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J R Pink

Publications and source records attributed to J R Pink.

At least 37 records · Page 2Linked to original sources
PubMed1989

Characterizing T-cell epitopes in vaccine candidates.

T-cell clones can be used to test in vitro whether synthetic peptides associate with major histocompatibility complex (MHC) antigens - a necessary condition for the peptides to be immunogenic. Generally, the T-cell clones have been obtained from immune donors, but J.R.L. Pink and F. Sinigaglia argue here that non-immune, human leukocyte antigen (HLA)-typed donors are a useful source of clones and antigen-presenting cells that can be used to assay systematically peptide-MHC associations.

Animals↗
PubMed1988

Nonpolymorphic regions of p190, a protein of the Plasmodium falciparum erythrocytic stage, contain both T and B cell epitopes.

Two conserved regions from the genetically polymorphic p190 molecule of the malaria parasite Plasmodium falciparum have previously been expressed in Escherichia coli as separate polypeptides (190.L and 190.M) or as a single fusion protein (190.N). In the present study we investigated whether human B and T lymphocytes recognize these conserved regions. The more amino-terminal region, 190.L (corresponding to residues 188-363 of the encoded protein sequence) reacted preferentially with sera from donors living in a malaria-endemic area. Also, EBV-transformed B cells, from a healthy donor living in a malaria-mesoendemic area, were fused with a human-mouse hybrid line (SPM4-0), yielding two hybridomas whose products recognized both 190.L and the fusion protein 190.N, but not the 190.M polypeptide. A large number of p190-specific T cell clones were obtained from PBMC of a noninfected donor, after in vitro stimulation with the recombinant fusion protein 190.N. The clones reacted with intact, parasite-derived p190, as well as either 190.L or 190.M. Four clones that recognized the more amino-terminal fragment also responded to infected E. According to these results the more amino-terminal conserved sequences of p190 have the requisites to be immunogenic in humans.

Adult↗
PubMed1988

Assessment in humans of a synthetic peptide-based vaccine against the sporozoite stage of the human malaria parasite, Plasmodium falciparum.

Eleven volunteers were injected with an anti-malaria (Plasmodium falciparum) sporozoite vaccine candidate consisting of a synthetic peptide, Ac-Cys-(NANP)3, coupled to tetanus toxoid (TT) and adsorbed to aluminum hydroxide. Two of the volunteers had no previously known exposure to TT. Eight volunteers made detectable antipeptide, anticircumsporozoite protein or antisporozoite antibodies, whose titers increased after multiple injections in four individuals. The maximum antisporozoite titer obtained in an immunofluorescence assay was 1280. In those individuals who produced antipeptide antibody, the overall correlation between IgG anti-Ac-Cys-(NANP)3 antibody in enzyme-linked immunosorbent assay and IgG antisporozoite reactivity in immunofluorescence was highly significant. However, the fine specificity of antibody varied among volunteers with two individuals producing mostly antipeptide antibody. Anti-TT antibody responses increased in all volunteers with the exception of that person who had the highest pretrial anti-TT titer; this individual was one of the two pre-TT-immunized volunteers who failed to produce anti-Ac-Cys-(NANP)3 or sporozoite antibody. For the two non-TT preimmunized volunteers, one produced an antisporozoite fluorescence titer of 320; the other made no detectable antibody against either Ac-Cys-(NANP)3 or sporozoites during a primary response. For the three volunteers monitored, after the first injection, significant T cell proliferative responses to (NANP)3 were observed, which increased up to 4 wk after immunization, when a second injection was given. Responsiveness then declined to background levels and did not reappear after further immunizations. In contrast, a marked TT-specific proliferation was observed for the duration of the study.

Adult↗
PubMed1988

Antibody-forming capacity of B cell-deficient chickens reconstituted with limiting numbers of B cell precursors.

To examine the antibody-forming capacity of neonatally cyclophosphamide-treated chickens reconstituted with limiting numbers of B cell precursors, we analyzed their antibody responses to six unrelated antigens. Our results demonstrate that about 10 x 10(6) bursal cells are needed in this adoptive cell transfer model to restore normal immune competence to B cell-deficient birds. From our earlier data we know that with low repopulating cell numbers (less than 10 x 10(6) cells) developing bursal follicles are of clonal origin. Ten million cells repopulate about 40%, i.e. about 4 x 10(3) of the bursal lymphoid follicles. Assuming the clonal origin of the follicles these results imply that the B cell system of birds receiving this dose is derived from less than 5 x 10(3) precursor cells. At the lowest reconstituting dose (1.25 x 10(6) cells) most birds do not respond to the antigens studied. However, their B cell system is derived from only about 500 precursor cells. Because the antibody repertoire of a normal chicken was estimated to be at least 10(6) our results suggest that each precursor gives rise to a large number (greater than 200) of immunoglobulin V-region gene variants during its clonal proliferation in the bursa. Our results are thus consistent with the proposed "hyperconversion" mechanism of generation of antibody diversity in the chicken and provide quantitative data useful for estimating what such somatic modification rates might be.

Animals↗
PubMed1988

Epitopes recognized by human T lymphocytes on malaria circumsporozoite protein.

The circumsporozoite protein of the malaria parasite Plasmodium falciparum contains regions of nonrepetitive sequences which are predicted to be T cell recognition sites. We synthesized peptides corresponding to three of these regions, and tested their ability to stimulate proliferation of peripheral blood lymphocytes from donors living in a malaria-endemic area, or from nonimmune donors. Cells from 15 out of 22 donors (including 4 of 6 nonimmune individuals) were stimulated by one or more of the peptides. T cell clones specific for one of the peptides were obtained and shown to recognize the native protein purified from sporozoites. These data help to identify T cell epitopes which could be incorporated into a malaria vaccine.

Animals↗
PubMed1988

Immunoglobulin diversification in embryonic chicken bursae and in individual bursal follicles.

Previous studies have shown that the same immunoglobulin (Ig) V lambda gene (V lambda 1) is rearranged in all chicken B cells, and that extensive sequence diversification of this gene occurs during B cell development in the bursa of Fabricius. We used two-dimensional gel electrophoresis to compare the heterogeneity of Ig lambda light chains produced by B cells at different stages of bursal development. Somatically diversified light chains were observed in Ig molecules produced by bursal cells as early as 15 days of embryonic incubation. The two principal species of light chain observed probably represent glycosylated and nonglycosylated forms of lambda chain encoded by alleles of a single lambda gene. Extensive diversification was observed during late embryogenesis. We also studied lambda light chain diversity in cyclophosphamide-treated birds repopulated with normal bursal cells. In these birds, individual bursal follicles are repopulated by single B cell precursors. Follicular cells derived from single B cell precursors were able to produce a spectrum of light chains almost as diverse as that of the total bursal cell population. We used two monoclonal anti-idiotype antibodies to study idiotype expression in individual normal or reconstituted follicles. About 30% of follicles contained 0.1% to 5% of lymphocytes which reacted with one or both of the antibodies. The results indicate that within individual bursal follicles bursa stem cells undergo Ig hyperdiversification.

Animals↗
PubMed1988

B cell precursors in chick embryos surgically bursectomized at 72 h of incubation.

To determine the presence of precursor B cells in chick embryos surgically bursectomized at 72 h of incubation (E-Bx) we studied chick chimeras that were produced by establishing parabiotic connections between blood vessels of chorioallantoic membranes of normal and surgically bursectomized chick embryos. Using sex chromosomes and a B cell alloantigen (Bu-1a) as markers we showed that chick embryos bursectomized at 72 h of incubation contain B cell precursors capable of colonizing the bursa of Fabricius and developing into B lymphocytes. The repopulation capacity of 14-day-old embryonic spleen cells from E-Bx recipients was tested by transferring them into age-matched X-irradiated Bu-1-disparate embryos. The results show that B cell precursors are present in 14-day spleen of chick embryos bursectomized at 72 h of incubation. These precursors carry the Bu-1 B cell alloantigen, suggesting that commitment to the B cell lineage can take place in the absence of bursa.

Age Factors↗
PubMed1988

Human T cells recognize polymorphic and non-polymorphic regions of the Plasmodium falciparum circumsporozoite protein.

In order to characterize T cell epitopes in the Plasmodium falciparum circumsporozoite (CS) protein sequence, we isolated T cell clones, from non-immune donors, which reacted with synthetic peptides corresponding to two predicted CS protein T cell epitopes. Peptide CS.T3 (corresponding to a non-polymorphic region of the CS protein, residues 378-398) was recognized in association with either DR2 or DRw9 restriction elements. T cell clones recognizing CS.T3 also reacted with the sporozoite-derived CS protein. Peptide CS.T2 corresponds to a polymorphic region (residues 325-341) of the CS protein. Unlike the CS.T3-specific clones, the CS.T2-specific clones did not recognize the CS protein. Since the CS.T2 peptide includes residues which are polymorphic in different P. falciparum isolates, we investigated whether these residues were critical for recognition of the peptide. We show here that a single amino acid substitution at a position of the CS protein which shows genetic polymorphism affects recognition of the sequence by human T cells. The implications of these data for malaria vaccine development are discussed.

Animals↗
PubMed1988

Susceptibility to tolerance induction of bursal and peripheral B cells.

We investigated cellular aspects of immunological tolerance to protein antigens in chickens by examining the immune responses of bursal and splenic cells from tolerant or normal chickens after transfer into cyclophosphamide (CP)-treated recipients. Newly-hatched chicks were made tolerant to bovine serum albumin (BSA) by injection of 100 mg of the antigen. When bursa cells from 4-day-old BSA-unresponsive chicks were transferred into CP-treated recipients, the reconstituted birds were able to respond to a subsequent injection of BSA almost as well as normal birds, and as well as CP-treated birds that had been reconstituted with normal bursa cells. To investigate whether the presence of the BSA antigen might affect recovery from tolerance, we injected CP-treated recipients with BSA at the time of transfer of bursal cells. The presence of the antigen prevented the recovery of the anti-BSA response in reconstituted birds. When spleen cells from 6.5-week-old unresponsive chicks were transferred into CP-treated recipients, no recovery of responsiveness to BSA could be demonstrated. A likely reason for the failure of splenic B cells to recover responsiveness on transfer is their inability to generate somatic variants of Ig genes in the same way as bursal stem cells. Thus, when the bursa involutes, the chicken's antibody repertoire may be frozen in a less adaptable state than that of a mammal.

Animals↗
PubMed1987

Plasmodium falciparum-specific human T cell clones: evidence for helper and cytotoxic activities.

This report describes the isolation, and the phenotypic and functional characterization of Plasmodium falciparum-specific human T lymphocyte clones (TLC) obtained from 2 acutely infected and 4 clinically immune donors. Approximately one third of the TLC obtained from the acutely infected patients had the phenotype CD8+/CD4-. No such clones were obtained from the clinically immune donors. P. falciparum-specific, major histocompatibility complex-restricted CD8+ clones can lyse unrelated tumor cell targets in the presence of anti-CD3 antibodies, suggesting that these clones have cytotoxic potential. Conversely no killing was obtained with two CD4+ P. falciparum-specific TLC, even in the presence of anti-CD3 antibody, In addition the helper function of a CD4+ clone was demonstrated in a T/B cell cooperation system.

Animals↗
PubMed1987

Plasmodium falciparum-specific human T cell clones: recognition of different parasite antigens.

T lymphocyte clones specific for malarial (Plasmodium falciparum) blood stage antigens were obtained from acutely infected patients or from donors living in a malaria-endemic area of West Africa. Thirty-four clones carrying the CD4 antigen, and one CD8+ clone, were tested in a proliferation assay for their capacity to recognize P. falciparum isolates of different geographical origins. Only one clone distinguished between different parasite isolates (it failed to react with a parasite isolate originating from East Africa, but did recognize West African and Asian isolates). All of the clones responded well to intact erythrocytes containing viable parasites, but some responded poorly to extracts of parasitized cells. Eight of 19 clones studied (all CD4+) recognized parasite antigens which had characteristic mobilities in sodium dodecyl sulfate-containing polyacrylamide gels. The antigens had apparent molecular weights of about 20,000, 35,000, 40,000, 120,000, 150,000-200,000 and 200,000. These results (together with a previous report of two clones recognizing an antigen of molecular weight about 50,000, Sinigaglia and Pink, EMBO J. 1985. 4:3819) show that T cells in infected individuals react with at least 6 different parasite proteins.

Animals↗
PubMed1987

A re-evaluation of the function of the bursa of Fabricius.

We will briefly outline mammalian B cell ontogeny to provide a comparison with the avian model. The earliest defined stage of mammalian B cell development is the pre-B cell (itself derived from a multipotent stem cell) which expresses heavy chains within the cytoplasm and is a large, rapidly dividing cell. This cell drops out of division, reduces in size and, over about 24 hours, rearranges Ig light chain V region gene. As a consequence of light chain rearrangement and expression, intact IgM is expressed on the cell surface and the cell leaves the bone marrow as a small virgin B lymphocyte with the capacity to respond to antigen (Opstelten and Osmond 1983). This pathway occurs throughout the life of the animal and so there is constant production of B cells from the bone marrow which are recently derived from sIg- precursors, which have, in turn, recently rearranged their Ig V region genes. Thus there is a constant influx of new V region gene combinations into the peripheral mammalian B cell pool. Furthermore, the mammalian B cell repertoire is very large with estimated germ line diversity approaching 5 X 10(7) as a consequence of various germ-line V region recombinations (Honjo 1983). The pre-bursal stem cell is present in the periphery of the embryo from before day 8 to about day 16 - 17 of embryonation. Most colonisation of bursal follicles probably occurs prior to day 13. Individual bursal follicles are populated by a low number (2-3) of precursor cells. These rapidly become committed for the expression of particular V region genes, probably as a consequence of productive V region recombination. This recombination is restricted to the embryo and may not require the bursal microenvironment for its induction. The available germ-line repertoire of heavy and light chain V region in the chicken is very limited and little diversity can be generated from this initial recombination event (see Weill et al 1986). By day 12 of embryonation, sIg+ cells are present in the bursa and from this time sIg+ cells rapidly divide within the medulla of the bursal follicle. By about day 18 of embryonation, there are no pre-bursal cells in the periphery and sIg+ cells begin to seed from the bursa. It is from about this time that the cortex appears within the bursal follicle and it is tempting to suggest that cells migrate from the medulla to the cortex where further cell division occurs prior to export into the periphery.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Ontogeny and tissue distribution of the chicken Bu-1a antigen.

By using a sensitive technique of immunofluorescence on polyethylene glycol-embedded tissue sections, we could better determine the distribution of L22+ cells in embryonic and adult chickens. L22 mAb was originally described as reacting with bursa and bursa-derived lymphocytes. We now present evidence to suggest that this mAb also reacts with a subset of macrophages found in bursa, thymus, spleen, liver, intestine and peritoneum. The L22+ cells appear early during embryonic life, simultaneously in yolk sac, bursa, thymus, spleen and bone marrow. At all steps of their ontogeny, thymocytes were L22-, while most, if not all, bursal lymphoid cells were L22+. Moreover, L22 antigen can be detected on haemopoietic cells probably precursors, before and during their entry into the bursal rudiment on Day 9 or 10 of embryonic life. We conclude from these data that L22 is not restricted to the B-cell lineage as it is shared with a subset of macrophages. Furthermore, our observations of L22+ cells during embryonic life favour the hypothesis of separate lineages for B-cell and T-cell precursors in chick embryo, which was previously put forward on the basis of different sets of experiments.

Animals↗