Roger Brookes

Protective mucosal immunity elicited by targeted iliac lymph node immunization with a subunit SIV envelope and core vaccine in macaques.

Prevention of sexually transmitted HIV infection was investigated in macaques by immunization with a recombinant SIV (simian immunodeficiency virus) envelope gp120 and core p27 vaccine. In two independent series of experiments, we used the novel targeted iliac lymph node (TILN) route of immunization, aiming close to the iliac lymph nodes draining the genitorectal mucosa. Rectal challenge with the SIVmac 32H J5 molecular clone in two series induced total protection in four out of seven macaques immunized by TILN, compared with infection in 13 of 14 unimmunized macaques or immunized by other routes (P = 0.025). The remaining three macaques showed either a decrease in viral load (>90%) or transient viremia, indicating that all seven TILN–immunized macaques showed total or partial protection (P = 0.001). Protection was associated with significant increase in the iliac lymph nodes of lgA antibody–secreting cells to p27 (P < 0.02), CD8–suppressor factor (P< 0.01), and the chemokines RANTES and MIP–1β (P< 0.01).

Homing of mononuclear cells from iliac lymphnodes to the genital and rectal mucosa in non‐human primates

The route of immunization may affect the type of immunity that is induced. The objectives of this investigation were to establish in the non‐human primate if the internal iliac lymph nodes (LN) function as an inductive site of immunity from which mononuclear cells home to the rectal and cervico‐vaginal mucosa. Rhesus macaques were immunized with simian immunodeficiency virus (SIV) core antigen p27 in the proximity of the iliac lymph nodes, and compared with the intramuscular (IM) (deltoid or gluteal), and axillary LN routes of immunization. The macaques were then challenged rectally or vaginally by a particulate SIVp27 antigen which was applied to the mucosal surface. The tracking dye PKH26 was injected near the immunizing LN or IM site and a week later the mucosal and lymphoid tissues were examined at autopsy. Preferential homing of PKH26‐labeled cells from the internal iliac LN to the rectal and vaginal mucosa was demonstrated by flow cytometry after targeted iliac LN (TILN) but not after intramuscular (deltoid) or axillary LN immunization. Homing of the subsets of cells revealed that labeled CD4, CD8 and B cells, as well as monocytes were found in the rectum, colon, vagina or cervix. The results of this investigation shows that the route of immunization may affect regional mucosal immunity. Furthermore, the internal iliac LN may function as an inductive immunological site from which CD4, CD8 and B cells may home preferentially to the rectal, cervical and vaginal mucosa, as well as to the related regional but not the unrelated distal LN.

Extended Culture Enhances Sensitivity of a Gamma Interferon Assay for Latent Mycobacterium tuberculosis Infection

ABSTRACT To test the hypothesis that prolonged culture would enhance the sensitivity of latent tuberculosis detection by a gamma interferon release assay, blood samples from 33 household contacts of Gambian tuberculosis patients were stimulated with Mycobacterium tuberculosis-specific antigens. After 24 h of culture, 66% were positive, compared to 93% after 6 days of culture.

The common mucosal immune system for the reproductive tract: basic principles applied toward an AIDS vaccine

Abstract The concept of the Collaborative Mucosal Immunization Research Group for AIDS (CMIG) was originally conceived by the AIDS Vaccine Branch, National Institute of Allergy and Infectious Diseases (NIAID) in order to provide support for a cooperative research environment for the development of mucosal immunity to AIDS. We have purposely organized five groups of investigators at five different locations to determine how effective mucosal immunity to AIDS can be optimally approached. CMIG recognizes that both rectal (homosexual) as well as vaginal (heterosexual) infections with HIV are two of the major ways that AIDS currently disseminates through the human population. Thus, we have chosen the SIV model of infection of rhesus macaques, but more importantly the CMIG have joined two of our five components in order to use the significant expertise developed for mucosal transmission of SIV and immunity. Thus, we have brought the extensive expertise with vaginal and rectal immunization and immunity to spread [Drs. Chris Miller and Marta Marthas, California Regional Primate Research Center (CRPRC), Davis and Drs. Thomas Lehner and Martin Cranage, United Medical and Dental School Guys Hospital, London and the Centre for Applied Microbiology and Research (Guys/CAMR)]. Two additional components were added in order to perform mucosal immune response studies required to develop and to optimize a mucosal vaccine. First, extensive CD4+ T helper (Th) cell (e.g., Th1 and Th2) and CD8+ T cell subset studies are a major effort of the coordinating group at the University of Alabama at Birmingham (Drs. Hiroshi Kiyono and Jerry R. McGhee). This component is closely interacting with both the CRPRC and Guys/CAMR components in terms of SIV-specific CD4+ and CD8+ T cell subset responses. For example, SIV-specific CTL responses are jointly examined using different techniques by CRPRC, Guys/CAMR and UAB investigators. Further, it is also important to examine a balance between SIV-specific and Th1 and Th2 cell responses following mucosal immunization since the Th cell-derived cytokines are essential for the induction of appropriate antigen-specific mucosal immune responses. This issue is currently being extensively examined by the CMIG effort and a summary of our findings is discussed in this review. A major question in mucosal immunity involves the functions of secretory IgA (S-IgA) antibodies and this area is of particular importance in rectal and reproductive tract immunity. A novel and exciting in vitro epithelial cell assay system is used to study how effectively S-IgA neutralizes SIV infection (Drs. John Huang, John Nedrud and Michael Lamm, Case Western Reserve, Cleveland). A clear advantage of this CMIG effort is the unique expertise in design of mucosal delivery systems for an AIDS vaccine. We are using state-of-the-art recombinant bacteria, i.e.. rSalmonella and rVibrios for mucosal immunization [Drs. Yichen Lu and Bryan Roberts, Virus Research Institute (VRI), Boston]. In addition, we are also testing other mucosal delivery systems including DNA vaccine, microspheres, cholera toxin (CT) and CT-B, recombinant poliovirus, and immune complexes. These studies represent the first efforts to induce not only Th cell mediated S-IgA responses, but also CTL responses to SIV in primates immunized with different mucosal vector delivery systems. In order to focus our effort for the induction of SIV-specific immune responses following mucosal immunization, investigators from the CMIG are attempting to understand the induction and regulation of antigen-specific immune responses in rhesus macaques mucosally immunized with different preparations of SIV vaccines.

Generation of diversity in the hierarchy of T-cell epitope responses following different routes of immunization with simian immunodeficiency virus protein.

ObjectivesTo examine whether the route of immunization determines the hierarchy of T-cell epitope proliferative responses in macaques. DesignMacaques were immunized with a recombinant simian immunodeficiency virus (SIV) p27 core protein by the intramuscular, male and female genital or rectal route, each of which was augmented by oral immunization, and by the novel targeted lymph-node immunization route. Overlapping peptides were used to identify the proliferative T-cell epitopes and to determine their hierarchy in the circulation, spleen and lymph nodes. MethodsT-cell epitope mapping of the proliferative responses was studied in short-term cell lines. Dendritic cells and macrophages were enriched by metrizamide gradient and adherence to plastic, respectively. ResultsIntramuscular immunization elicited in the circulating T cells a hierarchy of T-cell epitopes within four peptides in the following descending order of frequency: peptides 121–140 (57.9%), 41–60 (28.9%), 61–80 (18.9%) and 101–120 (5.4%). The hierarchy of these four T-cell epitope responses differed significantly with each of the five routes of immunization, when circulating (P<0.001), splenic (P<0.02-<0.001) or iliac lymph-node cells (P<0.001) were analysed. The effect of antigen-presenting cells was then investigated and enriched dendritic cells were more effective than macrophages in processing and presenting the p27 antigen and the immunodominant (121–140) and 61–80 T-cell epitopes. ConclusionsThe route of immunization may determine the hierarchy of T-cell epitopes in the lymph nodes draining the mucosa in the circulating and splenic lymphocytes. The diversity of T-cell epitopes may affect the control of HIV at different anatomical sites, the administration route of the vaccine, and selection of polypeptides or recombinant antigens for immunization.