Free Hb Levels Research Articles

Hemolysis Precedes Urine Color Change in Patients Undergoing Open-Heart Surgery on Cardiopulmonary Bypass.

Red-colored urine often occurs in patients in the perioperative period who undergo cardiac surgery using cardiopulmonary bypass (CPB). This urine color change has been utilized for approximating hemolysis during CPB without a proven relationship for ongoing hemolysis. This case series study aimed to examine the relationship between plasma free hemoglobin (Hb) levels and quantified measures of urine color. Ten patients were enrolled in this study. Blood and urine were collected for analyses for the following time points: before surgery, two hours after the initiation of CPB, every 30 min during CPB thereafter, and 0, 2, 4, 12, and 24 hours after the completion of CPB. We measured free Hb in plasma and urine using the azide-methemoglobin method. Photographs of urine were obtained, and the luminance of the three basic colors (red/green/blue) was analyzed by quantitative luminance contrast analysis to find a correlation for hemolysis. Median levels of plasma free Hb were 0.015 (0.010-0.080, n = 10) g/dL at baseline. During the CPB, increases in plasma free Hb levels were measured: median plasma free Hb levels were increased to0.100 g/dL(0.020-0.240, p = 0.039, vs. baseline, n = 9) at two hours into CPB, median and range, respectively.In contrast, increases in urinary free Hb levels and/or urine color changes were measured only after cessation of CPB in nine patients. Urine color change or elevation of urinary free Hb levels followed the elevation of plasma free Hb levels with considerable delay.

Red Blood Cell Abnormalities as the Mirror of SARS-CoV-2 Disease Severity: A Pilot Study.

PurposeUnraveling the pathophysiology of COVID-19 disease is of crucial importance for designing treatment. The purpose of this study is to investigate the effects of the disease on erythrocytes (RBCs) and to correlate the findings with disease severity.Materials and MethodsHospitalized patients (n = 36) with COVID-19 and control group of healthy volunteers (n = 18) were included in the study. Demographic data, clinical, laboratory and chest Computed Tomography (CT) findings at time of admission were recorded. Laboratory measurements included: Hemoglobin (H b), indirect billirubin, LDH, D-Dimers, and plasma free hemoglobin (plasma free-Hb). On RBCs were performed: osmotic fragility (MCF), Free-Hb after mechanical stress (Free-Hb-MECH), intracellular RBC concentration of calcium ions (iCa2+), intracellular ROS (iROS), G6PD, intracellular active caspase-3 (RBC-caspase-3), IgG immunoglobulins (RBC-IgGs), which are bound on RBCs’ senescent neo-antigen proteins and RBC surface phosphatidylserine (RBC-PS).ResultsThe percentage of males was 50 and 66% and the mean age was 65.16 ± 14.24 and 66.33 ± 13.48 years among patients and controls respectively (mean ± SD, p = 0.78). Upon admission patients’ PO2/FiO2 ratio was 305.92 ± 76.75 and distribution of infiltration extend on chest CT was: 0–25% (N = 19), 25–50%: (N = 7), and 50–75% (N = 9). Elevated hemolysis markers (LDH and plasma free-Hb) were observed in patients compared to the control group. Patients’ RBCs were more sensitive to mechanical stress, and exhibited significantly elevated apoptotic markers (iCa2+, RBC-PS). Plasma free Hb levels correlated with the extend of pulmonary infiltrates on chest CT in COVID-19 patients. Surprisingly, patients’ RBC-iROS were decreased, a finding possibly related with the increased G6PDH levels in this group, suggesting a possible compensatory mechanism against the virus. This compensatory mechanism seemed to be attenuated as pulmonary infiltrates on chest CT deteriorated. Furthermore, RBC-IgGs correlated with the severity of pulmonary CT imaging features as well as the abnormality of lung function, which are both associated with increased disease severity. Lastly, patients’ D-Dimers correlated with RBC surface phosphatidylserine, implying a possible contribution of the red blood cells in the thrombotic diathesis associated with the SARS-CoV-2 disease.ConclusionThis pilot study suggests that SARS-CoV-2 infection has an effect on red blood cells and there seems to be an association between RBC markers and disease severity in these patients.

Red blood cell homeostasis in children and adults with and without asymptomatic malaria infection in Burkina Faso

Asymptomatic malaria infections may affect red blood cell (RBC) homeostasis. Reports indicate a role for chronic hemolysis and splenomegaly, however, the underlying processes are incompletely understood. New hematology analysers provide parameters for a more comprehensive analysis of RBC hemostasis. Complete blood counts were analysed in subjects from all age groups (n = 1118) living in a malaria hyperendemic area and cytokines and iron biomarkers were also measured. Subjects were divided into age groups (<2 years, 2-4, 5-14 and ≥15 years old) and clinical categories (smear-negative healthy subjects, asymptomatic malaria and clinical malaria). We found that hemoglobin levels were similar in smear-negative healthy children and asymptomatic malaria children but significantly lower in clinical malaria with a maximum difference of 2.2 g/dl in children <2 years decreasing to 0.1 g/dl in those aged ≥15 years. Delta-He, presenting different hemoglobinization of reticulocytes and RBC, levels were lower in asymptomatic and clinial malaria, indicating a recent effect of malaria on erythropoiesis. Reticulocyte counts and reticulocyte production index (RPI), indicating the erythropoietic capacity of the bone marrow, were higher in young children with malaria compared to smear-negative subjects. A negative correlation between reticulocyte counts and Hb levels was found in asymptomatic malaria (ρ = -0.32, p<0.001) unlike in clinical malaria (ρ = -0.008, p = 0.92). Free-Hb levels, indicating hemolysis, were only higher in clinical malaria. Phagocytozing monocytes, indicating erythophagocytosis, were highest in clinical malaria, followed by asymptomatic malaria and smear-negative subjects. Circulating cytokines and iron biomarkers (hepcidin, ferritin) showed similar patterns. Pro/anti-inflammatory (IL-6/IL-10) ratio was higher in clinical than asymptomatic malaria. Cytokine production capacity of ex-vivo whole blood stimulation with LPS was lower in children with asymptomatic malaria compared to smear-negative healthy children. Bone marrow response can compensate the increased red blood cell loss in asymptomatic malaria, unlike in clinical malaria, possibly because of limited level and length of inflammation. Trial registration: Prospective diagnostic study: ClinicalTrials.gov identifier: NCT02669823. Explorative cross-sectional field study: ClinicalTrials.gov identifier: NCT03176719.

Effect of warming and infusion of red blood cell concentrates on markers of haemolysis: An ex vivo simulation study

BackgroundTransfusion of red blood cell (RBC) concentrates is a common procedure to restore blood volume and tissue oxygen delivery in patients with trauma. Although RBC warmers may prevent hypothermia, some warming or infusion equipment may lead to haemolysis and patient injury. ObjectivesThe aim of this study was to test the effect of (i) RBC warming and (ii) administration via manual vs. pump infusion on haemolysis. MethodsThis experimental ex vivo study studied haemolysis markers of RBC injury. The sample consisted of 90 RBC infusions in two simulations, randomly, 45 warmed RBC infusions and 45 nonwarmed RBC infusions, in two or three stages: before the intervention (baseline—warming, N= 45; nonwarming, N= 45), after water bath warming at 42 °C (warmed, N= 45), and then after the warmed or nonwarmed RBCs were infused by manual or pump infusion at a rate of 100 mL/h (infusion—warming, N= 45; nonwarming, N= 45). ResultsWarmed RBCs showed significantly lower total haemoglobin (Hb) and haematocrit levels and increase in free Hb levels, haemolysis levels, and lactate dehydrogenase (LDH) activity (all p<0.05) than baseline RBCs. Pump infusion RBCs were associated with reduced total Hb and increased free Hb, haemolysis, and potassium (K) levels (all p<0.05) compared with warmed RBCs. In contrast, manual infusion of warmed RBCs resulted in significantly reduced total Hb levels and increased LDH activity (both <0.05). After infusion, total Hb, free Hb, haematocrit, haemolysis, and LDH values were significantly different for warmed vs. nonwarmed RBCs (p<0.05). ConclusionsHaemolysis biomarkers increase with RBC warming and infusion, especially when using infusion pumps. Critically ill patients should be carefully monitored for possible complications during and after RBC infusion.

Elevated free hemoglobin and decreased haptoglobin levels are associated with adverse clinical outcomes, unfavorable physiologic measures, and altered inflammatory markers in pediatric cardiac surgery patients.

There are data suggesting that free hemoglobin (Hb), heme, and iron contribute to infection, thrombosis, multiorgan failure, and death in critically ill patients. These outcomes may be mitigated by haptoglobin. 164 consecutively treated children undergoing surgery for congenital heart disease were evaluated for associations between free Hb and haptoglobin and clinical outcomes, physiologic metrics, and biomarkers of inflammation RESULTS: Higher perioperative free Hb levels (and lower haptoglobin levels) were associated with mortality, nosocomial infection, thrombosis, hours of intubation and inotropes, increased interleukin-6, peak serum lactate levels, and lower nadir mean arterial pressures. The median free Hb in patients without infection (30 mg/dL; 29 interquartile range [IQR], 24-52 mg/dL) was lower than in those who became infected (39 mg/dL; IQR, 33-88 mg/ 31 dL; p = 0.0046). The median mechanical ventilation requirements were 19 (IQR, 7-72) hours in patients with higher levels of haptoglobin versus 48 (IQR, 18-144) hours in patients with lower levels (p = 0.0047). Transfusion dose, bypass duration, and complexity of surgery were all significantly correlated with Hb levels and haptoglobin levels. Multivariate analyses demonstrated that these variables were independently and significantly associated with outcomes. Elevated pre- and postoperative levels of free Hb and decreased levels of haptoglobin were associated with adverse clinical outcomes, inflammation, and unfavorable physiologic metrics. Transfusion, RACHS score, and duration of bypass were associated with increased free Hb and decreased haptoglobin. Further investigation of the role of hemolysis and haptoglobin as potential mediators or markers of outcomes is warranted.

Hydroxyurea reduces leukocyte interactions with the vessel wall in a haemolytic mouse model via a possible NO/cGMP-mediated effect

Background Haemolysis occurs in a number of haematologic and non-haematologic diseases, including sickle cell disease, malaria and sepsis. Elevated extracellular haemoglobin (Hb) can trigger specific events that are associated with adverse clinical outcomes. For example, in sickle cell disease, cell-free haemoglobin has been proposed to reduce nitric oxide (NO) bioavailability and induce vascular oxidative stress and, possibly, inflammation [1]. Hydroxyurea, a drug used commonly as a therapy for sickle cell disease, may exert some of its effects by acting as a NO donor, in vivo [2]. This study aimed to compare the effects of haemolytic and inflammatory stimuli on blood vessel leukocyte recruitment in vivo. The effects of the acute administration of hydroxyurea (HU) or an NO donor on these alterations were also investigated. Methods Inflammatory or hemolytic processes were induced in C57BL/6 mice (2-4 months old) and cremaster muscles of mice were prepared for intravital microscopy (IVM), according to protocols described in figure 1. Plasma-free Hb was measured with Drabkin’s solution. Results C57BL/6 mice that received water i.v. (H2O mice) presented marked vascular haemolysis after 15 min; levels of plasma Hb were more than doubled in H2O mice, compared to mice that receiving PBS i.v. (2.57±0.43g/dL, 1.08 ±0.08g/dL Hb, respectively, p<0.01, n=3-5), and resembled those of SCD mice (data not shown). In contrast, C57BL/6 mice (n=3) that received an inflammatory stimulus (TNFa) showed no alteration in plasma free Hb levels (1.29 ±0.43 g/dL, 1.23±0.09 g/dL; PBS and TNF-a, respectively). IVM demonstrated that both the inflammatory and haemolytic stimuli induced leukocyte adhesion to vessel walls (3.57±0.84; 11.07±0.92; 7.25±1.20 [100µm -1 ]f or PBS, TNF-a and H2O, respectively, p<0.05, N=15-30 venules), as well as leukocyte extravasation (1.07±0.17; 4.02±0.39; 2.97±0.49 [per 100 µm x 50 µm] for PBS, TNF-a and H2O, respectively, p<0.05, N=15-30 venules). Surprisingly, when HU was given simultaneously with TNF-a or following H2O, this drug was able to prevent leukocyte recruitment in both models, reducing both leukocyte adhesion (7.40±0.47; 2.79±0.34 [100 µm -1 ] for TNF-a+PBS and TNF-a+HU, respectively, p<0.0001, n=30-60 venules; and 7.84±0.73; 2.41±0.37 for H2O+PBS and H2O+HU, p<0.0001, n=20-30 venules) and leukocyte extravasation (3.88±0.26; 1.22±0.18 for TNF-a+PBS and TNF-a+HU, p<0.0001, n=30-60 venules and 2.98±0.32; 1.94±0.21 for H2O+PBS and H2O+HU, p<0.0001, n=20-30 venules) and increased leukocyte rolling (18.11±1.58; 25.48±5.01 for TNF-a+PBS and TNF-a+HU, respectively, p<0.05, n=30-60 venules and 11.56±1.47; 32.47±3.54 for H2O+PBS and H2O+HU, p<0.0001, n=20-30 venules). Additionally, DEANO was also able to reverse the inflammatory process installed by TNF-a, reducing leukocyte adhesion (10.24 ±1.49; 11.45±1.40 for TNF-a before and after vehicle control, n=21-22 venules; 7.63±0.63; 3.76±0.41 for TNF-a, before and after DEANO, p<0.01; n=29-35 venules) and extravasation (1.95±0.29; 3.23±0.44 before and after vehicle control, p<0.05, n=21-22 venules; 2.49±0.25; 2.24±0.24 before and after DEANO, n=29-35 venules).

Patients with sickle cell anemia on simple chronic transfusion protocol show sex differences for hemodynamic and hematologic responses to transfusion

Chronic transfusion therapy (CTT) is a mainstay for stroke prophylaxis in sickle cell anemia, but its effects on hemodynamics are poorly characterized. Transfusion improves oxygen-carrying capacity, reducing demands for high cardiac output, while decreasing hemoglobin (Hb)S%, reticulocyte count, and hemolysis. We hypothesized that transfusion would improve oxygen-carrying capacity, but that would be counteracted by a decrease in cardiac output due to increased hematocrit (Hct) and vascular resistance, leaving oxygen delivery unchanged. To test this hypothesis, we examined patients on CTT immediately before transfusion and again 12 to 120 hours after transfusion, using echocardiography and near infrared spectroscopy. Comparable increases in Hb and Hct and decreases in reticulocyte count and HbS with transfusion were observed in all patients, but males had a larger rebound of HbS%, reticulocyte count, and free Hb levels between transfusions. In males, transfusion decreased heart rate by 12%, stroke volume by 15%, and cardiac index by 24% while estimates for pulmonary and systemic vascular resistance increased, culminating in 6% decrease in oxygen delivery. In contrast, stroke volume and cardiac index and systemic and pulmonary vascular resistance did not change in women after transfusion, such that oxygen delivery improved 17%. In our sample population, males exhibit a paradoxical reduction in oxygen delivery in response to transfusion because the increase in vascular resistance is larger than the increase in oxygen capacity. This may result from an inability to adequately suppress their HbS% between transfusion cycles.

Sports anaemia: facts or fiction?

Sports anaemia: facts or fiction?

Hemolytic Effects of a Football-Specific Training Session in Elite Female Players

This study aimed to quantify the hemolytic responses of elite female football (soccer) players during a typical weekly training session. Ten elite female football players (7 field players [FPs] and 3 goalkeepers [GKs]) were recruited from the Australian National Women's Premier League and asked to provide a venous blood sample 30 min before and at the immediate conclusion of a typical weekly training session. During this training session, the players' movement patterns were monitored via a 5-Hz global positioning system. The blood samples collected during the training session were analyzed for iron status via serum ferritin (SF) analysis, and the hemolytic response to training, via serum free hemoglobin (Hb) and haptoglobin (Hp) measurement. 50% of the participants screened were found to have a compromised iron stores (SF<35 μg/L). Furthermore, the posttraining serum free Hb levels were significantly elevated (P=.011), and the serum Hp levels were significantly decreased (P=.005), with no significant differences recorded between the FPs and GKs. However, the overall distance covered and the movement speed were significantly greater in the FPs. The increases in free Hb and decreases in Hp levels provide evidence that a typical team-sport training session may result in significant hemolysis. This hemolysis may primarily be a result of running-based movements in FPs and/or the plyometric movements in GKs, such as diving and tackling.

Time-related hemolysis in stored shed mediastinal blood after cardiopulmonary bypass

Reinfusion of mediastinal shed blood during cardiopulmonary bypass reportedly reduces the need for homologous blood transfusion. Although the fragility of blood components is thought to be amplified by shear stress during cardiopulmonary bypass and processing, the time-related deterioration of red blood cells (RBCs) in stored shed blood has not been studied extensively. In this study, we examined time-related hemolysis in shed blood stored at different temperatures. We examined processed shed blood collected from 15 patients (11 men and 4 women; mean age±standard deviation, 71±9years) during cardiopulmonary bypass. The shed blood was collected and stored at 20°C (group A) or 4°C (group B). Stored blood collected by venipuncture at the end of the surgery was used as a control. Damage was assessed by measuring its free hemoglobin (Hb) levels, using a photometric assay. The free Hb levels in blood samples from each group were tested at 0, 3, 6, 12, 24, 36, and 48h after surgery. The free Hb levels (g/dl) at 0, 12, and 24h were 0.03±0.01, 0.05±0.02*, and 0.06±0.02* in group A; 0.03±0.02, 0.04±0.03, and 0.05±0.02* in group B; and 0.01±0.01, 0.01±0.01, and 0.01±0.01 in the control group (*p<0.05 vs. 0h after surgery). The free Hb levels in stored shed blood significantly increased after 12h in group A (20°C) and increased after 24h in group B (4°C), whereas in drawn blood, they did not significantly increase over the first 24h. Compared to storage at 20°C, storage at 4°C suppresses the increase in the free Hb levels.

Serum free hemoglobin as a novel potential biomarker for acute ischemic stroke

Currently, no practical biomarker is available for the diagnosis of acute ischemic stroke. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has being applied extensively to analyze large biomolecules such as proteins. The technique is likely to be of remarkable value as indicators of systemic processes such as atherosclerosis and stroke. The aim of this study is to identify potential protein biomarkers for ischemic stroke diagnosis utilizing MALDI-TOF MS. Serum samples obtained from acute ischemic stroke patients (n = 47) and controls (n = 34) were analyzed by MALDI-TOF MS. Stroke severity was assessed by the National Institutes of Health Stroke Scale (NIHSS), subtype by the Trial of Org 10172 in Acute Stroke Treatment (TOAST), functional outcome by the modified Rankin Scale (mRS) and infarct volume by the diffusion-weighted images. Risk factors and routine laboratory data of the stroke patients were registered prospectively. The peaks of hemoglobin (Hb) alpha-chain and beta-chain were differentially expressed between stroke patients and controls (p < 0.0001). Hb ions were detected in the samples collected from 33 (70.2%) stroke patients and 5 (14.7%) controls. The sensitivity is 70.2% and the specificity is 85.3%. Among stroke patients, there is no significant correlation (p > 0.05) between Hb peaks and the NIHSS, TOAST, mRS, stroke risk factors, infarct volume, infarct location and laboratory data. Serum free Hb may serve as a novel potential biomarker for the diagnosis of acute ischemic stroke. The clinical value of this potential biomarker may be clarified by further studies quantifying serum free Hb levels.

Severe Malaria and Leptospirosis Are Associated with a Deficiency of the Von Willebrand Factor Cleaving Protease, ADAMTS13

The thrombotic microangiopathy observed in severe malaria and leptospirosis shows resemblance to that of thrombotic thrombocytopenic purpura (TTP) and haemolytic uremic syndrome (HUS), respectively. The pathophysiology of these complications remains incompletely understood. TTP and (in some cases) HUS are associated with a deficiency of ADAMTS13, a metalloprotease that cleaves large prothrombotic VWF multimers. We hypothesized that ADAMTS13 may be deficient in cerebral malaria and leptospirosis, and that the severity of ADAMTS13 deficiency may be associated with the degree of endothelial activation. We investigated a series of patients with cerebral malaria (n=9) and leptospirosis (n=18). Patients with scrub typhus (n=12) representing a disease that is accompanied by endothelial activation, but without the clinical features of TTP-HUS, and healthy blood donors served as controls. Baseline characteristics did not significantly differ between study groups, except for BUN values which were higher in cerebral malaria than in leptospirosis (p-value 0.046). The Glasgow Coma Scale (GCS) score was evidently lower in cerebral malaria, with a median score of 4 (interquartile range 3–6) while subjects with leptospirosis, scrub typhus and normal controls had (near) maximum scores of 15. ADAMTS13 levels were significantly reduced in both cerebral malaria and leptospirosis, compared to scrub typhus and normal controls, with a significantly more severe deficiency in cerebral malaria than in leptospirosis (data in figure). VWF antigen and VWF propeptide levels were elevated in all three disease groups in comparison to healthy controls (p-values < 0.001), and most markedly increased in cerebral malaria. ADAMTS13 activity inversely correlated with both VWF antigen and VWF propeptide levels (p-values <0.001 and 0.005). No significant correlation was apparent between ADAMTS13 activity and hemoglobin (Hb) concentration or platelet count. Free Hb levels were not elevated in any of the patient groups, hence it seems highly unlikely that free Hb is involved in the observed reduction of ADAMTS13 activity in cerebral malaria and leptospirosis. The results of this study indicate that ADAMTS13 levels are decreased in patients with cerebral malaria and leptospirosis providing direct support to the hypothesis of a role of ADAMTS13 in the pathogenesis of the thrombotic microangiopathy associated with these diseases. Further research seems warranted to establish the role of ADAMTS13 deficiency in the development and the severity of these diseases, and to evaluate its prognostic significance. Potentially, ADAMTS13 function might serve as a risk marker of disease severity and it may be speculated that the administration of exogenous (recombinant) ADAMTS13 can be used as an adjunctive therapeutic strategy in the clinical management of patients with cerebral malaria and severe leptospirosis. [Display omitted]

Therapeutic Inhibition of Endothelial Cell Tissue Factor Expression In Vivo by Nitric Oxide and Arginine in Sickle Transgenic Mice.

Sickle cell anemia is accompanied by biochemical activation of the coagulation system and by clinical thromboses. Therefore, we have focused on the expression of tissue factor (TF), the trigger of the coagulation system, in this disease. Previous studies from our laboratories described abnormal TF expression by monocytes and by circulating endothelial cells in human sickle blood. More recently, we reported (Blood 104:840, 2004) that severe-phenotype (BERK) sickle mice abnormally express TF on pulmonary vein endothelium; mild-phenotype (NY1DD) sickle mice are normal in this regard but assume the abnormal phenotype after exposure to hypoxia (3 hr at 8% O2) followed by reoxygenation for 18 hr at ambient air (H/R). Monitoring the % of pulmonary veins positive for TF on triple stained tissue sections (for nuclei, for murine TF, for murine CD31 to identify endothelial cells), we have now evaluated the role of NO in TF expression, employing these two models. In the NY1DD H/R model, NO inhalation (34 ppm) administered during the entire reoxygenation period led to 68±18% reduction (P<.001) of TF expression. Consistent with this, dietary supplementation with arginine (substrate for nitric oxide synthase [NOS]) prior to H/R led to 93±25% reduction (P<.001) of the H/R induced TF response. Conversely, dietary supplementation with L-NAME (inhibitor of NOS) converted the NY1DD mouse to a TF positive phenotype (P=.047) without even requiring H/R. Interestingly, inhalation of NO was not protective in the NY1DD H/R model if given only during the hypoxia period itself or only during the last 3 hours of the reoxygenation period, but it was protective if given only during the first 3 hours of the reoxgygenation period (68±24% reduction; P=.0008). Thus, the timing of clinical intervention is probably critical. In the BERK model at ambient air, treatment with dietary arginine for 2 weeks diminished TF expression by 60±33% (P=.04), an effect that was countervailed by concurrent administration of L-NAME. Data will soon be available on eNOS over-expressing sickle mice. Also in BERK mice, L-NAME largely prevented the TF inhibiting therapeutic benefit of lovastatin, suggesting that lovastatin's beneficial effect on TF is mediated via NO biology. These results suggest that NO biology determines endothelial TF expression in vivo in sickle mice. We note that NO is reported to suppress endothelial TF expression in vitro. We find that sickle mice have elevated plasma free Hb levels (increased by 40% in NY1DD mice and 57% in BERK mice), a risk factor for diminished NO bioavailability. On the other hand, H/R in NY1DD mice is not associated with a further increase in free plasma Hb. So it seems that the NO biology of sickle mice is not simply explained by plasma Hb. NO biology seems to exert a major role in regulating endothelial TF expression in the sickle mouse. Long-term NO modulating therapy, such as dietary arginine or lovastatin, may be beneficial in terms of the coagulopathy and thrombosis risk of sickle patients and should be tested with this in mind.

Transfusions via hand-held syringes and small-gauge needles as risk factors for hyperkalemia.

Pediatric emergency RBC transfusions are often infused rapidly through 22-gauge (ga) or smaller needles or catheters using hand-held syringes. Data relating needle size, unit age, and infusion rate are needed to assess the risk of hemolysis and hyperkalemia in this setting. Multiple simulated transfusions were performed during storage of RBC units. Aliquots from five units were rapidly passed through needles (18, 20, 22-25 ga) using a hand-held syringe. Resulting plasma Hb and K+ concentrations were measured. Free Hb levels were used as a measure of needle-associated hemolysis (NAH). Passage through 18-ga and 20-ga needles caused no hemolysis, but rapid passage through 23-ga, 24-ga, and 25-ga did. RBCs stored less than 7 days showed significant K+ release with 23- to 25-ga needles. The greatest needle-associated K+ release was 10 mEq per L, on Day 5. Due to high K+ concentrations resulting from spontaneous efflux, K+ release from NAH was not detectable after 2 or more weeks of storage. Rapidly transfusing RBCs using hand-held syringes through 23-ga or smaller needles can cause hemolysis. In RBCs stored 2 weeks or more, NAH does not measurably increase K+ concentrations above that present from storage-related efflux. During rapid transfusions, RBC storage time is the primary risk factor for transfusion-associated hyperkalemia.

EVAHEART: an implantable centrifugal blood pump for long-term circulatory support.

We developed "EVAHEART": a compact centrifugal blood pump system as an implantable left ventricular assist device for long-term circulatory support. The 55 x 64 mm pump is made from pure titanium, and weighs 370 g. The entire blood-contacting surface is covered with an anti-thrombogenic coating of diamond like carbon (DLC) or 2-methacryloyloxyethyl phosphorylcholine (MPC) to improve blood compatibility. Flows exceeding 12 L/min against 100 mmHg pressure at 2600 rpm was measured. A low-temperature mechanical seal with recirculating cooling system is used to seal the shaft. EVAHEART demonstrated an acceptably low hemolysis rate with normalized index of hemolysis of 0.005 +/- 0.002 g/100L. We evaluated the pump in long-term in-vivo experiments with seven calves. Via left thoracotomy, we conducted left ventricular apex-descending aorta bypass, placing the pump in the left thoracic cavity. Pump flow rates was maintained at 5-9 L/min, pump power consumption remained stable at 9-10 W in all cases, plasma free Hb levels were less than 15 mg/dl, and the seal system showed good seal capability throughout the experiments. The calves were sacrificed on schedule on postoperative day 200, 222, 142, 90, 151, 155, and 133. No thrombi formed on the blood contacting surface with either the DLC or MPC coating, and no major organ thromboembolisms occurred except for a few small renal infarcts. EVAHEART centrifugal blood pump demonstrated excellent performance in long-term in-vivo experiments.

An implantable centrifugal blood pump with a recirculating purge system (Cool-Seal system).

A compact centrifugal blood pump has been developed as an implantable left ventricular assist system. The impeller diameter is 40 mm, and pump dimensions are 55 x 64 mm. This first prototype, fabricated from titanium alloy, resulted in a pump weight of 400 g including a brushless DC motor. The weight of a second prototype pump was reduced to 280 g. The entire blood contacting surface is coated with diamond like carbon (DLC) to improve blood compatibility. Flow rates of over 7 L/min against 100 mm Hg pressure at 2,500 rpm with 9 W total power consumption have been measured. A newly designed mechanical seal with a recirculating purge system (Cool-Seal) is used for the shaft seal. In this seal system, the seal temperature is kept under 40 degrees C to prevent heat denaturation of blood proteins. Purge fluid also cools the pump motor coil and journal bearing. Purge fluid is continuously purified and sterilized by an ultrafiltration unit which is incorporated in the paracorporeal drive console. In vitro experiments with bovine blood demonstrated an acceptably low hemolysis rate (normalized index of hemolysis = 0.005 +/- 0.002 g/100 L). In vivo experiments are currently ongoing using calves. Via left thoracotomy, left ventricular (LV) apex descending aorta bypass was performed utilizing an expanded polytetrafluoroethylene (ePTFE) vascular graft with the pump placed in the left thoracic cavity. In 2 in vivo experiments, the pump flow rate was maintained at 5-9 L/min, and pump power consumption remained stable at 9-10 W. All plasma free Hb levels were measured at less than 15 mg/dl. The seal system has demonstrated good seal capability with negligible purge fluid consumption (<0.5 ml/day). In both calves, the pumps demonstrated trouble free continuous function over 6 month (200 days and 222 days).

Blood Interaction with a Bioline Heparin Coated HIA-VAD: A Study on Calves

The blood compatibility of ventricular assist devices developed by the Helmholtz Institute Aachen (HA-VAD's) was tested on calves. Seven calves received a non-coated HIA-VAD (control) and three a Bioline heparin coated device. The circulatory support of these HIA-VAD's lasted one week. Mechanical blood cell trauma estimated by hematocrit (Hct), hemoglobin (total Hb) and free plasma hemoglobin (free Hb) levels did not differ in either group. All HIA-VAD's in the control group remained thrombus free, except on one occasion when an inflow cannula was obstructed by a thrombus located in the tip. After circulatory support, the animals in this group seemed clinically healthy. However, thrombus formation was observed in the three heparin coated HIA-VAD's. One animal in this group died from complications after re-operation for pneumothorax on the fifth day of support, whereas the other two animals seemed clinically healthy. In these three animals, a strong decrease in platelet numbers was measured even after 24 hours of support which recovered after 72 hours. This decrease in platelet numbers was associated with a lower degree of platelet aggregation ability stimulated by ADP (p < 0.05). Fibrin(ogen) degradation products (FDP) increased significantly immediately after the implantation procedure (p < 0.05). Fibrinogen levels initially decreased during the implantation procedure, but increased thereafter in both groups. The FDP levels remained high in this group, although the FDP levels in both groups were decreased after the implantation procedure. The ex vivo measured circulating heparin levels were lower in the heparin coated HIV-VAD group despite the equally administrated heparin doses in both animal groups. No differences were measured in either group with regard to white blood cell (WBC) numbers and complement hemolytic activity (CH50). Despite these hemostatic changes, no mechanical trauma could be demonstrated after seven days of circulatory support.

Le délai de péremption du sang décongelé: Nouvelles perspectives

In order to comply with the needs for delayed transfusion in current practice (rare blood types dispatched to other centers, thawed blood not finally transfused to a patient, blood bank supply of Rh negative blood), we have investigated preservative media for thawed blood. We have previously shown that thawed RBC concentrates resuspended in isologous or autologous plasma remain viable and functional for 72 hours. We have extended these investigations by resuspending thawed RBC concentrates in an original synthetic preservative medium designed in our laboratory, containing: (table; see text) The following parameters have been investigated before freezing and during 15 days following thawing and washing: sterility, pH and 2,3-DPG, ATP and energy charge, free Hb levels in the supernatant, 24 h post-transfusion life and half-life of 51Cr-labeled RBC. This preservative allows the conservation of thawed RBC for up to nine days at +4 degrees C. During that time, sterility is maintained, pH is 6.80, 2,-3-DPG levels are 50% of the original values, ATP levels are 100% of the original values, free Hb is 122 mg per blood unit, energy charge is 0.90; RBC labeled 7 days after thawing showed a 89% survival rate 24 h after transfusion and a half-life of 19 days.