resulted in balanced induction of IgG2 and IgG1 isotypes and antibody binding and neutralization titers were undiminished for more than 3 months after a single immunization. A single dose of this candidate, named VBI-2902a, protected Syrian golden hamsters from challenge with SARS-CoV-2 and supports the on-going clinical evaluation of VBI-2902a as a highly potent vaccine against COVID-19.

Prime-boost immunization with adenoviral and modified vaccinia virus Ankara vectors enhances the durability and polyfunctionality of protective malaria CD8+ T-cell responses.

Protection against liver-stage malaria relies on the induction of high frequencies of antigen-specific CD8+ T cells. We have previously reported high protective levels against mouse malaria, albeit short-lived, by a single vaccination with adenoviral vectors coding for a liver-stage antigen (ME.TRAP). Here, we report that prime-boost regimens using modified vaccinia virus Ankara (MVA) and adenoviral vectors encoding ME.TRAP can enhance both short- and long-term sterile protection against malaria. Protection persisted for at least 6 months when simian adenoviruses AdCh63 and AdC9 were used as priming vectors. Kinetic analysis showed that the MVA boost made the adenoviral-primed T cells markedly more polyfunctional, with the number of gamma interferon (INF-gamma), tumor necrosis factor alpha (TNF-alpha), and interleukin-2 (IL-2) triple-positive and INF-gamma and TNF-alpha double-positive cells increasing over time, while INF-gamma single-positive cells declined with time. However, IFN-gamma production prevailed as the main immune correlate of protection, while neither an increase of polyfunctionality nor a high integrated mean fluorescence intensity (iMFI) correlated with protection. These data highlight the ability of optimized viral vector prime-boost regimens to generate more protective and sustained CD8+ T-cell responses, and our results encourage a more nuanced assessment of the importance of inducing polyfunctional CD8(+) T cells by vaccination.

The relationship between human effector and memory T cells measured by ex vivo and cultured ELISPOT following recent and distal priming.

Maintenance of T-cell responses is an essential feature in protection from many infectious diseases that must be harnessed in vaccination. The relationship between effector T-cell responses and more durable and highly proliferative T-cell memory, particularly in humans, is not well understood. In this study, effector T-cell responses were measured by overnight ex vivo interferon-gamma (IFN-gamma) enzyme-linked immunosorbent spot-forming cell assay (ELISPOT), whereas memory T cells were measured by 10-day culture followed by IFN-gamma ELISPOT (cultured ELISPOT). We observed a significant correlation between IFN-gamma responses to CD4-stimulatory, but not to CD8-stimulatory, recall antigens measured by these assays, suggesting a divergence in regulation. In vaccine trial participants who received a prime-boost vaccination regimen comprising malaria antigens delivered by poxviruses, there was a correlation between ex vivo and cultured responses on day 7, but not 3 months post-vaccination, with the ratio of cultured : ex vivo response increasing over time. To compare responses revealed by cultured ELISPOT in more detail, tetramers comprising viral recall antigens were used to ascribe effector-memory and central-memory T-cell phenotypes through CCR7 and CD62L costaining. For CD8(+) responses the effector phenotype decreased during the initial culture period and memory populations remained high within the resulting 20-fold to 50-fold increased IFN-gamma-secreting or tetramer(+) population. This was less marked for CD4(+) responses, which had higher starting memory phenotype. Depletion of these central-memory T-cell populations generally ablated responses in cultured ELISPOT and reduced ex vivo responses. This study highlights differences between CD4(+) and CD8(+) effector and memory T cells, and the more complex phenotype of CD4(+) T cells.

MIG (CXCL9) is a more sensitive measure than IFN-gamma of vaccine induced T-cell responses in volunteers receiving investigated malaria vaccines.

For many years the IFN-gamma ex vivo ELISPOT has been a major assay for assessing human T-cell responses generated by malaria vaccines. The ELISPOT assay is a sensitive assay, but an imperfect correlate of protection against malaria. Monokine induced by gamma (MIG), or CXCL9, is a chemokine induced by IFN-gamma and has the potential to provide amplification of the IFN-gamma signal. MIG secretion could provide a measure of bio-active IFN-gamma and a functional IFN-gamma signalling pathway. We report that detecting MIG by flow cytometry and by RT-PCR can be more sensitive than the detection of IFN-gamma using these methods. We also find that there is little inter-individual variability in MIG secretion when detected by flow cytometry and that the MIG assay may be used to estimate the amount of bio-active IFN-gamma present. Measurement of MIG alongside IFN-gamma may provide a fuller picture of Th1 type responses post-vaccination.

IL-7-induced proliferation of peripheral Th17 cells is impaired in HAART-controlled HIV infection.

OBJECTIVES: Th17 cells are key regulators of functional immunity in mucosal tissues, including the gut-associated lymphoid tissue (GALT), an important site of immune impairment in HIV infection. During HIV infection, Th17 cells are lost in large numbers from the GALT. Despite the recovery of peripheral CD4 T cells that accompanies suppression of viral replication with HAART, Th17 cells in GALT are not completely restored. IL-7 is essential for the survival and proliferation of T cells, but its signaling through its receptor IL-7Rα (CD127), is impaired in CD8 T cells and thymocytes during HIV infection. We set out to determine if decreased CD127 expression or impaired CD127 signaling may be the cause of Th17 impairment in HAART-controlled HIV infection. DESIGN: Healthy and HIV donors on HAART were selected for this study of Th17 cell function in HIV. METHODS: Peripheral CD4 T cells and Th17 cells were isolated using magnetic beads, then stimulated with IL-7. CD127 expression and the phosphorylation of signaling molecules was determined using flow cytometry. Proliferation was determined with a CFSE dilution assay. RESULTS: CD127 was not decreased on Th17 cells from HAART-controlled HIV individuals, in fact, the percentage of Th17 cells that express CD127 was increased in treated HIV individuals. Furthermore, Th17 cells from HAART-controlled individuals, have normal IL-7-induced STAT5 and Bcl-2 responses, but vastly decreased proliferative responses. CONCLUSION: This reduced IL-7 responsiveness may explain the lack of Th17 cell recovery and ongoing systemic immune activation that persists despite well treated HIV infection.

Cord Blood IL-12 Confers Protection to Clinical Malaria in Early Childhood Life.

Using a well-designed longitudinal cohort, we aimed to identify cytokines that were protective against malaria and to explore how they were influenced by genetic and immunological factors. 349 Mozambican pregnant women and their newborn babies were recruited and followed up for malaria outcomes until 24 months of age. Six Th1 cytokines in cord blood were screened for correlation with malaria incidence, of which IL-12 was selected for further analyses. We genotyped IL-12 polymorphisms in children/mothers and evaluated the genotype-phenotype associations and genetic effects on IL-12 levels. Maternal IL-12 concentrations were also investigated in relation to Plasmodium infections and cord blood IL-12 levels. Our data showed that high background IL-12 levels were prospectively associated with a low incidence of clinical malaria, while IL-12 production after parasite stimulation had the opposite effect on malaria incidence. IL-12 genotypes (IL-12b rs2288831/rs17860508) and the haplotype CGTTAGAG distribution were related to malaria susceptibility and background IL-12 levels. Maternal genotypes also exhibited an evident impact on host genotype-phenotype associations. Finally, a positive correlation in background IL-12 levels between maternal and cord blood was identified. Thus, cord blood background IL-12 concentrations are important for protecting children from clinical malaria, likely mediated by both genotypes (children&mothers) and maternal immunity.

Low-level malaria infections detected by a sensitive polymerase chain reaction assay and use of this technique in the evaluation of malaria vaccines in an endemic area.

The feasibility of using a sensitive polymerase chain reaction (PCR) to evaluate malaria vaccines in small group sizes was tested in 102 adult Gambian volunteers who received either the malaria vaccine regimen FP9 ME-TRAP/MVA ME-TRAP or rabies vaccine. All volunteers received the antimalarial drugs primaquine and Lapdap plus artesunate to eliminate malaria parasites. Volunteers in a further group received an additional single treatment with sulfadoxine-pyrimethamine (SP) to prevent new infections. There was substantially lower T-cell immunogenicity than in previous trials with this vaccine regimen and no protection against infection in the malaria vaccine group. Using the primary endpoint of 20 parasites per mL, no difference was found in the prevalence of low-level infections in volunteers who received SP compared with those who did not, indicating that SP did not reduce the incidence of very low-density infection. However, SP markedly reduced the incidence of higher density infections. These findings support the feasibility and potential of this approach to screen pre-erythrocytic vaccines for efficacy against infection in small numbers of vaccinees in endemic areas.

IL-23 signaling in Th17 cells is inhibited by HIV infection and is not restored by HAART: Implications for persistent immune activation.

OBJECTIVES: HIV infection causes a profound depletion of gut derived Th17 cells, contributing to loss of mucosal barrier function and an increase in microbial translocation, thus driving systemic immune activation. Despite normalization of circulating CD4+ T cell counts with highly active antiretroviral therapy (HAART), Th17 frequency and function often remain impaired. Given the importance of interleukin (IL)-23 in the generation and stabilization of Th17 cells we hypothesized that impaired IL-23 signaling causes persistent Th17 dysfunction in HIV infection. METHODS: The effects of in vitro HIV infection on responses to IL-23 in Th17 cells were examined. These included the production of IL-17, phosphorylated STAT3 (pSTAT3) and the transcription of retinoic acid orphan receptor C (RORC) gene. Blood derived Th17 cells from untreated and HAART-treated HIV-infected individuals were also examined for the IL-23 induced production of phosphorylated STAT3 (pSTAT3) and the expression of the IL-23 receptors. RESULTS: In vitro HIV infection significantly inhibited IL-17 production and IL-23 induced pSTAT3 while expression of RORC RNA was unaffected. Th17 cells isolated from untreated and HAART-treated HIV-infected individuals showed complete loss of IL-23 induced pSTAT3 without a decrease in the expression of the IL-23 receptors. CONCLUSIONS: This study is the first to demonstrate an effect of HIV on the IL-23 signaling pathway in Th17 cells. We show that in vitro and in vivo HIV infection results in impaired IL-23 signaling which is not reversed by HAART nor is it a result of reduced receptor expression, suggesting that HIV interferes with IL-23-activated signaling pathways. These findings may explain the inability of HAART to restore Th17 frequency and function and the resulting persistent chronic immune activation observed in HIV infected individuals.

Transcriptional changes induced by candidate malaria vaccines and correlation with protection against malaria in a human challenge model.

INTRODUCTION: The complexity of immunity to malaria is well known, and clear correlates of protection against malaria have not been established. A better understanding of immune markers induced by candidate malaria vaccines would greatly enhance vaccine development, immunogenicity monitoring and estimation of vaccine efficacy in the field. We have previously reported complete or partial efficacy against experimental sporozoite challenge by several vaccine regimens in healthy malaria-naïve subjects in Oxford. These include a prime-boost regimen with RTS,S/AS02A and modified vaccinia virus Ankara (MVA) expressing the CSP antigen, and a DNA-prime, MVA-boost regimen expressing the ME TRAP antigens. Using samples from these trials we performed transcriptional profiling, allowing a global assessment of responses to vaccination. METHODS: We used Human RefSeq8 Bead Chips from Illumina to examine gene expression using PBMC (peripheral blood mononuclear cells) from 16 human volunteers. To focus on antigen-specific changes, comparisons were made between PBMC stimulated with CSP or TRAP peptide pools and unstimulated PBMC post vaccination. We then correlated gene expression with protection against malaria in a human Plasmodium falciparum malaria challenge model. RESULTS: Differentially expressed genes induced by both vaccine regimens were predominantly in the IFN-γ pathway. Gene set enrichment analysis revealed antigen-specific effects on genes associated with IFN induction and proteasome modules after vaccination. Genes associated with IFN induction and antigen presentation modules were positively enriched in subjects with complete protection from malaria challenge, while genes associated with haemopoietic stem cells, regulatory monocytes and the myeloid lineage modules were negatively enriched in protected subjects. CONCLUSIONS: These results represent novel insights into the immune repertoires involved in malaria vaccination.