Biomedical subjects

V Udhayakumar

Publications and source records attributed to V Udhayakumar.

At least 19 recordsLinked to original sources

PubMed2000

Immunity to placental malaria. II. Placental antigen-specific cytokine responses are impaired in human immunodeficiency virus-infected women.

An association was demonstrated recently between elevated in vitro production of interferon (IFN)-gamma by intervillous blood mononuclear cells (IVBMCs) and protection against placental malaria (PM). Because human immunodeficiency virus (HIV)-infected pregnant women have increased susceptibility to PM, loss of the IFN-gamma response in these women may impair their ability to control PM. Measurement of cytokines in culture supernatants by ELISA revealed that IFN-gamma responses by HIV-positive IVBMCs were impaired, especially after malarial antigen stimulation. Interleukin (IL)-4 and IL-10 responses also were reduced in HIV-positive persons, the latter more so in HIV-positive, PM-positive persons. In contrast, tumor necrosis factor-alpha production generally was enhanced in PM-positive and HIV-positive persons. Overall, cytokine production was reduced in HIV-positive persons with CD4 T cell counts <500/microL, particularly in response to malarial antigen. Thus, HIV-mediated cytokine dysregulation and impairment of the protective IFN-gamma response may contribute to the increased susceptibility of HIV-positive pregnant women to malaria.

Adult↗

PubMed2000

Development, expression, and murine testing of a multistage Plasmodium falciparum malaria vaccine candidate.

A synthetic gene encoding twelve B cell epitopes, six T-cell proliferative epitopes, and three cytotoxic T lymphocyte (CTL) epitopes from nine stage-specific antigens, representing the sporozoite, liver stage, asexual blood-stage, and sexual-stage antigens of Plasmodium falciparum, was constructed by assembling overlapping oligonucleotides followed by PCR extension and annealing. A three-step PCR protocol using twelve long oligonucleotides was employed to generate a 1053 base-pair synthetic gene, the identity of which was confirmed by sequencing. This synthetic gene, named CDC/NII MAL VAC-1, was cloned, and the recombinant protein was expressed in the Baculovirus Expression Vector System (BEVS). The selection of malarial epitopes for inclusion in this vaccine construct was based on immunoepidemiological studies in malaria endemic area, in vitro, and in vivo protection studies in model systems. The 41 kDa BEVS-expressed recombinant protein reacted with mouse antibodies specific for individual B cell epitopes in the vaccine construct and with sera from clinically immune Kenyan adults. An immunization study in three strains of mice that differ at the H-2 locus demonstrated that the BEVS-expressed recombinant protein is immunogenic; the candidate vaccine antigen induced high titer antibodies, and lymphocyte proliferative and IFN-gamma responses. These results demonstrate that individual B and T cell epitopes can be assembled to create synthetic genes that encode proteins capable of eliciting specific antibody and T cell responses.

Adult↗

PubMed2000

Field studies of cytotoxic T lymphocytes in malaria infections: implications for malaria vaccine development.

The search for a cytotoxic T lymphocyte (CTL)-inducing malaria vaccine has moved forward from epitope identification to planning stages of safety and immunogenicity trials of candidate vaccines. Development of CTL-inducing vaccine candidates has taken center stage based on the observation that CTL-mediated protection might be the dominant mechanism by which sterile immunity is achieved in irradiated sporozoite immunization experiments in humans and laboratory animals. However, studies in naturally infected individuals living in endemic areas, as reviewed here by Michael Aidoo and Venkatachalam Udhayakumar, have revealed that CTL induction might be influenced by factors such as parasite variants, host genes, other infections and transmission patterns. The influence of these factors on CTL induction has been demonstrated individually and in various combinations in controlled animal experiments. However, in naturally infected humans, they are presented in a complex host-parasite-environment interaction, in a manner that is not easily achieved in laboratory-based experiments. Understanding these interactions is crucial for the development and testing of CTL-inducing vaccines for humans.

Amino Acid Sequence↗

PubMed2000

Immunologic memory in the placenta: a lymphocyte recirculation hypothesis.

The placenta is an immunologically unique organ where a balance between maternal immunity and fetoplacental well-being must be maintained for successful pregnancy to occur. The intervillous blood is important in this context, yet little is known about local immunologic processes, particularly how placenta-specific memory immune responses are maintained. Using malaria as an illustrative case, we describe an hypothetical model in which recirculation of memory T lymphocytes from the intervillous blood to local lymphoid tissue facilitates maintenance of local memory immunity. This explains how memory cells might be retained when the placenta is expelled at parturition and thus remain available for rapid recall from the local lymphoid tissue to the intervillous space when exposure to the same antigenic stimulus occurs in subsequent pregnancies. Study of cell-mediated immunity to infections like malaria in the intervillous blood and the use of animal models will be necessary to provide proof for this hypothesis.

Female↗

PubMed2000

Host-pathogen interactions in emerging and re-emerging infectious diseases: a genomic perspective of tuberculosis, malaria, human immunodeficiency virus infection, hepatitis B, and cholera.

On exposure to a pathogen, a host may resist infection, become subclinically infected, or progress through several stages from mild to severe infection. Chronic sequelae may or may not occur. Host factors, particularly host genes, influence many of these stages. We have used a model of the continuum of pathogenesis of infectious diseases to consider the effect of host genes on five pathogens of significant public health burden: Mycobacterium tuberculosis, Plasmodium species, human immunodeficiency virus, hepatitis B virus, and Vibrio cholerae. The relationships between these infections and polymorphisms in human leukocyte antigen, cytokines, other immune response, or pathogen receptor genes are reviewed. We discuss gene-gene interactions and their effects in complex settings, such as coinfections with several pathogens. Priorities for prevention and control of these pathogens include vaccines and antimicrobial drugs. Research on how host genes can influence vaccine responses and the efficacy of drugs or other interventions, as well as further research into the relationship of host genes to infectious disease outcomes, may lead to new strategies for prevention and control.

Cholera↗

PubMed1999

A low interleukin-10 tumor necrosis factor-alpha ratio is associated with malaria anemia in children residing in a holoendemic malaria region in western Kenya.

The balance between Th1 cytokines (tumor necrosis factor [TNF]-alpha, interferon [IFN]-gamma) and Th2 cytokines (interleukin [IL]-10, -4) may be critical in the development of severe falciparum malaria. Therefore, plasma concentrations of these cytokines were determined in children with various manifestations of malaria. Plasma levels of IFN-gamma and IL-4 were undetectable in most children. However, TNF-alpha and IL-10 were significantly elevated in children with high-density parasitemia and malaria anemia compared with children in control groups. In children with mild malaria, IL-10, but not TNF-alpha, was significantly elevated. While the highest concentrations of TNF-alpha were found in children with malaria anemia, IL-10 levels were highest in children with high-density uncomplicated malaria. The mean ratio of IL-10 to TNF-alpha was significantly higher in children with mild and high-density parasitemia (4.64, P<.005) than in children with malaria anemia (1.77). Thus, higher levels of IL-10 over TNF-alpha may prevent development of malaria anemia by controlling the excessive inflammatory activities of TNF-alpha.

Anemia↗

PubMed1999

Immunity to placental malaria. I. Elevated production of interferon-gamma by placental blood mononuclear cells is associated with protection in an area with high transmission of malaria.

In areas in which malaria is holoendemic, primigravidae and secundigravidae, compared with multigravidae, are highly susceptible to placental malaria (PM). The nature of gravidity-dependent immune protection against PM was investigated by measuring in vitro production of cytokines by placental intervillous blood mononuclear cells (IVBMC). The results demonstrated that interferon (IFN)-gamma may be a critical factor in protection against PM: production of this cytokine by PM-negative multigravid IVBMC was elevated compared with PM-negative primigravid and secundigravid and PM-positive multigravid cells. Low IFN-gamma responsiveness to malarial antigen stimulation, most evident in the latter group, was balanced by increased interleukin (IL)-4 production, suggesting that counter-regulation of these two cytokines may be a crucial determinant in susceptibility to PM. A counter-regulatory relationship between IL-10 and tumor necrosis factor-alpha was also observed in response to malarial antigen stimulation. These data suggest that elevated production of IFN-gamma, as part of a carefully regulated cytokine network, is important in the control of PM.

Adult↗

PubMed1999

Reply

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Journal Article↗

PubMed1999

Differential effect and interaction of monocytes, hyperimmune sera, and immunoglobulin G on the growth of asexual stage Plasmodium falciparum parasites.

Using a flow cytometry-based parasite growth inhibition assay (GIA) and an antibody-dependent cellular inhibition (ADCI) assay, we have assessed the differential effect and interaction of monocytes, immune sera, and purified immunoglobulins from Kenyan adults on the growth of Plasmodium falciparum parasites in vitro. We found that monocytes from 14 different normal, healthy, non-malaria-exposed donors had varying effects on parasite growth, i.e., inhibition or enhancement of parasitemia, suggesting heterogeneity in anti-parasitic activities of monocytes from individual donors. Twenty-two serum samples collected from clinically immune adults from western Kenya inhibited growth of P. falciparum after 48 hr in culture. In contrast, all IgG preparations, except one, purified from the same serum samples enhanced parasite growth. In ADCI experiments, of the 22 purified IgG samples used, 11 showed ADCI activities with specific growth inhibition (SGI) of more than 10%, with the highest at 27.6%, and the remaining 11 IgG samples had an SGI of less than 10%. Our results also showed that the ratio of IgG1 to IgG3 antibodies, as determined by an indirect immunofluorescence assay, was higher in the high ADCI response group than in the low response group, suggesting that a higher concentration of IgG1 antibodies with a higher IgG1/IgG3 ratio might be associated with ADCI activities. The present study has resulted in the development of simple, reproducible flow cytometry-based GIA and ADCI assays, and also provides baseline information for further investigation of the role of ADCI activity in naturally acquired immune protection against malaria.

Adult↗

PubMed1999

Longitudinal cohort study of the epidemiology of malaria infections in an area of intense malaria transmission I. Description of study site, general methodology, and study population.

A large-scale longitudinal cohort project was initiated in western Kenya in June 1992. The primary purpose of the project was to study Plasmodium falciparum malaria in a highly endemic area using a comprehensive and multidisciplinary approach, which included epidemiology, entomology, and immunology. Between June 1992 and July 1994, pregnant women living in 15 rural villages were identified during a monthly census and 1,164 were enrolled. The women were followed-up throughout their pregnancy and they, along with their newborn infants and direct siblings of the infants' less than 15 years of age, were monitored over time. As of May 1995, 1,017 infants had been born to these women. This paper presents the design and general methodology used in this study and describes the initial experience with intense monitoring of a large population over a prolonged period.

Child, Preschool↗

PubMed1998

Postpartum coma.

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Coma↗

PubMed1998

A longitudinal investigation of IgG and IgM antibody responses to the merozoite surface protein-1 19-kiloDalton domain of Plasmodium falciparum in pregnant women and infants: associations with febrile illness, parasitemia, and anemia.

This study was aimed at delineating characteristics of naturally acquired immunity against the merozoite surface antigen-1 (MSP-1) of Plasmodium falciparum, a candidate malaria vaccine antigen. A case/control study was performed on 75 case/control pairs of infants with febrile illness at the time of the first detected infection indicating a clinical case. The presence and level of antibodies at one month prior to the first infection and at the time of the first infection in the afebrile group was significantly higher than in the febrile group. Decreased parasite density and decreased infection-related loss of hemoglobin was seen in infants with anti-MSP-1(19kD) IgG antibodies. In addition, mothers who were positive for the presence of these antibodies conferred protection against placental infection and infection in their infants. In this study, development of anti-MSP-1(19kD) antibody responses in 24 infants were studied longitudinally using monthly serum samples collected from birth until approximately one year of age. In addition, umbilical cord blood sera and respective mothers' sera were analyzed. Longitudinal studies of antibody responses revealed several short-lived IgG and IgM peaks throughout an infant's first year that correlated with detection of parasitemia. The protection against parasitemia and febrile illness was observed in infants when anti-MSP-1(19kD) antibodies were present; when infants were negative for IgG, they had a 10-times greater risk of becoming parasitemic. These data from a longitudinal and prospective study of malaria suggest a protective role for anti-MSP-1(19kD) antibodies in infants and pregnant women.

Anemia↗

PubMed1997

A simple perfusion technique for isolation of maternal intervillous blood mononuclear cells from human placentae.

A noninvasive perfusion method for the recovery of maternal placental (intervillous) blood for use in immunologic assays is described. 60% of the perfused blood samples tested for fetal red blood cell (RBC) contamination were found to be pure maternal blood; in the remainder, fetal RBC contamination, with a single exception, was less than 6%. The intervillous mononuclear cells (IVBMC) isolated from this blood were of predominantly maternal origin as demonstrated by a polymerase chain reaction-based DNA typing technique. The number of IVBMC obtained was within the range of 9 to 55 X 10(6) cells. Phenotypic analysis of IVBMC surface antigens revealed that 61% of the cells were CD3 + T-cells and 18% were CD19 + B-cells. The CD4 + and CD8 + T-lymphocyte subsets accounted for 28 and 26% of the IVBMC, respectively. The IVBMC were functionally competent as evidenced by in vitro lymphoproliferation and cytokine production in response to mitogen and PPD stimulation. This technique allows for rapid and safe isolation of large numbers of IVBMC which are functionally active up to 12 h post-delivery, thus representing a significant improvement over previously described methods. It should facilitate more vigorous research in the study of uteroplacental immunity and infectious disease research, particularly in field settings where sample collection and laboratory facilities are distant.

Chorionic Villi↗

PubMed1997

Cytotoxic T cell reactivity and HLA-B35 binding of the variant Plasmodium falciparum circumsporozoite protein CD8+ CTL epitope in naturally exposed Kenyan adults.

In this study, we have investigated the extent of natural polymorphism in the CD8+ cytotoxic T lymphocyte (CTL) determinant (amino acids 368-390) of circumsporozoite (CS) protein of Plasmodium falciparum field isolates from a holoendemic region of Kenya, and determined how this variation affects the CTL reactivities in clinically immune adults and binding specificities to human histocompatibility leukocyte antigen (HLA)-B35. Among the eight variant sequences that were found in this region, four were new and not seen in parasites from other geographical regions. When synthetic peptides corresponding to the eight variants were used to test the presence of CTL response in different donors, a different spectrum of CTL reactivity to these variants was noticed. While CTL from some donors recognized the P1 sequence (the most prevalent type of sequence) but not P8 (another major variant), other donors showed a reverse pattern of reactivity. Although none of the donors was able to recognize all the variants, CTL responses to all the eight variant sequences were found in this population. An octamer peptide with P1 sequence KPKDELDY in this polymorphic determinant was known to bind HLA-B35. When we tested the effect of natural variation in this octamer sequence on HLA-B35 binding, it became evident that SP13 with D --> N substitution retained its binding specificity to HLA-B35. On the other hand, the SP12 octamer sequence which had two substitutions did not bind HLA-B35. The most interesting finding was the observation that a D --> G substitution at position 374 rescued the binding ability of SP14, which otherwise could not bind to this HLA molecule due to E --> Q amino acid substitution at position 372. To our knowledge, this is the first demonstration showing that a natural polymorphism can rescue the binding specificity to an HLA-class I molecule that was lost due to another natural amino acid substitution. Altogether, these results demonstrate that natural polymorphism in the CS protein affects both the CTL reactivity and the ability to bind to HLA-B35.

Adult↗

PubMed1997

Rapid onset of malaria-induced mortality by immunizations with lipo-peptides: an experimental model to study deleterious immune responses and immunopathology in malaria.

We have recently shown that circumsporozoite (CS) protein-based cytotoxic T-cell epitope of Plasmodium berghei coupled to monoplamitic and tripalmitic acid was able to induce cytotoxic T-cell responses. In the present study, we investigated whether lipopeptide derivatized CS protein B and T helper epitopes in different combinations will be able to induce protective immune responses against sporozoite challenge. Several P. berghei CS peptides with monopalmitic fatty acid tails were prepared, suspended in an oil-in-water emulsion, and used to immunize and boost female A/J mice. The mice were challenged iv. with viable sporozoites of P. berghei (ANKA) two weeks after the last immunization. While immunization with some of these vaccine formulations induced protective immune responses, others shifted the typical bimodal pattern of P. berghei sporozoite induced death toward a rapid onset of death in a peptide specific manner. Therefore, demonstration that immunization with formulations of malarial peptides can cause enhanced malaria-related death provides an experimental model to delineate characteristics of deleterious immune responses.

Amino Acid Sequence↗

PubMed1997

Induction of protective antibodies in Saimiri monkeys by immunization with a multiple antigen construct (MAC) containing the Plasmodium vivax circumsporozoite protein repeat region and a universal T helper epitope of tetanus toxin.

Previous attempts in inducing protective immunity against Plasmodium vivax in human volunteers and nonhuman primates with recombinant circumsporozoite (CS) proteins have been unsuccessful, largely due to the failure of generating antibodies against the protective B epitope AGDR in the CS protein repeat region. We report here an immunization study in Saimiri monkeys with a multiple antigen construct (MAC) containing the P. vivax CS protein repeat region and a T helper epitope of tetanus toxin formulated in different adjuvants. Monkeys immunized three times with MAC in copolymer P1005, copolymer P1005 plus RaLPS, or MF-75 had titers of antibodies against CS repeat, sporozoites and the protective B epitope AGDR significantly higher than those immunized with MAC in alum or PBS (P < 0.05). Antibody levels in animals that received P1005 were maintained at high level for 7 months after the last immunization. Upon challenge with 10000 sporozoites 2 weeks after the last immunization, 75% (three of four) of monkeys from the alum group, 50% (three of six) of monkeys from the P1005 plus RaLPS group, 40% (two of five) of monkeys from the P1005 group, 33% (two of six) of monkeys from the MF-75 group, and 17% (one of six) of monkeys from the MAC alone group were fully protected. When immunized animals were challenged again with 30000 sporozoites 22 weeks after the last immunization. 40% (two of five) monkeys from the P1005 group were fully protected. The remaining (three) in this group developed low parasitemia (< 2000 parasites mm-3 of blood) after significantly longer prepatent period (P < 0.05). In addition, 17% (one of six) of monkeys each from the P1005 plus RaLPS and MF-75 groups were also fully protected. Protected animals had higher levels of prechallenge anti-AGDR antibody titers than unprotected (1933 vs 281 for the first challenge, P > 0.05; 21527 vs 196 for the rechallenge, P < 0.05). Anti-AGDR antibody titers were positively correlated with the prepatent period of infected animals (r = 0.42 for the first challenge, P > 0.05; r = 0.60 for the rechallenge, P < 0.05) and negatively correlated with the peak parasitemia (r = -0.39 for the first challenge, P < 0.05; r = 0.50 for the rechallenge, P < 0.05). The results suggested that when combined with the use of potent adjuvants and T helper epitopes, MAC subunit vaccines may potentially offer protection against malaria infection.

Amino Acid Sequence↗

PubMed1996

Quantitation of RT-PCR amplified cytokine mRNA by aequorin-based bioluminescence immunoassay.

We described here a bioluminescence-based immunoassay for the quantitation of RT-PCR amplified cytokine mRNA. This technique uses a standard RT-PCR procedure, with the following modifications. The forward primer in the PCR reaction is labeled with a 5' biotin molecule. Following PCR, a digoxigenin-conjugated oligonucleotide probe is hybridized to the target biotin-labeled DNA template. The hybridized duplex is captured onto a streptavidin-coated microtiter plate. The bound product is quantitated by adding digoxigenin-specific antibodies conjugated with the photoprotein aequorin. The amount of specific DNA captured onto the plate is quantitated by triggering the bioluminescence reaction through the addition of calcium ions. This technique detected as low as 40 amol of amplified cytokine products, or 500 copies of templates when 27 PCR cycles were used. The high sensitivity of this technique enables the quantitation of target DNA during the exponential phase of the PCR reaction. The aequorin-bioluminescence assay is an alterative non-radioactive method for the quantitation of PCR products.

Aequorin↗

PubMed1996

Plasmodium falciparum: involvement of additional receptors in the cytoadherence of infected erythrocytes to microvascular endothelial cells.

The involvement of additional ligands in the cytoadhesion of PRBC to endothelial cells was studied by the use of human microvascular endothelial cells (HMEC-1), brain microvascular endothelial cells (HBEC-51), umbilical vein endothelial (HUVEC), and C32 melanoma cells as well as soluble CD36, ICAM-1, and thrombospondin in the adhesion assays. Immunostaining showed that ICAM-1 and thrombospondin were expressed by all cell lines, whereas CD36 and VCAM-1 were expressed constitutively only by C32 melanoma cells and HBEC-51, respectively; none of these cells had basal expression of E-selectin. Bindings of the parental HB3 parasite strain to HMEC-1 and HUVEC were higher than that to HBEC-51 and C32 melanoma cells. Selections by panning the parental HB3 through HMEC-1 (HB3EC-6 line) or C32 melanoma cells (HB3C32-6 to HMEC-1 was higher than that to C32 melanoma cells. Antibody or peptide blockade against CD36, ICAM-1, and thrombospondin or preincubation of target cells with TNF-alpha and IFN-gamma did not significantly alter the binding intensity of HB3EC-6 to HMEC-1 and HB3C32-6 to C32 melanoma cells. Preincubation of HMEC-1 with IL-4, however, reduced its binding with HB3EC-6. In vitro selection did not enhance the binding of PRBC to plate-bound CD36 or thrombospondin; binding to ICAM-1 was negligible. The binding of both selected lines was inhibited by dextran sulfate and sulfatides, but not by chondroitin sulfate A. These results suggested that in addition to CD36 and thrombospondin, sulfated glycoconjugates were probably concurrently utilized by these PRBC as receptors. Experiments with freshly isolated Kenyan parasites indicated that they also exhibited a similar mechanism of binding to endothelial cells.

Animals↗