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J Gysin

Publications and source records attributed to J Gysin.

At least 19 recordsLinked to original sources
PubMed2006

Cytoadhesion of Plasmodium falciparum-infected erythrocytes and the infected placenta: a two-way pathway.

Malaria is undoubtedly the world's most devastating parasitic disease, affecting 300 to 500 million people every year. Some cases of Plasmodium falciparum infection progress to the deadly forms of the disease responsible for 1 to 3 million deaths annually. P. falciparum-infected erythrocytes adhere to host receptors in the deep microvasculature of several organs. The cytoadhesion of infected erythrocytes to placental syncytiotrophoblast receptors leads to pregnancy-associated malaria (PAM). This specific maternal-fetal syndrome causes maternal anemia, low birth weight and the death of 62,000 to 363,000 infants per year in sub-Saharan Africa, and thus has a poor outcome for both mother and fetus. However, PAM and non-PAM parasites have been shown to differ antigenically and genetically. After multiple pregnancies, women from different geographical areas develop adhesion-blocking antibodies that protect against placental parasitemia and clinical symptoms of PAM. The recent description of a new parasite ligand encoded by the var2CSA gene as the only gene up-regulated in PAM parasites renders the development of an anti-PAM vaccine more feasible. The search for a vaccine to prevent P. falciparum sequestration in the placenta by eliciting adhesion-blocking antibodies and a cellular immune response, and the development of new methods for evaluating such antibodies should be key priorities in mother-child health programs in areas of endemic malaria. This review summarizes the main molecular, immunological and physiopathological aspects of PAM, including findings related to new targets in the P. falciparum var gene family. Finally, we focus on a new methodology for mimicking cytoadhesion under blood flow conditions in human placental tissue.

Animals↗
PubMed2001

Role of nonimmune IgG bound to PfEMP1 in placental malaria.

Infections with Plasmodium falciparum during pregnancy lead to the accumulation of parasitized red blood cells (infected erythrocytes, IEs) in the placenta. IEs of P. falciparum isolates that infect the human placenta were found to bind immunoglobulin G (IgG). A strain of P. falciparum cloned for IgG binding adhered massively to placental syncytiotrophoblasts in a pattern similar to that of natural infections. Adherence was inhibited by IgG-binding proteins, but not by glycosaminoglycans or enzymatic digestion of chondroitin sulfate A or hyaluronic acid. Normal, nonimmune IgG that is bound to a duffy binding-like domain beta of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) might at the IE surface act as a bridge to neonatal Fc receptors of the placenta.

Animals↗
PubMed2001

Molecular mechanisms of Plasmodium falciparum placental adhesion.

In natural Plasmodium falciparum infections, parasitized erythrocytes (PEs) circulate in the peripheral blood for a period corresponding roughly to the first part of the erythrocytic life cycle (ring stage). Later, in blood-stage development, parasite-encoded adhesion molecules are inserted into the erythrocyte membrane, preventing the circulation of the PEs. The principal molecule mediating PE adhesion is P. falciparum erythrocyte membrane protein 1 (PfEMP1), encoded by the polymorphic var gene family. The population of parasites is subject to clonal antigenic variation through changes in var expression, and a single PfEMP1 variant is expressed at the PE surface in a mutually exclusive manner. In addition to its role in immune evasion, switches in PfEMP1 expression may be associated with fundamental changes in parasite tissue tropism in malaria patients. A switch from CD36 binding to chondroitin sulphate A (CSA) binding may lead to extensive sequestration of PEs in placenta syncytiotrophoblasts. This is probably a key event in malaria pathogenesis during pregnancy. The CSA-binding phenotype of mature PEs is linked to another distinct adhesive phenotype: the recently described CSA-independent cytoadhesion of ring-stage PEs. Thus, a subpopulation of PEs that sequentially displays these two different phenotypes may bind to an individual endothelial cell or syncytiotrophoblast throughout the asexual blood-stage cycle. This suggests that non-circulating (cryptic) parasite subpopulations are present in malaria patients.

Animals↗
PubMed2000

Characterisation of the chondroitin sulphate of Saimiri brain microvascular endothelial cells involved in Plasmodium falciparum cytoadhesion.

Cytoadhesion of Plasmodium falciparum-infected erythrocytes (IRBC) to chondroitin-4-sulphate (CSA) is inhibited by soluble CSA in vitro on Saimiri brain microvascular endothelial cells (SBEC) and in vivo in P. falciparum-infected Saimiri monkeys. We tested whether the SBEC model was appropriate for studying CSA-binding IRBC using four cell lines. All SBEC expressed a chondroitin sulphate (CS), with a composition of CSA. The mean sizes of these CSA were 20.5, 22, 23, 32.5 and 36 kDa for SBEC 3A and C2, CHO, SBEC 1D and 17, respectively. We found that cytoadhesion of the Palo-Alto (FUP)1 CSA-binding phenotype, selected by panning on SBEC 17, was specifically inhibited in a dose-dependent manner by all the purified CSA. The extent of inhibition depended on the cellular origin of the tested CSA. SBEC 17 CSA was 33 times more efficient than CHO-CSA and 21 times more efficient than the 50 kDa commercial bovine trachaea CSA. Dynabeads coated with a total extract of SBEC 1D CS-proteoglycans interacted with CSA- but not with CD36- or ICAM-1-binding IRBC. These Dynabeads also interacted specifically with the PfEMP1 DBL-3 domain, on the surface of CHO transfectants, but not with the CIDR-1 domain. Thrombomodulin was involved in IRBC adhesion to all SBEC whereas CD44 was only expressed by SBEC 1D and 17. These two CSA-proteoglycans have also been detected at the surface of human endothelial cells. Thus, the two homologous models, SBEC/Saimiri sciureus, are useful and reliable tools for the evaluation of new anti-CSA adhesion treatments and anti-disease vaccines for pregnant women.

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PubMed2000

Recombinant human thrombomodulin(csa+): a tool for analyzing Plasmodium falciparum adhesion to chondroitin-4-sulfate.

The proteoglycan thrombomodulin has been shown to be involved, via its chondroitin-sulfate moiety, in the cytoadhesion of chondroitin-4-sulfate-binding-Plasmodium falciparum-infected erythrocytes to endothelial cells and syncytiotrophoblasts. We cloned and expressed in CHO and COS-7 cells a gene encoding soluble human recombinant thrombomodulin, with a chondroitin-4-sulfate moiety. This system is complementary to the in vitro cell models currently used to study the chondroitin-4-sulfate-binding phenotype. It also provides a means of overcoming the lack of specificity observed in interactions of infected erythrocytes with modified chondroitin-4-sulfate. This thrombomodulin displayed normal activity in coagulation, indicating that it was in a functional conformation. The recombinant protein, whether produced in CHO or COS-7 cells, inhibited cytoadhesion to Saimiri brain microvascular endothelial cells 1D infected with Palo-Alto(FUP)1 parasites selected for chondroitin-4-sulfate receptor preference. Thus, the recombinant protein was produced with a chondroitin-sulfate moiety, identified as a chondroitin-4-sulfate, in both cell types. In both cases, the recombinant protein bound to the chondroitin-4-sulfate phenotype, but not to CD36- and ICAM-1-binding parasites. The chondroitin-4-sulfate was 36 kDa in size for CHO and 17.5 kDa for COS-7 cells. There was, however, no difference in the capacities of the recombinant proteins produced by the two cell types to inhibit the cytoadhesion of infected erythrocytes. Thrombomodulin immobilized on plastic or coupled to Dynabeads was used to purify specifically the infected erythrocytes that bind to chondroitin-4-sulfate. These infected erythrocytes were cultured to establish parasite lines of this phenotype. We then showed that the thrombomodulin, labeled with FITC, could be used to detect this phenotype in blood samples. Finally, the direct binding of infected erythrocytes to immobilized thrombomodulin was used to screen for anti-chondroitin-4-sulfate-binding antibodies.

Animals↗
PubMed2000

Cytoadhesion of Plasmodium falciparum ring-stage-infected erythrocytes.

A common pathological characteristic of Plasmodium falciparum infection is the cytoadhesion of mature-stage-infected erythrocytes (IE) to host endothelium and syncytiotrophoblasts. Massive accumulation of IE in the brain microvasculature or placenta is strongly correlated with severe forms of malaria. Extensive binding of IE to placental chondroitin sulfate A (CSA) is associated with physiopathology during pregnancy. The adhesive phenotype of IE correlates with the appearance of Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) at the erythrocyte surface (approximately 16 h after merozoite invasion), so that only early blood-stage (ring-stage) IE appear in the peripheral blood. Here, we describe results that challenge the existing view of blood-stage IE biology by demonstrating the specific adhesion of IE, during the early ring-stage, to endothelial cell lines from the brain and lung and to placental syncytiotrophoblasts. Later, during blood-stage development of these IE, trophozoites switch to an exclusively CSA cytoadhesion phenotype. Therefore, adhesion to an individual endothelial cell or syncytiotrophoblast may occur throughout the blood-stage cycle, indicating the presence in malaria patients of noncirculating (cryptic) parasite subpopulations. We detected two previously unknown parasite proteins on the surface of ring-stage IE. These proteins disappear shortly after the start of PfEMP1-mediated adhesion.

Adult↗
PubMed1999

Plasmodium falciparum domain mediating adhesion to chondroitin sulfate A: a receptor for human placental infection.

Malaria during the first pregnancy causes a high rate of fetal and neonatal death. The decreasing susceptibility during subsequent pregnancies correlates with acquisition of antibodies that block binding of infected red cells to chondroitin sulfate A (CSA), a receptor for parasites in the placenta. Here we identify a domain within a particular Plasmodium falciparum erythrocyte membrane protein 1 that binds CSA. We cloned a var gene expressed in CSA-binding parasitized red blood cells (PRBCs). The gene had eight receptor-like domains, each of which was expressed on the surface of Chinese hamster ovary cells and was tested for CSA binding. CSA linked to biotin used as a probe demonstrated that two Duffy-binding-like (DBL) domains (DBL3 and DBL7) bound CSA. DBL7, but not DBL3, also bound chondroitin sulfate C (CSC) linked to biotin, a negatively charged sugar that does not support PRBC adhesion. Furthermore, CSA, but not CSC, blocked the interaction with DBL3; both CSA and CSC blocked binding to DBL7. Thus, only the DBL3 domain displays the same binding specificity as PRBCs. Because protective antibodies present after pregnancy block binding to CSA of parasites from different parts of the world, DBL-3, although variant, may induce cross-reactive immunity that will protect pregnant women and their fetuses.

Animals↗
PubMed1999

Ex vivo desequestration of Plasmodium falciparum-infected erythrocytes from human placenta by chondroitin sulfate A.

We performed ex vivo experiments with Plasmodium falciparum-infected human placentas from primi- and multigravida women from Cameroon. All women, independent of their gravida status, had anti-chondroitin sulfate A (CSA) adhesion antibodies which cross-reacted with heterologous strains, such as FCR3 and Palo-Alto(FUP)1, which were selected for CSA binding. These antibodies, directed against the surface of infected erythrocytes obtained by flushing with CSA (IRBC(CSA)), were restricted to the immunoglobulin G3 isotypes. Massive desequestration of parasites was achieved with soluble CSA but not with anti-ICAM-1 and anti-CD36 monoclonal antibodies. All of the CSA-flushed parasites were analyzed immediately by using in vitro assays of binding to Saimiri brain endothelial cells (SBEC) expressing various adhesion receptors. Parasites derived from all six placentas displayed the CSA adhesion phenotype. However, only partial inhibition of adhesion was observed in the presence of soluble CSA or when Sc1D SBEC were treated with chondroitinase ABC. These results suggest that an additional adhesive molecule of IRBC(CSA) which binds to an unidentified receptor is present in the placenta. This new phenotype was lost once the parasites adapted to in vitro culture. We observed additional differences in the CSA adhesion phenotype between placental parasites and in vitro-cultured parasites panned on endothelial cells carrying CSA. The minimum size of fractionated CSA required for a significant inhibition of placental IRBC(CSA) adhesion to Sc1D cells was 1 to 2 kDa, which contrasts with the 4-kDa size necessary to reach equivalent levels of inhibition with panned IRBC(CSA) of this phenotype. All placental IRBC(CSA) cytoadhered to Sc17 SBEC, which express only the CSA receptor. Panning of IRBC(CSA) on these cells resulted in a significant quantitative increase of IRBC cytoadhering to the CSA of Sc1D cells but did not change their capacity for adhesion to CSA on normal placenta cryosections. Our results indicate that the CSA binding phenotype is heterogeneous and that several distinct genes may encode P. falciparum-CSA ligands with distinct binding properties.

Animals↗
PubMed1999

Plasmodium falciparum-infected erythrocytes: a mutational analysis of cytoadherence via murine thrombomodulin.

The pathophysiologic events leading to organ damage in Plasmodium falciparum malaria infections involve adhesion and sequestration of parasite-infected erythrocytes (PRBC) to the vascular endothelium and syncytiotrophoblast. Several potential receptors to which the PRBCs may bind have recently been identified, one of which is thrombomodulin (TM). TM has been implicated particularly in mediating sequestration of P. falciparum-infected erythrocytes in the placenta and brain, two sites of disease associated with high morbidity. In order to establish that binding of parasite-infected red blood cells to TM is dependent on its containing chondroitin-4-sulfate (CSA), we have mutated the CSA-attachment site of murine TM, and expressed this mutant form (TMsergly) in COS-7 cells. In cytoadhesion assays, we demonstrate that, in contrast to wild-type TM which contains CSA and supports the adhesion of 1466 PRBCs/mm2, TMser-gly does not contain CSA and adhesion of PRBCs to those cells expressing TMser-gly is entirely abrogated (200 PRBCs/mm2). These studies further confirm that the CSA of TM may play a role in the pathophysiology of malaria by providing a binding site for PRBCs.

Animals↗
PubMed1998

Antigenic variation in malaria: in situ switching, relaxed and mutually exclusive transcription of var genes during intra-erythrocytic development in Plasmodium falciparum.

Members of the Plasmodium falciparum var gene family encode clonally variant adhesins, which play an important role in the pathogenicity of tropical malaria. Here we employ a selective panning protocol to generate isogenic P.falciparum populations with defined adhesive phenotypes for CD36, ICAM-1 and CSA, expressing single and distinct var gene variants. This technique has established the framework for examining var gene expression, its regulation and switching. It was found that var gene switching occurs in situ. Ubiquitous transcription of all var gene variants appears to occur in early ring stages. However, var gene expression is tightly regulated in trophozoites and is exerted through a silencing mechanism. Transcriptional control is mutually exclusive in parasites that express defined adhesive phenotypes. In situ var gene switching is apparently mediated at the level of transcriptional initiation, as demonstrated by nuclear run-on analyses. Our results suggest that an epigenetic mechanism(s) is involved in var gene regulation.

ADP-ribosyl Cyclase↗
PubMed1998

Biological and biochemical characteristics of cytoadhesion of Plasmodium falciparum-infected erythrocytes to chondroitin-4-sulfate.

The cytoadhesion of Plasmodium falciparum laboratory strains and clones to Saimiri brain microvascular endothelial cells (SBEC 17), with chondroitin-4-sulfate (CSA) as the only adhesion receptor, was tested. Only one strain had significant cytoadhesion. However, CSA-specific infected erythrocytes (IRBCs) were detected in all strains after selection of a CSA-specific subpopulation by culturing the few adherent IRBCs. This demonstrates the lack of sensitivity of cytoadhesion microassays for detecting small quantities of CSA-specific IRBCs in cultures or field isolates. Cytoadhesion to CSA is maximal at 24 h of the cycle and decreases with the onset of schizogony, reaching a minimum just before reinvasion. This fluctuation must be taken into account in comparisons of the cytoadhesion of different strains or isolates. The minimum size of CSA for active inhibition was 4 kDa, and a mass of 9 kDa was required for inhibition similar to that obtained with the 50-kDa CSA. In contrast to cytoadhesion to CSA, which is pH independent or maximal at physiological pH (depending on the target endothelial cells), adhesion to CD36 and intercellular adhesion molecule 1 was pH dependent, requiring acidic conditions to be maximal in all cases. Cytoadhesion to CSA may trigger the occlusion of microvessels and cause the acidosis necessary for the other receptors to be fully efficient. If this key role in the mechanisms of sequestration were to be confirmed in vivo, prevalence studies of the CSA cytoadhesion phenotype would have to be reevaluated, because simple cytoadhesion assays do not detect CSA-specific parasites present in very low numbers, and these parasites might then be undetected in the peripheral blood but present in organs in which sequestration occurs, such as the placenta (M. Fried and P. E. Duffy, Science 272:1502-1504, 1996).

Animals↗
PubMed1998

[Plasmodium falciparum and chondroitin-4-sulfate: the new key couple in sequestration].

Some complications of Plasmodium falciparum infection such as cerebral malaria and pregnancy-associated malaria may be partially due to cytoadherence of erythrocytes infected by mature parasites on microvascular endothelial cells or placental syncytiotrophoblasts. Recently a new cytoadherence receptor, chondroitin-4-sulphate (CSA), was identified first on endothelial cells in primates and then on CHO cells and purified receptors. Further study has implicated CSA in cytoadherence of infected red blood cells to syncytiotrophoblasts in human placenta and Saimiri sciureus monkeys. In solution the minimal size for full inhibitory effect is approximately 9 kDa. Injection of CSA in Plasmodium falciparum-infected Saimiri monkeys resulted in specific release of sequestered erythrocytes infected by mature parasites. An added interest of these findings is that CSA, a glycosaminoglycan, is already in clinical use for treatment of degenerative joint disease. Current data on the parasite ligand for CSA indicates that it is not co-expressed with other cytoadherence ligands and that its binding activity decreases as the parasite matures from the 20th to 40th hour of the cycle. Since one or more var genes encoding the CSA ligand have been identified, it is likely that peptides will be obtained quickly and used either for direct inhibition of cytoadherence on CSA or for development of an anti-sequestration vaccine.

Animals↗
PubMed1997

Presence of the parasitophorous duct in Plasmodium falciparum and P. vivax parasitized Saimiri monkey red blood cells.

Although the exchange of metabolites between the intraerythrocytic malaria parasite and the external medium has been studied extensively, the transport of molecules across the erythrocyte cytoplasmic membrane and cytoplasm and the parasitophorous vacuolar membrane needs to be investigated more fully to be completely understood. Recently, the concept of the parasitophorous duct, establishing a continuity between the environment and the vacuolar space surrounding the intraerythrocytic parasite, has been suggested to provide an explanation of how macromolecules can cross two membranes in a cell devoid of an endocytic system. This concept is highly controversial and has been suspected to be an in vitro artefact. In this article, Bruno Pouvelle and Jürg Gysin present evidence of the existence of the parasitophorous duct in Saimiri monkey Plasmodium falciparum- and P. vivax-infected erythrocytes, with a series of ex vivo experiments showing stage and species dependent variations of the characteristics of this structure.

Journal Article↗
PubMed1997

Immunogenicity and efficacy trials with Plasmodium falciparum recombinant antigens identified as targets of opsonizing antibodies in the naive squirrel monkey Saimiri sciureus.

We have previously shown that Plasmodium falciparum recombinant antigens PfEB200, R23, and Pfi72 consistently inhibit opsonization of infected red blood cells by protective hyperimmune Saimiri sera, indicating that they present target epitopes involved in the phagocytosis of infected red blood cells. We report here an analysis of the immune response elicited in naive squirrel monkeys injected with the individual recombinant antigens or with a mixture of the three antigens combined with a synthetic peptide. In the three administration protocols investigated, there was no evidence for the production of antibody contributing to the phagocytosis of infected red blood cells, contrasting with the increase of opsonizing antibodies elicited by these antigens in monkeys with a prior (> or = 500 days) experience with malaria infection. However, the recombinant antigens were highly immunogenic, inducing specific antibody responses to P. falciparum and to the recombinant antigens. When the monkeys immunized with the antigen combination were challenged with blood-stage parasites, there was substantial protection: three of seven immunized animals self-cured and two others experienced a delayed peak of parasitemia. Taken together with our previous findings, these results suggest that PfEB200, R23, and Pfi72 constitute interesting vaccine candidates, and show that the presence of antibodies promoting phagocytosis of infected red blood cells is not a prerequisite for protection after immunization with these antigens in the Saimiri model.

Animals↗
PubMed1997

Chondroitin-4-sulfate impairs in vitro and in vivo cytoadherence of Plasmodium falciparum infected erythrocytes.

BACKGROUND: Chondroitin-4-sulfate (CSA) was recently described as a Plasmodium falciparum cytoadherence receptor present on Saimiri brain microvascular and human lung endothelial cells. MATERIALS AND METHODS: To specifically study chondroitin-4-sulfate-mediated cytoadherence, a parasite population was selected through panning of the Palo-Alto (FUP) 1 P. falciparum isolate on monolayers of Saimiri brain microvascular endothelial cells (SBEC). Immunofluorescence showed this SBEC cell line to be unique for its expression of CSA-proteoglycans, namely CD44 and thrombomodulin, in the absence of CD36 and ICAM-1. RESULTS: The selected parasite population was used to monitor cytoadherence inhibition/dissociating activities in Saimiri sera collected at different times after intramuscular injection of 50 mg CSA/kg of body weight. Serum inhibitory activity was detectable 30 min after injection and persisted for 8 hr. Furthermore, when chondroitin-4-sulfate was injected into monkeys infected with Palo-Alto (FUP) 1 P. falciparum, erythrocytes containing P. falciparum mature forms were released into the circulation. The cytoadherence phenotype of circulating infected red blood cells (IRBC) was determined before and 8 hr after inoculation of CSA. Before inoculation, in vitro cytoadherence of IRBCs was not inhibited by CSA. In contrast, in vitro cytoadherence of circulating infected erythrocytes obtained 8 hr after CSA inoculation was inhibited by more than 90% by CSA. CONCLUSIONS: In the squirrel monkey model for infection with P. falciparum, chondroitin-4-sulfate impairs in vitro and in vivo cytoadherence of parasitized erythrocytes.

Animals↗