Mechanical clearance of red blood cells by the human spleen: Potential therapeutic applications of a biomimetic RBC filtration method.

During their lifespan, circulating RBC are frequently checked for their deformability. This mechanical quality control operates essentially in the human spleen. RBC unable to squeeze though narrow splenic slits are retained and cleared from the blood circulation. Under physiological conditions this prevents microvessels from being clogged by senescent, rigid RBC. Retention of poorly deformable RBC is an important determinant of pathogenesis in malaria and may also impact the clinical benefit of transfusion. Modulating the splenic retention of RBC has already been proposed to support therapeutic approaches in these research fields. To this aim, the development of microplates for high throughput filtration of RBC through microsphere layers (microplate-based microsphiltration) has been undertaken. This review focuses on potential therapeutic applications provided by this technology in malaria chemotherapy and transfusion.

Travelers with cutaneous leishmaniasis cured without systemic therapy.

BACKGROUND: Cutaneous leishmaniasis (CL) is a disfiguring but not life-threatening disease. Because antileishmanial drugs are potentially toxic, the World Health Organization (WHO) recommends simple wound care or local therapy as first-line treatment, followed or replaced by systemic therapy if local therapy fails or cannot be performed. METHODS: To determine the feasibility and impact of the recommended approach, we analyzed the results of a centralized referral treatment program in 135 patients with parasitologically proven CL. RESULTS: Infections involved 10 Leishmania species and were contracted in 29 different countries. Eighty-four of 135 patients (62%) were initially treated without systemic therapy. Of 109 patients with evaluable charts, 23 of 25 (92%) treated with simple wound care and 37 of 47 (79%) treated with local antileishmanial therapy were cured by days 42-60. In 37 patients with large or complex lesions, or preexisting morbidities, or who had not been cured with local therapy, the cure rate with systemic antileishmanial agents was 60%. Systemic adverse events were observed in 15 patients, all receiving systemic therapy. CONCLUSIONS: In this population of CL patients displaying variable degrees of complexity and severity, almost two-thirds of patients could be initially managed without systemic therapy. Of these, 60 were cured before day 60. The WHO-recommended stepwise approach favoring initial local therapy therefore resulted in at least 44% of all patients being cured without exposure to the risk of systemic adverse events. Efforts are needed to further simplify local therapy of CL and to improve the management of patients with complex lesions and/or preexisting comorbidities.

[Cutaneous leishmaniasis in France: towards the end of injectable therapy?]. / La leishmaniose cutanée en France: vers la fin des traitements injectables?

Today no drug likely to be efficient on the majority of human-infecting species, well tolerated, and easy to administer is available for the treatment of human cutaneous leishmaniasis. But recent progress has been made. Efficient against visceral leishmaniasis, orally administered miltefosine may supplant pentavalent antimonials for the treatment of cutaneous leishmaniasis acquired in the New World. Right now, the reference treatment is still parenteral pentavalent antimonials 20 mg Sbv/kg/d for a duration that may probably be reduced from 20 to 10 days. The benefit/risk ratio of pentamidine still compares well with that of pentavalent antimonials for the treatment of lesions due to species belonging to the L. panamensis/L. guyanensis/L. shawi group. Pentamidine, which is easier to handle than antimonials, remains the reference treatment for cases from areas where these species predominate. Oral fluconazole is an improvement, readily available for cases from L. major foci. If its efficacy is confirmed in other foci and against other species, mechanisms will have to be implemented to make this therapeutic improvement affordable to poor patients in endemic countries. The development of an efficient and well tolerated topical treatment is still warranted. A new formulation of aminosidine is currently under evaluation. One can hope that the treatment of cutaneous leishmaniasis will soon become simpler, both for patients and doctors. For the benefits of this simplification to be rapidly affordable to all patients, the pharmaceutical and clinical research outlay must be maintained.

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.

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.

Antigenic variation at the surface of infected erythrocytes in plasmodium falciparum

Os membros da família dos genes var de Plasmodium falciparum codificam para receptores que desempenham um papel importante na patogenicidade da malária. O mecanismo responsável pela seleçäo da expressäo dos diferentes membros da família dos genes var ("switching") tem sido estdado utilizando populações de parasitas clonados, selecionados por suas características adesivas. O parasita expressa um único gene var o estágio de trofozoíto do seu ciclo de vida. Análises dos sítios de expressäo, ativos ou inativos, dos genes var demonstraram que o controle da expressäo ocorre durante a transcriçäo e a ativaçäo destes genes ocorre "in situ". Observamos que näo há sobreposiçäo no repertório dos genes var para diferentes isolados de laboratório, sugerindo desta maneira a existência de mecanismos para a geraçäo de diversidade desta família gênica. Experimentos de "fluorescence in situ hybridization" (FISH) mostraram que as extremidades dos cromossomos de P. falciparum estäo fisicamente associados e que esta formaçäo é importante para a geraçäo da diversidade dos genes var.

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.

[First case of cutaneous leishmaniasis from Leishmania infantum in Corsica]. / Premier cas de leishmaniose cutanée à Leishmania infantum en Corse.

Canine leishmaniasis is endemic in Corsica. Sporadic cases of visceral leishmaniasis due to Leishmania infantum have been reported in humans, but no case of cutaneous leishmaniasis has been reported to date. In August 1994, a 42-year-old woman living in Orleans (France) presented with two nodular lesions on the face. These lesions appeared two months after a stay near the Gulf of Ajaccio in Corsica. Histopathological examination revealed intracellular amastigotes. The culture isolate was identified as Leishmania infantum zymodeme MON-29. Epidemiological data and climatic conditions prevailing during the patient's stay suggest that contamination took place in Corsica.

Therapeutic effect of reference antileishmanial agents in murine visceral leishmaniasis due to Leishmania infantum.

A sensitive, culture-based, microtitration technique has recently been developed for determining parasite burdens in organs recovered from Balb/c mice infected with Leishmania infantum. In the present study, this technique was used to examine the efficacy of three, first-line, antileishmanial agents in reducing parasite burdens and eradicating parasites from target organs in mice. Treatment with meglumine antimoniate (50 mg SbV/kg.day) significantly reduced the parasite burdens in the livers and lungs (by about 10-fold and > 100-fold, respectively) but not those in the spleens. Although use of a higher dose of meglumine antimoniate (200 mg SbV/kg.day) resulted in an even more dramatic reduction in the parasite burdens in the livers, it had no significant effect on the burdens in the spleens. Treatment with amphotericin B (0.8 mg/kg every other day) resulted in significant reductions in the parasite burdens in the livers, spleens and lungs of infected mice. Although low doses of aminosidine (20 mg/kg.day) had no effect, high doses (200 mg/kg.day) resulted in undetectable parasite burdens in the livers, for at least 100 days post-treatment, and marked reductions in burdens in the spleens. These results are consistent with previous data from studies using animal models of visceral leishmaniasis. Thanks to the sensitivity of the technique, culture microtitration revealed that none of the drug schedules achieved the elimination of all parasites in all target organs. The murine model used mimics some important features of HIV/Leishmania infantum co-infections in humans.

Culture microtitration: a sensitive method for quantifying Leishmania infantum in tissues of infected mice.

We developed a microtitration method to determine the parasite burdens in homogenized organs of mice infected with Leishmania infantum. This method proved more sensitive than direct enumeration of amastigotes in stained organs, was appropriate for describing the kinetics of infection, and can be considered for physiopathological or pharmaceutical experimental studies.