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Biomedical subjects

K Marsh

Publications and source records attributed to K Marsh.

At least 181 records · Page 10Linked to original sources
PubMed1988

A highly conserved amino-acid sequence in thrombospondin, properdin and in proteins from sporozoites and blood stages of a human malaria parasite.

As a consequence of gene cloning and DNA sequencing several gene families are emerging in the field of cell-cell recognition. These include immunoglobulins, integrins, certain extracellular glycoproteins and a family of functionally unrelated proteins which include factor B. We report here the cloning and sequencing of a gene from Plasmodium falciparum, coding for a protein we call thrombospondin related anonymous protein (TRAP), which shares certain sequence motifs common to other well-characterized proteins. The most significant homology is based around the sequence Trp-Ser-Pro-Cys-Ser-Val-Thr-Cys-Gly (WSPCSVTCG), present in three copies in region I of thrombospondin (TSP), six copies in properdin (P) and one copy in all the circumsporozoite (CS) proteins sequenced so far. TRAP also shares with certain extracellular glycoproteins, including TSP, the cell-recognition signal Arg-Gly-Asp (RGD), which has been shown to be crucial in the interaction of several extracellular glycoproteins with members of the integrin superfamily. Unlike the CS protein, TRAP is expressed during the erythrocytic stage of the parasite life cycle.

Amino Acid Sequence↗
PubMed1988

Plasmodium falciparum: the behavior of clinical isolates in an in vitro model of infected red blood cell sequestration.

An in vitro model of Plasmodium falciparum-infected red blood cell sequestration which uses C32 amelanotic melanoma cells as targets has been used to examine the binding capacity of infected red blood cells from subjects with naturally acquired P. falciparum infections of varying severity. The binding of infected red blood cells (IRBCs) to melanoma cells was specific to cells containing mature parasites. Variations in target cell density and in conditions of growth had significant effects on binding. Binding was pH dependent, being maximum at a pH of 6.9. Using standardized conditions the binding capacity of individual isolates of P. falciparum could be measured with a high degree of reproducibility. Binding capacity of IRBCs from 51 subjects between the ages of 6 months and 15 years varied between 12 and 1254 IRBCs per 100 melanoma cells when RBC suspensions at a 1% parasitemia and 4% hematocrit were used. Variation in binding was not related to the level of peripheral parasitemia of the isolate or to differences in adaptation to culture conditions. The binding capacity of parasitized cells from subjects with cerebral malaria did not differ from that of IRBCs from subjects with less serious clinical manifestations.

Adolescent↗
PubMed1988

Anti-sporozoite antibodies and immunity to malaria in a rural Gambian population.

A conserved repeated epitope, (NANP)3, of the circumsporozoite protein of Plasmodium falciparum has been identified previously as a putative target for artificially induced immunity to malaria. We examined the role of humoral responses to this epitope in acquired immunity to malaria in a rural African population. Seropositivity to (NANP)3 was slow to develop (9% positive in subjects aged 1-11 years; 88% in those of 30 years and above), and responses in younger subjects were transient. The poor response in younger subjects did not appear to be due to immunosuppression by concomitant blood stage parasitization. The relationship between levels of anti-(NANP)3 antibodies and parasitaemia changed from positive to negative with age. 126 subjects age 1-11 years were followed through an entire transmission season; those who were seropositive at the beginning ended the season with lower parasite rates (20% vs 59%) and experienced fewer episodes of clinical malaria (0.43 vs 0.67). However, the trend towards increasing susceptibility to clinical malaria in subjects entering the transmission season with lower levels of anti-(NANP)3 antibodies was modest, and combined cross-sectional and longitudinal data indicated that the humoral response to (NANP)3 did not play a major role in the development of immunity to clinical malaria in the population we studied.

Age Factors↗
PubMed1988

Design, synthesis, and properties of (4S)-7-(4-amino-2-substituted-pyrrolidin-1-yl)quinolone-3-carboxylic acids.

The quinolonecarboxylic acids constitute a class of extremely potent and orally active broad-spectrum antibacterial agents. These compounds have been shown to inhibit DNA gyrase, a key enzyme in bacterial DNA replication. The 7-(3-aminopyrrolidinyl)quinolone A-60969 (1) is a particularly potent member of this class and is currently undergoing clinical evaluation. We have studied a series of enantiomerically homogeneous (4S)-7-(4-amino-2-substituted-pyrrolidinyl)quinolones in an effort to utilize the 2-position of the pyrrolidine moiety to improve upon the solubility and pharmacokinetic properties of this class of compounds while still maintaining potent antibacterial activity. We have found that the absolute stereochemistry at the 2-position of the pyrrolidine ring is critical to the maintenance of such activity. In this paper, we report the full details of the asymmetric synthesis and the in vitro and in vivo structure-activity relationships of this series of compounds as well as the physiochemical properties, such as water solubility and log P, associated with the structural modifications. We also discuss the pharmacokinetic properties of several of these compounds in mice and the pharmacokinetics of 59, which has the best overall properties of agents in this study, in dog.

Animals↗
PubMed1988

Plasmodium falciparum glycolipid synthesis: constant and variant molecules of isolates and of strains with differing knob and cytoadherence phenotype.

Plasmodium falciparum-infected erythrocytes were metabolically labeled with tritiated glucosamine. Lipid extracts were analyzed by high-performance thin-layer chromatography to compare labeled molecules of eleven isolates from patients, six cytoadherent in vitro strains, and two knobbed and two knobless strains from Aotus monkeys. Up to nineteen labeled bands were identified. Glycolipid GL1, previously identified in Malayan Camp, was present in all isolates and strains. Other molecules, between CG and GM1 and between GM1 and GD1a, varied in mobility or presence. There was no apparent association between labeled molecules and the presence of knobs or the property of cytoadherence.

Animals↗
PubMed1987

Hypoglycaemia in African children with severe malaria.

Hypoglycaemia, defined as a plasma glucose concentration below 2.2 mmol/l, developed in 15 of 47 prospectively studied Gambian children with severe chloroquine-sensitive falciparum malaria. 5 of these hypoglycaemic children died compared with 1 in the normoglycaemic group (p = 0.02). In contrast to previous observations in quinine-treated adults, in whom hypoglycaemia was associated with hyperinsulinaemia, plasma concentrations of insulin were appropriately low and plasma ketones were high. Raised plasma concentrations of lactate and alanine suggested impairment of hepatic gluconeogenesis. In African children, hypoglycaemia is an important and treatable manifestation of severe malaria and is unrelated to antimalarial treatment.

Acute Disease↗
PubMed1987

Mortality and morbidity from malaria among children in a rural area of The Gambia, West Africa.

Mortality and morbidity from malaria were measured among 3000 children under the age of 7 years in a rural area of The Gambia, West Africa. Using a post-mortem questionnaire technique, malaria was identified as the probable cause of 4% of infant deaths and of 25% of deaths in children aged 1 to 4 years. The malaria mortality rate was 6.3 per 1000 per year in infants and 10.7 per 1000 per year in children aged 1 to 4 years. Morbidity surveys suggested that children under the age of 7 years experienced about one clinical episode of malaria per year. Calculation of attributable fractions showed that malaria may be responsible for about 40% of episodes of fever in children. Although the overall level of parasitaemia showed little seasonal variation, the clinical impact of malaria was highly seasonal; all malaria deaths and a high proportion of febrile episodes were recorded during a limited period at the end of the rainy season.

Age Factors↗
PubMed1987

Comparative analysis of the Plasmodium falciparum histidine-rich proteins HRP-I, HRP-II and HRP-III in malaria parasites of diverse origin.

Plasmodium falciparum-infected erythrocytes (IRBC) synthesize 3 histidine-rich proteins: HRP-I or the knob-associated HRP, HRP-II and HRP-III or SHARP. In order to distinguish these proteins immunochemically we prepared monoclonal antibodies which react with HRP-I, HRP-II and HRP-III, and rabbit antisera against synthetic peptides derived from the HRP-II and HRP-III sequences. A comparative analysis of diverse P. falciparum parasites was made using these antibodies and immunoprecipitation or Western blotting. HRP-I (Mr 80,000-115,000) was identified in all knob-positive P. falciparum parasites including isolates examined directly from Gambian patients. However, this protein was of lower abundance in these isolates and in 6 knob-positive, culture-adapted parasites compared to Aotus monkey-adapted parasites or culture-adapted parasites studied previously. HRP-II (Mr 60,000-105,000) was identified in all P. falciparum parasites regardless of knob-phenotype, and was recovered from culture supernatants as a secreted water-soluble protein. Within IRBC, HRP-II was found as a complex of several closely spaced bands. Cell surface radio-iodination of IRBC from several isolates and immunoprecipitation with a rabbit antiserum against the HRP-II repeat sequence identified HRP-II as a surface-exposed protein. Like HRP-I, the abundance of HRP-II was lower in the Gambian isolates than with Aotus monkey-adapted parasites studied earlier. Neither HRP-I nor HRP-II were identified in a knob-positive isolate of P. malariae collected from a Gambian patient. Analogues of these HRP were also absent from asexual parasites of diverse primate and murine malaria species screened with this panel of antibodies. HRP-III (Mr 40,000-55,000) was distinguished by its lower apparent size and by specific reaction with rabbit antibody against its 5-mer repeat sequence. HRP-III was of lowest abundance compared with the other two HRP. These antibody reagents and distinguishing properties should prove useful in studies on the separate functions of the 3 P. falciparum HRP.

Adult↗
PubMed1987

The accuracy of the clinical histories given by mothers of seriously ill African children.

The mothers of 87 Gambian children with a potentially fatal illness were interviewed at the time that their children were admitted to hospital and attempts were made to establish a diagnosis using the mothers' history alone. In 66 cases (76%) initial diagnoses corresponded to the diagnoses established finally by clinical and laboratory investigations. Diagnoses established at second interviews held with 51 mothers 1 month after their children had left hospital were accurate in 88% of cases. Gambian mothers can describe accurately a serious illness in their children and they can, therefore, be relied upon to give accurate information about an illness from which a child has died.

Adult↗
PubMed1987

Transport of an Mr approximately 300,000 Plasmodium falciparum protein (Pf EMP 2) from the intraerythrocytic asexual parasite to the cytoplasmic face of the host cell membrane.

The profound changes in the morphology, antigenicity, and functional properties of the host erythrocyte membrane induced by intraerythrocytic parasites of the human malaria Plasmodium falciparum are poorly understood at the molecular level. We have used mouse mAbs to identify a very large malarial protein (Mr approximately 300,000) that is exported from the parasite and deposited on the cytoplasmic face of the erythrocyte membrane. This protein is denoted P. falciparum erythrocyte membrane protein 2 (Pf EMP 2). The mAbs did not react with the surface of intact infected erythrocytes, nor was Pf EMP 2 accessible to exogenous proteases or lactoperoxidase-catalyzed radioiodination of intact cells. The mAbs also had no effect on in vitro cytoadherence of infected cells to the C32 amelanotic melanoma cell line. These properties distinguish Pf EMP 2 from Pf EMP 1, the cell surface malarial protein of similar size that is associated with the cytoadherent property of P. falciparum-infected erythrocytes. The mAbs did not react with Pf EMP 1. In one strain of parasite there was a significant difference in relative mobility of the 125I-surface-labeled Pf EMP 1 and the biosynthetically labeled Pf EMP 2, further distinguishing these proteins. By cryo-thin-section immunoelectron microscopy we identified organelles involved in the transit of Pf EMP through the erythrocyte cytoplasm to the internal face of the erythrocyte membrane where the protein is associated with electron-dense material under knobs. These results show that the intraerythrocytic malaria parasite has evolved a novel system for transporting malarial proteins beyond its own plasma membrane, through a vacuolar membrane and the host erythrocyte cytoplasm to the erythrocyte membrane, where they become membrane bound and presumably alter the properties of this membrane to the parasite's advantage.

Animals↗
PubMed1987

Ultrastructure of the erythrocytic stages of Plasmodium malariae.

This report describes the fine structure of the erythrocytic stages of Plasmodium malariae. Erythrocytic parasites from a naturally acquired human infection and an experimentally infected chimpanzee were morphologically indistinguishable and structurally similar to other primate malarias. New findings included observations of highly structured arrays of merozoite surface coat proteins in the cytoplasm of early schizonts and on the surface of budding merozoites and the presence of knobs in the membranes of Maurer's clefts. Morphological evidence is presented suggesting that proteins are transported between the erythrocyte surface and intracellular parasites via two routes: one associated with Maurer's clefts for transport of membrane-associated knob material and a second associated with caveolae in the host cell membrane for the import or export of host- or parasite-derived substances through the erythrocyte cytoplasm.

Adolescent↗
PubMed1987

Thrombospondin binding by parasitized erythrocyte isolates in falciparum malaria.

Toward understanding the pathogenesis of vascular sequestration in falciparum malaria, we investigated binding of Plasmodium falciparum parasitized erythrocyte isolates to thrombospondin and other adhesive proteins. Blood samples with rings from 12 patients with falciparum malaria were cultured 30 hr until parasites were mature trophozoites and schizonts. All parasitized erythrocyte isolates bound to thrombospondin, but not to fibronectin, laminin, vitronectin, or factor VIII/von Willebrand factor. Parasitized erythrocyte binding varied among isolates, ranging from 192 to 6,725 per mm2, average 2,953. There was good correlation between trophozoite plus schizont % parasitemia and thrombospondin binding (r = 0.884, P less than 0.001). In two patients with stupor, 3,642 and 2,864 parasitized erythrocytes bound per mm2, in proportion to parasitemia, suggesting cerebral malaria is not due to increased binding affinity. These results indicate there is a conserved function among isolates from this geographic region, known to be antigenically diverse at the parasitized erythrocyte membrane surface. These results support the hypothesis that specific binding to an endothelial receptor, possibly involving thrombospondin, plays a role in vascular sequestration in falciparum malaria.

Adult↗
PubMed1986

Bed-nets (mosquito-nets) and morbidity from malaria.

A study was undertaken in the Farafenni area of The Gambia to determine the relation between morbidity from malaria in children and the use of bed-nets (mosquito-nets). From comparisons of parasite and spleen rates in bed-net users and in non-users it seemed that bed-nets had a strong protective effect. However, the prevalence of malaria in the study population was also influenced by ethnic group and place of residence, and the association of bed-net use with these two confounding factors accounted for some of the differences observed between bed-net users and non-users. Nevertheless, a significant inverse correlation between splenomegaly and the use of bed-nets remained. This suggests that bed-nets give Gambian children some protection against malaria and that the use of bed-nets, either untreated or treated with an insecticide such as permethrin, should be investigated further as a means of malaria control in Africa.

Adolescent↗
PubMed1986

Parasite-infected-cell-agglutination and indirect immunofluorescence assays for detection of human serum antibodies bound to antigens on Plasmodium falciparum-infected erythrocytes.

Two methods are described for detecting the binding of serum antibodies from adults in an endemic malarious area (The Gambia) to surface antigens on Plasmodium falciparum-infected erythrocytes. An antibody-mediated parasite-infected-cell-agglutination assay (without secondary antibody) and an indirect immunofluorescence assay employing an anti-Fc secondary reagent were used to detect bound antibody. The surface of erythrocytes containing mature parasites bound antibody, but the surface of uninfected cells or cells containing early parasite stages did not react. Serum from 'non-immune' Europeans did not agglutinate infected erythrocytes, however, in the immunofluorescence test with anti-Ig and anti-F(ab')2 secondary reagents we could detect the binding of IgG antibody from 'non-immune' European serum to a small proportion of infected cells. In contrast to the results with freshly collected isolates, antibodies from sera of Gambian adults did not bind to the surface of infected cells from five different culture-adapted isolates of P. falciparum. These assays are suitable for studies on the antigenic diversity of erythrocyte antigens in natural infections and specific antibody responses to these antigens in infected patients.

Antibodies↗
PubMed1986

Antigens induced on erythrocytes by P. falciparum: expression of diverse and conserved determinants.

Red blood cells that are infected with the malaria parasite Plasmodium falciparum express new antigens on their surface. In a study of these antigens in the erythrocytes of naturally infected children in the Gambia, an antibody-mediated agglutination assay revealed an extreme degree of antigenic diversity. Serum samples from each of ten children in the convalescent stage of malaria infection reacted with infected cells from the same child but generally not with infected cells from the other children. The Gambian children's erythrocytes also expressed shared determinants: sera from Gambian adults often reacted with the surface of infected cells from all of the children and were shown by adsorption and elution experiments to contain antibodies that recognized several isolates. Conserved determinants exposed on infected erythrocytes may be important for development of antimalarial immunity either naturally or through vaccination.

Adult↗