Microfluidic study of retention and elimination of abnormal red blood cells by human spleen with implications for sickle cell disease.

The spleen clears altered red blood cells (RBCs) from circulation, contributing to the balance between RBC formation (erythropoiesis) and removal. The splenic RBC retention and elimination occur predominantly in open circulation where RBCs flow through macrophages and inter-endothelial slits (IESs). The mechanisms underlying and interconnecting these processes significantly impact clinical outcomes. In sickle cell disease (SCD), blockage of intrasplenic sickled RBCs is observed in infants splenectomized due to acute splenic sequestration crisis (ASSC). This life-threatening RBC pooling and organ swelling event is plausibly triggered or enhanced by intra-tissular hypoxia. We present an oxygen-mediated spleen-on-a-chip platform for in vitro investigations of the homeostatic balance in the spleen. To demonstrate and validate the benefits of this general microfluidic platform, we focus on SCD and study the effects of hypoxia on splenic RBC retention and elimination. We observe that RBC retention by IESs and RBC-macrophage adhesion are faster in blood samples from SCD patients than those from healthy subjects. This difference is markedly exacerbated under hypoxia. Moreover, the sickled RBCs under hypoxia show distinctly different phagocytosis processes from those non-sickled RBCs under hypoxia or normoxia. We find that reoxygenation significantly alleviates RBC retention at IESs, and leads to rapid unsickling and fragmentation of the ingested sickled RBCs inside macrophages. These results provide unique mechanistic insights into how the spleen maintains its homeostatic balance between splenic RBC retention and elimination, and shed light on how disruptions in this balance could lead to anemia, splenomegaly, and ASSC in SCD and possible clinical manifestations in other hematologic diseases.

A combined computational and experimental investigation of the filtration function of splenic macrophages in sickle cell disease.

Being the largest lymphatic organ in the body, the spleen also constantly controls the quality of red blood cells (RBCs) in circulation through its two major filtration components, namely interendothelial slits (IES) and red pulp macrophages. In contrast to the extensive studies in understanding the filtration function of IES, fewer works investigate how the splenic macrophages retain the aged and diseased RBCs, i.e., RBCs in sickle cell disease (SCD). Herein, we perform a computational study informed by companion experiments to quantify the dynamics of RBCs captured and retained by the macrophages. We first calibrate the parameters in the computational model based on microfluidic experimental measurements for sickle RBCs under normoxia and hypoxia, as those parameters are not available in the literature. Next, we quantify the impact of key factors expected to dictate the RBC retention by the macrophages in the spleen, namely, blood flow conditions, RBC aggregation, hematocrit, RBC morphology, and oxygen levels. Our simulation results show that hypoxic conditions could enhance the adhesion between the sickle RBCs and macrophages. This, in turn, increases the retention of RBCs by as much as four-fold, which could be a possible cause of RBC congestion in the spleen of patients with SCD. Our study on the impact of RBC aggregation illustrates a 'clustering effect', where multiple RBCs in one aggregate can make contact and adhere to the macrophages, leading to a higher retention rate than that resulting from RBC-macrophage pair interactions. Our simulations of sickle RBCs flowing past macrophages for a range of blood flow velocities indicate that the increased blood velocity could quickly attenuate the function of the red pulp macrophages on detaining aged or diseased RBCs, thereby providing a possible rationale for the slow blood flow in the open circulation of the spleen. Furthermore, we quantify the impact of RBC morphology on their tendency to be retained by the macrophages. We find that the sickle and granular-shaped RBCs are more likely to be filtered by macrophages in the spleen. This finding is consistent with the observation of low percentages of these two forms of sickle RBCs in the blood smear of SCD patients. Taken together, our experimental and simulation results aid in our quantitative understanding of the function of splenic macrophages in retaining the diseased RBCs and provide an opportunity to combine such knowledge with the current knowledge of the interaction between IES and traversing RBCs to apprehend the complete filtration function of the spleen in SCD.

Retention of uninfected red blood cells causing congestive splenomegaly is the major mechanism of anemia in malaria.

Splenomegaly frequently occurs in patients with Plasmodium falciparum (Pf) or P. vivax (Pv) malarial anemia, but mechanisms underlying this co-occurrence are unclear. In malaria-endemic Papua, Indonesia, we prospectively analyzed red blood cell (RBC) concentrations in the spleen and spleen-mimetic retention in 37 subjects splenectomized for trauma or hyperreactive splenomegaly, most of whom were infected with Plasmodium. Splenomegaly (median 357 g [range: 80-1918 g]) was correlated positively with the proportion of red-pulp on histological sections (median 88.1% [range: 74%-99.4%]; r = .59, p = .0003) and correlated negatively with the proportion of white-pulp (median 8.3% [range: 0.4%-22.9%]; r = -.50, p = .002). The number of RBC per microscopic field (>95% uninfected) was correlated positively with spleen weight in both Pf-infected (r = .73; p = .017) and Pv-infected spleens (r = .94; p = .006). The median estimated proportion of total-body RBCs retained in Pf-infected spleens was 8.2% (range: 1.0%-33.6%), significantly higher than in Pv-infected (2.6% [range: 0.6%-23.8%]; p = .015) and PCR-negative subjects (2.5% [range: 1.0%-3.3%]; p = .006). Retained RBCs accounted for over half of circulating RBC loss seen in Pf infections. The proportion of total-body RBC retained in Pf- and Pv-infected spleens correlated negatively with hemoglobin concentrations (r = -.56, p = .0003), hematocrit (r = -.58, p = .0002), and circulating RBC counts (r = -.56, p = .0003). Splenic CD71-positive reticulocyte concentrations correlated with spleen weight in Pf (r = 1.0; p = .003). Retention rates of peripheral and splenic RBCs were correlated negatively with circulating RBC counts (r = -.69, p = .07 and r = -.83, p = .008, respectively). In conclusion, retention of mostly uninfected RBC in the spleen, leading to marked congestion of the red-pulp, was associated with splenomegaly and is the major mechanism of anemia in subjects infected with Plasmodium, particularly Pf.

Rapid clearance of storage-induced microerythrocytes alters transfusion recovery.

Permanent availability of red blood cells (RBCs) for transfusion depends on refrigerated storage, during which morphologically altered RBCs accumulate. Among these, a subpopulation of small RBCs, comprising type III echinocytes, spheroechinocytes, and spherocytes and defined as storage-induced microerythrocytes (SMEs), could be rapidly cleared from circulation posttransfusion. We quantified the proportion of SMEs in RBC concentrates from healthy human volunteers and assessed correlation with transfusion recovery, investigated the fate of SMEs upon perfusion through human spleen ex vivo, and explored where and how SMEs are cleared in a mouse model of blood storage and transfusion. In healthy human volunteers, high proportion of SMEs in long-stored RBC concentrates correlated with poor transfusion recovery. When perfused through human spleen, 15% and 61% of long-stored RBCs and SMEs were cleared in 70 minutes, respectively. High initial proportion of SMEs also correlated with high retention of RBCs by perfused human spleen. In the mouse model, SMEs accumulated during storage. Transfusion of long-stored RBCs resulted in reduced posttransfusion recovery, mostly due to SME clearance. After transfusion in mice, long-stored RBCs accumulated predominantly in spleen and were ingested mainly by splenic and hepatic macrophages. In macrophage-depleted mice, splenic accumulation and SME clearance were delayed, and transfusion recovery was improved. In healthy hosts, SMEs were cleared predominantly by macrophages in spleen and liver. When this well-demarcated subpopulation of altered RBCs was abundant in RBC concentrates, transfusion recovery was diminished. SME quantification has the potential to improve blood product quality assessment. This trial was registered at www.clinicaltrials.gov as #NCT02889133.

Erythropoietin downregulates red blood cell clearance, increasing transfusion efficacy in severely anemic recipients.

Red blood cells (RBC) transfusion is used to alleviate symptoms and prevent complications in anemic patients by restoring oxygen delivery to tissues. RBC transfusion efficacy, that can be measured by a rise in hemoglobin (Hb) concentration, is influenced by donor-, product-, and recipient-related characteristics. In some studies, severe pre-transfusion anemia is associated with a greater than expected Hb increment following transfusion but the biological mechanism underpinning this relationship remains poorly understood. We conducted a prospective study in critically ill patients and quantified Hb increment following one RBC transfusion. In a murine model, we investigated the possibility that, in conjunction with the host erythropoietic response, the persistence of transfused donor RBC is improved to maintain a highest RBC biomass. We confirmed a correlation between a greater Hb increment and a deeper pre-transfusion anemia in a cohort of 17 patients. In the mouse model, Hb increment and post-transfusion recovery were increased in anemic recipients. Post-transfusion RBC recovery was improved in hypoxic mice or those receiving an erythropoiesis-stimulating agent and decreased in those treated with erythropoietin (EPO)-neutralizing antibodies, suggesting that EPO signaling is necessary to observe this effect. Irradiated recipients also showed decreased post-transfusion RBC recovery. The EPO-induced post-transfusion RBC recovery improvement was abrogated in irradiated or in macrophage-depleted recipients, but maintained in splenectomized recipients, suggesting a mechanism requiring erythroid progenitors and macrophages, but which is not spleen-specific. Our study highlights a physiological role of EPO in downregulating post-transfusion RBC clearance, contributing to maintain a vital RBC biomass to rapidly cope with hypoxemia.

How the spleen reshapes and retains young and old red blood cells: A computational investigation.

The spleen, the largest secondary lymphoid organ in humans, not only fulfils a broad range of immune functions, but also plays an important role in red blood cell's (RBC) life cycle. Although much progress has been made to elucidate the critical biological processes involved in the maturation of young RBCs (reticulocytes) as well as removal of senescent RBCs in the spleen, the underlying mechanisms driving these processes are still obscure. Herein, we perform a computational study to simulate the passage of RBCs through interendothelial slits (IES) in the spleen at different stages of their lifespan and investigate the role of the spleen in facilitating the maturation of reticulocytes and in clearing the senescent RBCs. Our simulations reveal that at the beginning of the RBC life cycle, intracellular non-deformable particles in reticulocytes can be biomechanically expelled from the cell upon passage through IES, an insightful explanation of why this peculiar "pitting" process is spleen-specific. Our results also show that immature RBCs shed surface area by releasing vesicles after crossing IES and progressively acquire the biconcave shape of mature RBCs. These findings likely explain why RBCs from splenectomized patients are significantly larger than those from nonsplenectomized subjects. Finally, we show that at the end of their life span, senescent RBCs are not only retained by IES due to reduced deformability but also become susceptible to mechanical lysis under shear stress. This finding supports the recent hypothesis that transformation into a hemolyzed ghost is a prerequisite for phagocytosis of senescent RBCs. Altogether, our computational investigation illustrates critical biological processes in the spleen that cannot be observed in vivo or in vitro and offer insights into the role of the spleen in the RBC physiology.

Surveillance of leishmaniasis cases from 15 European centres, 2014 to 2019: a retrospective analysis.

BackgroundSurveillance of human leishmaniasis in Europe is mostly limited to country-specific information from autochthonous infections in the southern part. As at the end of 2021, no integrated analysis has been performed for cases seen across centres in different European countries.AimTo provide a broad perspective on autochthonous and imported leishmaniasis cases in endemic and non-endemic countries in Europe.MethodsWe retrospectively collected records from cutaneous, mucosal and visceral leishmaniasis cases diagnosed in 15 centres between 2014 and 2019. Centres were located in 11 countries: Belgium, France, Germany, Italy, the Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and the United Kingdom. Data on country of infection, reason for travelling, infecting species, age and sex were analysed.ResultsWe obtained diagnostic files from 1,142 cases, of which 76%, 21% and 3% had cutaneous, visceral, and mucosal disease, respectively. Of these, 68% were men, and 32% women, with the median age of 37 years (range: 0-90) at diagnosis. Visceral leishmaniasis was mainly acquired in Europe (88%; 167/190), while cutaneous leishmaniasis was primarily imported from outside Europe (77%; 575/749). Sixty-two percent of cutaneous leishmaniasis cases from outside Europe were from the Old World, and 38% from the New World. Geographic species distribution largely confirmed known epidemiology, with notable exceptions.ConclusionsOur study confirms previous reports regarding geographic origin, species, and traveller subgroups importing leishmaniasis into Europe. We demonstrate the importance of pooling species typing data from many centres, even from areas where the aetiology is presumably known, to monitor changing epidemiology.

Computational modeling of biomechanics and biorheology of heated red blood cells.

Because of their compromised deformability, heat denatured erythrocytes have been used as labeled probes to visualize spleen tissue or to assess the ability of the spleen to retain stiff red blood cells (RBCs) for over three decades, e.g., see Looareesuwan et al. N. Engl. J. Med. (1987). Despite their good accessibility, it is still an open question how heated RBCs compare to certain diseased RBCs in terms of their biomechanical and biorheological responses, which may undermine their effective usage and even lead to misleading experimental observations. To help answering this question, we perform a systematic computational study of the hemorheological properties of heated RBCs with several physiologically relevant static and hemodynamic settings, including optical-tweezers test, relaxation of prestretched RBCs, RBC traversal through a capillary-like channel and a spleen-like slit, and a viscometric rheology test. We show that our in silico RBC models agree well with existing experiments. Moreover, under static tests, heated RBCs exhibit deformability deterioration comparable to certain disease-impaired RBCs such as those in malaria. For RBC traversal under confinement (through microchannel or slit), heated RBCs show prolonged transit time or retention depending on the level of confinement and heating procedure, suggesting that carefully heat-treated RBCs may be useful for studying splenic- or vaso-occlusion in vascular pathologies. For the rheology test, we expand the existing bulk viscosity data of heated RBCs to a wider range of shear rates (1-1000 s-1) to represent most pathophysiological conditions in macro- or microcirculation. Although heated RBC suspension shows elevated viscosity comparable to certain diseased RBC suspensions under relatively high shear rates (100-1000 s-1), they underestimate the elevated viscosity (e.g., in sickle cell anemia) at low shear rates (<10 s-1). Our work provides mechanistic rationale for selective usage of heated RBC as a potentially useful model for studying the abnormal traversal dynamics and hemorheology in certain blood disorders.

Intravenous Artesunate for the Treatment of Severe Imported Malaria: Implementation, Efficacy, and Safety in 1391 Patients.

BACKGROUND: Intravenous artesunate is the World Health Organization-recommended first-line treatment for severe malaria worldwide, but it is still not fully licensed in Europe. Observational studies documenting its safety and efficacy in imported malaria are thus essential. METHODS: We prospectively collected clinical and epidemiological features of 1391 artesunate-treated patients among 110 participant centers during the first 7 years (2011-2017) of a national program implemented by the French Drug Agency. RESULTS: Artesunate became the most frequent treatment for severe malaria in France, rising from 9.9% in 2011 to 71.4% in 2017. Mortality was estimated at 4.1%. Treatment failure was recorded in 27 patients, but mutations in the Kelch-13 gene were not observed. Main reported adverse events (AEs) were anemia (136 cases), cardiac events (24, including 20 episodes of conduction disorders and/or arrhythmia), and liver enzyme elevation (23). Mortality and AEs were similar in the general population and in people with human immunodeficiency virus, who were overweight, or were pregnant, but the only pregnant woman treated in the first trimester experimented a hemorrhagic miscarriage. The incidence of post-artesunate-delayed hemolysis (PADH) was 42.8% when specifically assessed in a 98-patient subgroup, but was not associated with fatal outcomes or sequelae. PADH was twice as frequent in patients of European compared with African origin. CONCLUSIONS: Artesunate was rapidly deployed and displayed a robust clinical benefit in patients with severe imported malaria, despite a high frequency of mild to moderate PADH. Further explorations in the context of importation should assess outcomes during the first trimester of pregnancy and collect rare but potentially severe cardiac AEs.

Evaluation of splenic accumulation and colocalization of immature reticulocytes and Plasmodium vivax in asymptomatic malaria: A prospective human splenectomy study.

BACKGROUND: A very large biomass of intact asexual-stage malaria parasites accumulates in the spleen of asymptomatic human individuals infected with Plasmodium vivax. The mechanisms underlying this intense tropism are not clear. We hypothesised that immature reticulocytes, in which P. vivax develops, may display high densities in the spleen, thereby providing a niche for parasite survival. METHODS AND FINDINGS: We examined spleen tissue in 22 mostly untreated individuals naturally exposed to P. vivax and Plasmodium falciparum undergoing splenectomy for any clinical indication in malaria-endemic Papua, Indonesia (2015 to 2017). Infection, parasite and immature reticulocyte density, and splenic distribution were analysed by optical microscopy, flow cytometry, and molecular assays. Nine non-endemic control spleens from individuals undergoing spleno-pancreatectomy in France (2017 to 2020) were also examined for reticulocyte densities. There were no exclusion criteria or sample size considerations in both patient cohorts for this demanding approach. In Indonesia, 95.5% (21/22) of splenectomy patients had asymptomatic splenic Plasmodium infection (7 P. vivax, 13 P. falciparum, and 1 mixed infection). Significant splenic accumulation of immature CD71 intermediate- and high-expressing reticulocytes was seen, with concentrations 11 times greater than in peripheral blood. Accordingly, in France, reticulocyte concentrations in the splenic effluent were higher than in peripheral blood. Greater rigidity of reticulocytes in splenic than in peripheral blood, and their higher densities in splenic cords both suggest a mechanical retention process. Asexual-stage P. vivax-infected erythrocytes of all developmental stages accumulated in the spleen, with non-phagocytosed parasite densities 3,590 times (IQR: 2,600 to 4,130) higher than in circulating blood, and median total splenic parasite loads 81 (IQR: 14 to 205) times greater, accounting for 98.7% (IQR: 95.1% to 98.9%) of the estimated total-body P. vivax biomass. More reticulocytes were in contact with sinus lumen endothelial cells in P. vivax- than in P. falciparum-infected spleens. Histological analyses revealed 96% of P. vivax rings/trophozoites and 46% of schizonts colocalised with 92% of immature reticulocytes in the cords and sinus lumens of the red pulp. Larger splenic cohort studies and similar investigations in untreated symptomatic malaria are warranted. CONCLUSIONS: Immature CD71+ reticulocytes and splenic P. vivax-infected erythrocytes of all asexual stages accumulate in the same splenic compartments, suggesting the existence of a cryptic endosplenic lifecycle in chronic P. vivax infection. Findings provide insight into P. vivax-specific adaptions that have evolved to maximise survival and replication in the spleen.

Metabolic rejuvenation upgrades circulatory functions of red blood cells stored under blood bank conditions.

BACKGROUND: Red blood cells (RBC) change upon hypothermic conservation, and storage for 6 weeks is associated with the short-term clearance of 15% to 20% of transfused RBCs. Metabolic rejuvenation applied to RBCs before transfusion replenishes energetic sources and reverses most storage-related alterations, but how it impacts RBC circulatory functions has not been fully elucidated. STUDY DESIGN AND METHODS: Six RBC units stored under blood bank conditions were analyzed weekly for 6 weeks and rejuvenated on Day 42 with an adenine-inosine-rich solution. Impact of storage and rejuvenation on adenosine triphosphate (ATP) levels, morphology, accumulation of storage-induced microerythrocytes (SMEs), elongation under an osmotic gradient (by LORRCA), hemolysis, and phosphatidylserine (PS) exposure was evaluated. The impact of rejuvenation on filterability and adhesive properties of stored RBCs was also assessed. RESULTS: Rejuvenation of RBCs restored intracellular ATP to almost normal levels and decreased the PS exposure from 2.78% to 0.41%. Upon rejuvenation, the proportion of SME dropped from 28.2% to 9.5%, while the proportion of normal-shaped RBCs (discocytes and echinocytes 1) increased from 47.7% to 67.1%. In LORCCA experiments, rejuvenation did not modify the capacity of RBCs to elongate and induced a reduction in cell volume. In functional tests, rejuvenation increased RBC filterability in a biomimetic splenic filter (+16%) and prevented their adhesion to endothelial cells (-87%). CONCLUSION: Rejuvenation reduces the proportion of morphologically altered and adhesive RBCs that accumulate during storage. Along with the improvement in their filterability, these data show that rejuvenation improves RBC properties related to their capacity to persist in circulation after transfusion.

Systematic analysis of direct antiglobulin test results in post-artesunate delayed haemolysis.

BACKGROUND: Post-artesunate delayed haemolysis (PADH) is common after severe malaria episodes. PADH is related to the "pitting" phenomenon and the synchronous delayed clearance of once-infected erythrocytes, initially spared during treatment. However, direct antiglobulin test (DAT) positivity has been reported in several PADH cases, suggesting a contribution of immune-mediated erythrocyte clearance. The aim of the present study was to compare clinical features of cases presenting a positive or negative DAT. METHODS: Articles reporting clinical data of patients diagnosed with PADH, for whom DAT had been performed, were collected from PubMed database. Data retrieved from single patients were extracted and univariate analysis was performed in order to identify features potentially related to DAT results and steroids use. RESULTS: Twenty-two studies reporting 39 PADH cases were included: median baseline parasitaemia was 20.8% (IQR: 11.2-30) and DAT was positive in 17 cases (45.5%). Compared to DAT-negative individuals, DAT-positive patients were older (49.5 vs 31; p = 0.01), had a higher baseline parasitaemia (27% vs 17%; p = 0.03) and were more commonly treated with systemic steroids (11 vs 3 patients, p = 0.002). Depth and kinetics of delayed anaemia were not associated with DAT positivity. CONCLUSIONS: In this case series, almost half of the patients affected by PADH had a positive DAT. An obvious difference between the clinical courses of patients presenting with a positive or negative DAT was lacking. This observation suggests that DAT result may not be indicative of a pathogenic role of anti-erythrocytes antibodies in patients affected by PADH, but it may be rather a marker of immune activation.

Mechanics of diseased red blood cells in human spleen and consequences for hereditary blood disorders.

In red blood cell (RBC) diseases, the spleen contributes to anemia by clearing the damaged RBCs, but its unique ability to mechanically challenge RBCs also poses the risk of inducing other pathogenic effects. We have analyzed RBCs in hereditary spherocytosis (HS) and hereditary elliptocytosis (HE), two typical examples of blood disorders that result in membrane protein defects in RBCs. We use a two-component protein-scale RBC model to simulate the traversal of the interendothelial slit (IES) in the human spleen, a stringent biomechanical challenge on healthy and diseased RBCs that cannot be directly observed in vivo. In HS, our results confirm that the RBC loses surface due to weakened cohesion between the lipid bilayer and the cytoskeleton and reveal that surface loss may result from vesiculation of the RBC as it crosses IES. In HE, traversing IES induces sustained elongation of the RBC with impaired elasticity and fragmentation in severe disease. Our simulations thus suggest that in inherited RBC disorders, the spleen not only filters out pathological RBCs but also directly contributes to RBC alterations. These results provide a mechanistic rationale for different clinical outcomes documented following splenectomy in HS patients with spectrin-deficient and ankyrin-deficient RBCs and offer insights into the pathogenic role of human spleen in RBC diseases.

Propensity Score Analysis of Artesunate Versus Quinine for Severe Imported Plasmodium falciparum Malaria in France.

BACKGROUND: Little is known on the use of artesunate compared with quinine for the treatment of imported malaria cases in nonendemic countries with a high level of care. Therefore, we compared the 2 treatments in terms of mortality and hospital and intensive care unit (ICU) discharge rates. METHODS: We analyzed the cohort of all severe imported malaria patients reported to the French National Reference Center from 2011 to 2017. After controlling for differences between quinine- and artesunate-treated individuals using the inverse probability of treatment weighting method, 28-day mortality rate was compared between the groups as well as hospital and ICU discharge rates using Kaplan-Meier estimation and weighted Cox proportional hazard models. RESULTS: Overall, 1544 patients were enrolled. Fifty patients died, 18 in the quinine group (n = 460) and 32 in the artesunate group (n = 1084), corresponding to death rates of 3.9% and 2.9%, respectively. No difference was evident between quinine and artesunate either in mortality or in hospital discharge rate, with hazard ratios (HRs) of 1.03 (95% confidence interval [CI], 0.47-2.25) and 1.12 (95% CI, 0.94-1.34), respectively. Artesunate was associated with a faster ICU discharge rate (HR, 1.18. 95% CI, 1.02-1.36). CONCLUSIONS: In a country with a high level of care, artesunate was associated with a shorter length of stay in the ICU, which supports the actual therapeutic transition; however, no difference was found in terms of mortality or in hospital discharge rates between artesunate- and quinine-treated patients.

Plasmodium falciparum Clearance Is Pitting-Dependent With Artemisinin-Based Drugs but Pitting-Independent With Atovaquone-Proguanil or Mefloquine.

Pitting, the removal of dead parasites from their host erythrocyte, has been studied in patients with severe malaria treated parenterally with quinine or artesunate, and was recently shown to contribute to delayed hemolysis, a frequent adverse event of artesunate. We quantified pitting in 81 travelers treated with oral antimalarial therapy. Pitting rate was high (55.8%) with artemisinin-based combinations, but <10% with the nonartemisinin drugs quinine, mefloquine, and atovaquone-proguanil. This may, in part, explain the slower parasite clearance in patients treated with antimalarial drugs lacking an artemisinin component, as well as the absence of posttreatment hemolysis with these drugs.

Reexamination of the chromium-51-labeled posttransfusion red blood cell recovery method.

BACKGROUND: The chromium-51-labeled posttransfusion recovery (PTR) study has been the gold-standard test for assessing red blood cell (RBC) quality. Despite guiding RBC storage development for decades, it has several potential sources for error. METHODS: Four healthy adult volunteers each donated an autologous, leukoreduced RBC unit, aliquots were radiolabeled with technetium-99m after 1 and 6 weeks of storage, and then infused. Subjects were imaged by single-photon-emission computed tomography immediately and 4 hours after infusion. Additionally, from subjects described in a previously published study, adenosine triphosphate levels in transfusates infused into 52 healthy volunteers randomized to a single autologous, leukoreduced, RBC transfusion after 1, 2, 3, 4, 5, or 6 weeks of storage were correlated with PTR and laboratory parameters of hemolysis. RESULTS: Evidence from one subject imaged after infusion of technetium-99m-labeled RBCs suggests that, in some individuals, RBCs may be temporarily sequestered in the liver and spleen immediately following transfusion and then subsequently released back into circulation; this could be one source of error leading to PTR results that may not accurately predict the true quantity of RBCs cleared by intra- and/or extravascular hemolysis. Indeed, adenosine triphosphate levels in the transfusates correlated more robustly with measures of extravascular hemolysis in vivo (e.g., serum iron, indirect bilirubin, non-transferrin-bound iron) than with PTR results or measures of intravascular hemolysis (e.g., plasma free hemoglobin). CONCLUSIONS: Sources of measurement error are inherent in the chromium-51 PTR method. Transfusion of an entire unlabeled RBC unit, followed by quantifying extravascular hemolysis markers, may more accurately measure true posttransfusion RBC recovery.

Artenimol-piperaquine in children with uncomplicated imported falciparum malaria: experience from a prospective cohort.

BACKGROUND: Although malaria remains one of the major public health threats in inter-tropical areas, there is limited understanding of imported malaria in children by paediatricians and emergency practitioners in non-endemic countries, often resulting in misdiagnosis and inadequate treatment. Moreover, classical treatments (atovaquone-proguanil, quinine, mefloquine) are limited either by lengthy treatment courses or by side effects. Since 2010, the World Health Organization (WHO) has recommended the use of oral artemisinin-based combination therapy for the treatment of uncomplicated Plasmodium falciparum malaria worldwide. The benefits of artenimol-piperaquine in children have been validated in endemic countries but experience remains limited in cases of imported malaria. METHODS: This prospective observational study in routine paediatric care took place at the Emergency Department, Robert-Debré Hospital (Paris, France) from September 2012 to December 2014. Tolerance and efficacy of artenimol-piperaquine in children presenting with the following inclusion criteria were assessed: P. falciparum positive on thin or thick blood smear; and the absence of WHO-defined features of severity. RESULTS: Among 83 children included in this study, treatment with artenimol-piperaquine was successful in 82 children (98.8%). None of the adverse events were severe and all were considered mild with no significant clinical impact. This also applied to cardiological adverse events despite a significant increase of the mean post-treatment QTc interval. CONCLUSION: Artenimol-piperaquine displays a satisfying efficacy and tolerance profile as a first-line treatment for children with imported uncomplicated falciparum malaria and only necessitates three once-daily oral intakes of the medication. Comparative studies versus artemether-lumefantrine or atovaquone-proguanil would be useful to confirm the results of this study.

Biomechanics of red blood cells in human spleen and consequences for physiology and disease.

Red blood cells (RBCs) can be cleared from circulation when alterations in their size, shape, and deformability are detected. This function is modulated by the spleen-specific structure of the interendothelial slit (IES). Here, we present a unique physiological framework for development of prognostic markers in RBC diseases by quantifying biophysical limits for RBCs to pass through the IES, using computational simulations based on dissipative particle dynamics. The results show that the spleen selects RBCs for continued circulation based on their geometry, consistent with prior in vivo observations. A companion analysis provides critical bounds relating surface area and volume for healthy RBCs beyond which the RBCs fail the "physical fitness test" to pass through the IES, supporting independent experiments. Our results suggest that the spleen plays an important role in determining distributions of size and shape of healthy RBCs. Because mechanical retention of infected RBC impacts malaria pathogenesis, we studied key biophysical parameters for RBCs infected with Plasmodium falciparum as they cross the IES. In agreement with experimental results, surface area loss of an infected RBC is found to be a more important determinant of splenic retention than its membrane stiffness. The simulations provide insights into the effects of pressure gradient across the IES on RBC retention. By providing quantitative biophysical limits for RBCs to pass through the IES, the narrowest circulatory bottleneck in the spleen, our results offer a broad approach for developing quantitative markers for diseases such as hereditary spherocytosis, thalassemia, and malaria.