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Red cell clearance in Malaria

Red blood cells are vital for tissue homeostasis, display multiple blood group systems, and play key roles in immunity, gas transport, vasodilation, detoxification, and metabolism.  We explore the pathophysiological mechanisms of red blood cell alteration and elimination in malaria. In the context of a long and fruitful collaboration between Indonesian-Australian colleagues and BioTiGR team members, our group has contributed to show that red blood cells infected by P. falciparum and P. vivax accumulate in the spleen of asymptomatic subjects in endemic areas (Kho, Qotrunnada et al. 2021) In close collaboration with the French National Reference Center for Malaria, we have confirmed that pitting is a major mechanism of parasite clearance (Ndour, Lopera-Mesa et al. 2015, Wojnarski, Mouri et al. 2019) and is tightly associated with anemia following treatment of malaria with artemisinin derivatives Jaureguiberry, Ndour et al. 2014). The quantification of pitted RBC enables the prediction of post-artemisinin delayed anemia (Ndour, Larreche et al. 2017). Ongoing clinical studies explore immunological and mechanical processes leading to the premature and excessive clearance of red blood cell in patients before and after treatment with antimalarial drugs (Wojnarski, Mouri et al. 2019). Several tools mimicking the spleen sensing and filtering function such as microsphiltration, microfluidic, mice model or ektacytometry (Duez, Holleran et al. 2015, Picot, Ndour et al. 2015) lead us to contribute to the development of innovative assays for gametocytes transmission blocking strategies and to explore the slow clearance of artemisinin resistant parasites.

Team Members

Papa Alioune Ndour (team leader)

Aissatou Bailo Diallo

Valentine Carret

Souheila Ben-Moumen

Research Topics

A biomimetic microfluidic chip to study the circulation and mechanical retention of red blood cells in the spleen.
A biomimetic microfluidic chip to study the circulation and mechanical retention of red blood cells in the spleen.

Picot Julien, Ndour Papa Alioune, Lefevre Sophie D, El Nemer Wassim, Tawfik Harvey, Galimand Julie, Da Costa Lydie, Ribeil Jean-Antoine, de Montalembert Mariane, Brousse Valentine, Le Pioufle Bruno, Buffet Pierre, Le Van Kim Caroline, Français Olivier
American journal of hematology, 90 (2015) 

Red blood cells (RBCs) are deformable and flow through vessels narrower than their own size. Their deformability is most stringently challenged when they cross micrometer-wide slits in the spleen. In several inherited or acquired RBC disorders, blockade of small vessels by stiff RBCs can trigger organ damage, but a functional spleen is expected to clear these abnormal RBCs from the circulation before they induce such complications. We analyzed flow behavior of RBCs in a microfluidic chip that replicates the mechanical constraints imposed on RBCs as they cross the human spleen. Polymer microchannels obtained by soft lithography with a hydraulic diameter of 25 μm drove flow into mechanical filtering units where RBCs flew either slowly through 5- to 2-μm-wide slits or rapidly along 10-μm-wide channels, these parallel paths mimicking the splenic microcirculation. Stiff heated RBCs accumulated in narrow slits seven times more frequently than normal RBCs infused simultaneously. Stage-dependent retention of Plasmodium falciparum-infected RBCs was also observed in these slits. We also analyzed RBCs from patients with hereditary spherocytosis and observed retention for those having the most altered mechanical properties as determined by ektacytometry. Thus, in keeping with previous observations in vivo and ex vivo, the chip successfully discriminated poorly deformable RBCs based on their distinct mechanical properties and on the intensity of the cell alteration. Applications to the exploration of the pathogenesis of malaria, hereditary spherocytosis, sickle cell disease and other RBC disorders are envisioned.

American journal of hematology, 2015, vol.90, p.

Picot Julien, Ndour Papa Alioune, Lefevre Sophie D, El Nemer Wassim, Tawfik Harvey, Galimand Julie, Da Costa Lydie, Ribeil Jean-Antoine, de Montalembert Mariane, Brousse Valentine, Le Pioufle Bruno, Buffet Pierre, Le Van Kim Caroline, Français Olivier

Hidden Biomass of Intact Malaria Parasites in the Human Spleen.
Hidden Biomass of Intact Malaria Parasites in the Human Spleen.

Kho Steven, Qotrunnada Labibah, Leonardo Leo, Andries Benediktus, Wardani Putu A I, Fricot Aurelie, Henry Benoit, Hardy David, Margyaningsih Nur I, Apriyanti Dwi, Puspitasari Agatha M, Prayoga Pak, Trianty Leily, Kenangalem Enny, Chretien Fabrice, Safeukui Innocent, Del Portillo Hernando A, Fernandez-Becerra Carmen, Meibalan Elamaran, Marti Matthias, Price Ric N, Woodberry Tonia, Ndour Papa A, Russell Bruce M, Yeo Tsin W, Minigo Gabriela, Noviyanti Rintis, Poespoprodjo Jeanne R, Siregar Nurjati C, Buffet Pierre A*, Anstey Nicholas M*
The New England journal of medicine, 384 (2021) 

The New England journal of medicine, 2021, vol.384, p.

Kho Steven, Qotrunnada Labibah, Leonardo Leo, Andries Benediktus, Wardani Putu A I, Fricot Aurelie, Henry Benoit, Hardy David, Margyaningsih Nur I, Apriyanti Dwi, Puspitasari Agatha M, Prayoga Pak, Trianty Leily, Kenangalem Enny, Chretien Fabrice, Safeukui Innocent, Del Portillo Hernando A, Fernandez-Becerra Carmen, Meibalan Elamaran, Marti Matthias, Price Ric N, Woodberry Tonia, Ndour Papa A, Russell Bruce M, Yeo Tsin W, Minigo Gabriela, Noviyanti Rintis, Poespoprodjo Jeanne R, Siregar Nurjati C, Buffet Pierre A*, Anstey Nicholas M*

Measuring the HRP2 protein in blood from artesunate-treated malaria patients predicts post-artesunate delayed hemolysis.
Measuring the HRP2 protein in blood from artesunate-treated malaria patients predicts post-artesunate delayed hemolysis.

Ndour Papa Alioune, Larréché Sébastien, Mouri Oussama, Argy Nicolas, Gay Frédérick, Roussel Camille, Jauréguiberry Stéphane, Perillaud Claire, Langui Dominique, Biligui Sylvestre, Chartrel Nathalie, Mérens Audrey, Kendjo Eric, Ghose Aniruddha, Hassan Md Mahtab Uddin, Hossain Md Amir, Kingston Hugh W F, Plewes Katherine, Dondorp Arjen M, Danis Martin, Houzé Sandrine, Bonnefoy Serge, Thellier Marc, Woodrow Charles J, Buffet Pierre A
Science translational medicine, 9 (2017) 

Artesunate, the recommended drug for severe malaria, rapidly clears the malaria parasite from infected patients but frequently induces anemia-called post-artesunate delayed hemolysis (PADH)-for which a simple predictive test is urgently needed. The underlying event in PADH is the expulsion of artesunate-exposed parasites from their host erythrocytes by pitting. We show that the histidine-rich protein 2 (HRP2) of the malaria parasite persists in the circulation of artesunate-treated malaria patients in Bangladesh and in French travelers who became infected with malaria in Africa. HRP2 persisted in whole blood (not plasma) of artesunate-treated patients with malaria at higher levels compared to quinine-treated patients. Using an optimized membrane permeabilization method, HRP2 was observed by immunofluorescence, Western blotting, and electron microscopy to persist in once-infected red blood cells from artesunate-treated malaria patients. HRP2 was deposited at the membrane of once-infected red blood cells in a pattern similar to that for ring erythrocyte surface antigen (RESA), a parasite invasion marker. On the basis of these observations, we developed a semiquantitative titration method using a widely available HRP2-based rapid diagnostic dipstick test. Positivity on this test using a 1:500 dilution of whole blood from artesunate-treated patients with malaria collected shortly after parasite clearance predicted subsequent PADH with 89% sensitivity and 73% specificity. These results suggest that adapting an existing HRP2-based rapid diagnostic dipstick test may enable prediction of PADH several days before it occurs in artesunate-treated patients with malaria.

Science translational medicine, 2017, vol.9, p.

Ndour Papa Alioune, Larréché Sébastien, Mouri Oussama, Argy Nicolas, Gay Frédérick, Roussel Camille, Jauréguiberry Stéphane, Perillaud Claire, Langui Dominique, Biligui Sylvestre, Chartrel Nathalie, Mérens Audrey, Kendjo Eric, Ghose Aniruddha, Hassan Md Mahtab Uddin, Hossain Md Amir, Kingston Hugh W F, Plewes Katherine, Dondorp Arjen M, Danis Martin, Houzé Sandrine, Bonnefoy Serge, Thellier Marc, Woodrow Charles J, Buffet Pierre A

Postartesunate delayed hemolysis is a predictable event related to the lifesaving effect of artemisinins.
Postartesunate delayed hemolysis is a predictable event related to the lifesaving effect of artemisinins.

Jauréguiberry Stéphane, Ndour Papa A, Roussel Camille, Ader Flavie, Safeukui Innocent, Nguyen Marie, Biligui Sylvestre, Ciceron Liliane, Mouri Oussama, Kendjo Eric, Bricaire François, Vray Muriel, Angoulvant Adéla, Mayaux Julien, Haldar Kasturi, Mazier Dominique, Danis Martin, Caumes Eric, Thellier Marc, Buffet Pierre
Blood, 124 (2014) 

Patients with severe malaria treated with artesunate sometimes experience a delayed hemolytic episode. Artesunate (AS) induces pitting, a splenic process whereby dead parasites are expelled from their host erythrocytes. These once-infected erythrocytes then return to the circulation. We analyzed hematologic parameters in 123 travelers treated with AS for severe malaria. Among 60 nontransfused patients observed for more than 8 days, 13 (22%) had delayed hemolysis. The peak concentration of circulating once-infected erythrocytes was measured during the first week in 21 patients and was significantly higher in 9 patients with delayed hemolysis than in 12 with other patterns of anemia (0.30 vs 0.07; P = .0001). The threshold of 180 million once-infected erythrocytes per liter discriminated patients with delayed hemolysis with 89% sensitivity and 83% specificity. Once-infected erythrocyte morphology analyzed by using ImageStream in 4 patients showed an 8.9% reduction in their projected area, an alteration likely contributing to their shorter lifespan. Delayed clearance of infected erythrocytes spared by pitting during AS treatment is an original mechanism of hemolytic anemia. Our findings consolidate a disease framework for posttreatment anemia in malaria in which delayed hemolysis is a new entity. The early concentration of once-infected erythrocytes is a solid candidate marker to predict post-AS delayed hemolysis.

Blood, 2014, vol.124, p.

Jauréguiberry Stéphane, Ndour Papa A, Roussel Camille, Ader Flavie, Safeukui Innocent, Nguyen Marie, Biligui Sylvestre, Ciceron Liliane, Mouri Oussama, Kendjo Eric, Bricaire François, Vray Muriel, Angoulvant Adéla, Mayaux Julien, Haldar Kasturi, Mazier Dominique, Danis Martin, Caumes Eric, Thellier Marc, Buffet Pierre

Plasmodium falciparum clearance is rapid and pitting independent in immune Malian children treated with artesunate for malaria.
Plasmodium falciparum clearance is rapid and pitting independent in immune Malian children treated with artesunate for malaria.

Ndour Papa Alioune, Lopera-Mesa Tatiana M, Diakité Seidina A S, Chiang Serena, Mouri Oussama, Roussel Camille, Jauréguiberry Stéphane, Biligui Sylvestre, Kendjo Eric, Claessens Antoine, Ciceron Liliane, Mazier Dominique, Thellier Marc, Diakité Mahamadou, Fairhurst Rick M, Buffet Pierre A
The Journal of infectious diseases, 211 (2015) 

[In Plasmodium falciparum-infected patients treated with artemisinins, parasitemia declines through so-called pitting, an innate splenic process that transforms infected red blood cells (iRBCs) into once-infected RBCs (O-iRBCs).,We measured pitting in 83 French travelers and 42 Malian children treated for malaria with artesunate.,In travelers, O-iRBCs peaked at 107.7% initial parasitemia. In Malian children aged 1.5-4 years, O-iRBCs peaked at higher concentrations than in children aged 9-13 years (91.60% vs 31.95%; P = .0097). The parasite clearance time in older children was shorter than in younger children (P = .0001), and the decline in parasitemia in children aged 1.5-4 years often started 6 hours after treatment initiation, a lag phase generally absent in infants and older children. A 6-hour lag phase in artificial pitting of artesunate-exposed iRBCs was also observed in vitro. The proportion of iRBCs recognized by autologous immunoglobulin G (IgG) correlated with the parasite clearance time (r = -0.501; P = .0006) and peak O-iRBC concentration (r = -0.420; P = .0033).,Antimalarial immunity correlates with fast artemisinin-induced parasite clearance and low pitting rates. In nonimmune populations, artemisinin-induced P. falciparum clearance is related to pitting and starts after a 6-hour lag phase. In immune populations, passively and naturally acquired immune mechanisms operating faster than pitting may exist. This mechanism may mitigate the emergence of artemisinin-resistant P. falciparum in Africa.]

The Journal of infectious diseases, 2015, vol.211, p.

Ndour Papa Alioune, Lopera-Mesa Tatiana M, Diakité Seidina A S, Chiang Serena, Mouri Oussama, Roussel Camille, Jauréguiberry Stéphane, Biligui Sylvestre, Kendjo Eric, Claessens Antoine, Ciceron Liliane, Mazier Dominique, Thellier Marc, Diakité Mahamadou, Fairhurst Rick M, Buffet Pierre A

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Biotigr Lab
Team 4 UMR-S 1134 INSERM
Université de Paris

Hôpital NECKER – Enfants Malades
149 rue de Sèvres
75015 Paris, France

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pierre.buffet@inserm.fr
mickael.marin@inserm.fr

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