Plasmapheresis may improve clinical condition in sickle cell disease through its effects on red blood cell rheology
Authors: Elie Nader, Philippe Connes, Yann Lamarre, Céline Renoux, Philippe Joly, Marie‐Dominique Hardy‐Dessources, Giovanna Cannas, Nathalie Lemonne, Samir K. Ballas
Journal: American Journal of Hematology (2017)

Acute chest syndrome (ACS) is the leading cause of death in patients with sickle cell disease (SCD). Its etiology is multifactorial and remains poorly understood. We report the case of a patient with homozygous SCD who developed severe ACS and who was successfully managed with plasma exchange. Additionally, we performed in vitro experiments to test the effects of plasma and several plasma molecules on the rheology of red blood cell (RBC) of SCD patients. A 19-year-old woman, known to have homozygous SCD, was admitted to the Emergency Department (ED) with an acute vaso-occlusive crisis (VOC) and fever. The pain was typical of her previous crises and was throbbing in nature involving her low back, chest, and abdomen with a score of 10/10. She denied cough, nausea, vomiting, and diarrhea. Past medical history was significant for frequent VOCs that required treatment with parenteral opioids in the ED and/or hospital. She also had history of asthma and recurrent ACS at least three times. The last episode of ACS was about 3 months before the current admission and was first treated with supportive care, simple transfusion, and antibiotics but later required two consecutive RBC exchanges (RCX) with suboptimal resolution. Noteworthy is that the first RCX provided no improvement despite the fact that hemoglobin S (HbS) decreased to 27% and the second RCX decreased HbS further to 14%. Physical examination was significant for temperature (37.2°C), heart rate (106 bpm), and diffuse tenderness over the abdomen, low back, and extremities. There were no wheezes, rales, or rhonchi on lung examination. Laboratory findings on admission included hemoglobin = 5.9 g/dL, hematocrit = 17.7%, mean cell volume = 78 fL, white blood cell count = 32 500/µL, platelet count = 387 000/µL, and reticulocyte = 8.5%. Her steady state values included hemoglobin = 8.6 g/dL, hematocrit 26.3%, mean cell volume = 80 fl, white blood cell count = 14 000/µL, platelet count = 494 000/µL, reticulocyte = 7.2%, fetal hemoglobin = 24%, and RBC deformability index = 0.26 (normal control value = 0.59). Biochemical parameters were within the normal range with the exception of total bilirubin = 4.2 mg/mL and lactate dehydrogenase level = 394 IU. Molecular diagnostics showed 3α-genes (ie, heterozygous α-thalassemia) and homozygosity for the Benin βS haplotype. Two days after admission, her condition worsened with fever (38.3°C) and dyspnea. She was treated with supportive care, antibiotics, and simple transfusion with 2 units of RBC over 2 days. Her HbS dropped to 20%. Her hypoxia, however, worsened, and she required intubation with ventilator support in the intensive care unit. Radiography was consistent with ACS. Because she had already achieved a reduction in HbS, a single volume plasma exchange (PEX) was performed using the Cobe Spectra and fresh frozen plasma as the replacement fluid (due to moderate prolongation of her prothrombin and partial thromboplastin times). Her oxygenation improved soon after PEX, and she was extubated 4 days later. The course of events in this patient suggests that PEX could have caused the rapid resolution of her ACS, possibly from normalization of plasma parameters. We tested the effects of sickle and healthy plasma on RBC rheology. In vitro experiments were performed (ethical approval: CPP Est-Lyon/L14–127) to measure RBC deformability and aggregation in 12 patients with SCD (steady state) and 12 controls (AA) either in their autologous plasma or after plasma switching (blood matching for A-B-O and Rhesus group; Figure 1). RBC deformability in autologous plasma was higher in AA than in SCD individuals (Figure 1A, P < .001). After plasma switching, SCD-RBC deformability improved (P < .05) while the deformability of AA RBCs re-suspended in SCD plasma decreased (P < .05). RBC aggregation (M and M1 indexes) was higher in SCD than in AA individuals (Figure 1B,C, P < .05). When SCD RBCs were mixed with AA plasma, both M and M1 decreased (P < .01 and P < .01, respectively) while aggregation of AA RBCs mixed with SCD plasma increased (P < .01 and P < .05 for M and M1, respectively). These findings demonstrate that plasma modulates the rheological properties of RBCs in SCD. Effects of SCD and normal plasma on the rheology of sickle and normal RBCs and effects of plasma molecules on RBC aggregates strength in SCD patients. Blood was centrifuged for separation of RBCs and plasma (10 min, 2100g, 25°C). Plasma was stored at 4°C and RBC pellet was washed two times with PBS-glucose buffer (5 mM glucose). A-C, The pellet was then re-suspended in autologous or heterologous plasma at hematocrit 40% to perform hemorheological measurements. RBC deformability was measured by ektacytometry (LORRCA MaxSis, Mechatronics, The Netherland). RBC aggregation was determined by light transmission with the Myrenne aggregometer (Myrenne GmbH, Roetgen, Germany). Two RBC aggregation indexes were measured: M = RBC aggregation at stasis; M1 = RBC aggregation at low shear rate. A, RBC deformability at 3 Pa; B, M RBC aggregation index; C, M1 RBC aggregation index. D, The RBC pellet was re-suspended in autologous plasma at hematocrit 30% with either 3 mg/mL of fibrinogen (FIB, Sigma-Aldrich), 1 μg/mL of thrombospondin (THBS1; R&D Systems), 272 μg/mL of von Willebrand factor (vWF; R&D Systems) or without any other molecule (control condition, CTL) and incubated for 30 min at 25°C. Then, RBC aggregates strength was measured by syllectometry (LORRCA, Mechatronics, The Netherland). A two-way ANOVA with repeated measures + post hoc Tukey test was used. Significant differences: *P < .05; **P < .01; ***P < .001 Plasma from SCD patients contains high levels of several inflammatory mediators and adhesion molecules. Fibrinogen (FIB) is the leading factor promoting RBC aggregation and its level increases during VOCs.1 Thrombospondin 1 (THBS1) and von-Willebrand factor (vWF) could also modulate RBC aggregation through their abilities to bind several RBC membrane receptors2 but this hypothesis has never been tested. Although we could not measure the plasma levels of FIB, THBS1, and vWF in this study, we tested their effects on the RBC aggregates strength of 9 patients with SCD at steady state (ethical approval: CPP Sud/Ouest Outre Mer III/2012-A00701-42; Figure 1D). We demonstrated that all three molecules increased RBC aggregates strength compared to a control condition (P < .001 for THBS1 and vWF, P < .05 for FIB, Figure 1D). Decreased RBC deformability and increased RBC aggregation/aggregates strength affect blood flow and may participate to the occurrence of ACS in patients with SCD.3 Our results suggest that replacing sickle by healthy plasma could improve the rheology of RBCs because of the normalization of several plasma parameters such as pro-aggregant molecules or oxidative stress. PEX has not been used very often in SCD but few reports have shown the beneficial clinical effects in acute situations such as multi-organ failure, VOC, and ACS.4-6 It is unclear whether PEX was responsible for the patient's clinical improvement rather than the antibiotic treatment or the natural course of the disease. Indeed, the extent to which PEX can serve as an adjunct to conventional RCX in ACS, or even serve as a replacement fluid for RCX or simple transfusion warrants further investigation. Nothing to report. Samir K. Ballas http://orcid.org/0000-0002-7566-1466