Sickle Cell Trait Donors Research Articles

Assessment of amino acids and metabolites in the supernatant of stored concentrates blood from sickle cell trait (SCT) and reference (non-SCT) donors.

Sickle cell trait (SCT) persons are significant donors, and discarding these blood units reduces their supplies, mainly in the third-world countries. This work focused on 12 metabolites associated with the red blood cell (RBC) storage lesion and 23 amino acids in the supernatants of packed RBC units from SCT and reference (non-SCT) donors stored in the same conditions. All samples of RBC concentrates were collected and separated from the storage of Colsan (Beneficient Association of Blood Collection), where they were routinely processed and separated as packed RBC units and stored in the refrigerator (2°-6°C). The supernatant samples of each packed RBC bag were separated by centrifugation at days 1, 7, 14, 21, 28 and 35 of storage and kept at -80°C till the metabolite analysis together. The quantitation of metabolites and amino acids examined in the supernatant of SCT and reference donors showed no statistical differences along the cold storage. Lactic acid and malic acid releases occur in three phases during RBC storage. Basic and acid amino acids and corresponding amides have low and stable values during the first 14 days of storage, followed by a steep increase. Our metabolomic results give elements that seem not to contraindicate the transfusion of RBC with SCT, besides its more structural fragility.

Donors with repeated blood product discards for filtration problems, clots or hemolysis: Causes and follow-up.

Sanquin donor medicine department is informed when donations or their components are rejected. This can occur isolated or frequently. It is undesirable because the donations cannot be used and there may be an underlying medical cause. Based on regional approaches, a uniform procedure was developed. Information about whole blood, plasma- plateletpheresis donations from which one or more components were rejected for filtration time (>2 h), hemolysis or clots were extracted from blood bank information system. After rejection of two successive components or donations or total ≥3 the donor is contacted. Depending on the medical history and investigation by the family doctor, the donor carrier is re-evaluated. We looked for the causes of the discarded products and performed a survey among blood services regarding polices with discarded products. One or more components from 1742 of about 2.2 million successful donations (0.08%) were rejected. The highest percentage of rejection was seen in plateletpheresis (1.5%), all for clots. No underlying medical causes were found. 24 whole blood donors were found to have sickle cell trait (SCT) and were permanently deferred. The policies for follow-up after discarded products or acceptance of SCT donors vary between the 16 blood banks. Six organizations do not follow-up donors and seven accept SCT for blood or plasma donation. Informing donors with repeated discarded products avoids the non-use of donations. Causes of repeated discarded products can be found by follow-up of donors. The results of the survey indicate a large discrepancy in policies applied worldwide.

Feasibility of peripheral blood stem cell collection from sickle cell trait donors with an intensified G-CSF regimen.

Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative treatment for SCD and bone marrow from an HLA-matched sibling is currently the standard of care. Haploidentical HSCT from a family donor with a TCR αβ/CD19 depleted graft (T-haplo) is an increasingly successful alternative, which requires the generation of G-CSF stimulated peripheral stem cell (PBSC) from haploidentical relatives. These sickle cell trait (SCT) donors reported to develop SCD-related complications in conditions of severe stress. In this retrospective analysis, we compared the safety and efficacy of PBSC mobilization with a G-CSF intensified mobilization regimen in SCT donors with a conventional G-CSF mobilization regimen in healthy donors. The reported adverse events were similar during intensified G-CSF mobilization, apheresis, and shortly after stem cell apheresis in SCT and control donors. In SCT and control donors, we were able to mobilize high yields of CD34+ stem cells and the harvested CD34+ cell count was comparable with control donors. Peripheral stem cell mobilization using an intensified G-CSF regimen is safe, and well tolerated among SCT donors. SCT donors are a valid alternative for collection of peripheral CD34+ stem cells for T-cell-depleted haploidentical stem cell transplantation.

Storage related haematological and biochemical changes in Plasmodium falciparum infected and sickle cell trait donor blood

BackgroundIn sub-Saharan Africa where sickle cell trait (SCT) and malaria is prevalent, significant proportions of blood donors may be affected by one or more of these abnormalities. The haemato-biochemical properties of SCT and asymptomatic malaria in donor blood have not been evaluated. This study evaluated the haemato-biochemical impact of SCT and asymptomatic malaria infections in citrate-phosphate-dextrose-adenine (CPDA-1) stored donor blood units.MethodsFifty-milliliters of sterile CPDA-1 anti-coagulated blood were drained into the sample pouch attached to the main blood bag. Ten units each of sickle cell/malaria negative, sickle cell and malaria positive blood were analyzed. Baseline and weekly haematological profiling and week 1, 3 and 5 concentrations of plasma haemoglobin, % haemolysis, sodium, potassium and chloride and lactate dehydrogenase (LDH) were assayed. Differences between baseline and weekly data were determined using one-way analysis of variance (ANOVA) and Kruskal-Wallis test, whereas differences between baseline parameters and week 1–3 data pairs were determined using paired t-test. P-value < 0.05 was considered statistically significant.ResultsStorage of SCT and malaria infected blood affected all haematological cell lines. In the SCT donors, red blood cells (RBC) (4.75 × 1012/L ± 1.43baseline to 3.49 × 1012/L ± 1.09week-5), haemoglobin (14.45 g/dl ± 1.63baseline to 11.43 g/dl ± 1.69week-5) and haematocrit (39.96% ± 3.18baseline to 33.22% ± 4.12week-5) were reduced. In the asymptomatic malaria group, reductions were observed in RBC (5.00 × 1012/L ± 0.75baseline to 3.72 × 1012/L ± 0.71week-5), haemoglobin (14.73 g/dl ± 1.67baseline to 11.53 g/dl ± 1.62week-5), haematocrit (42.72% ± 5.16baseline to 33.38% ± 5.80week-5), mean cell haemoglobin concentration (35.48 g/dl ± 1.84baseline to 35.01 g/dl ± 0.64week-5) and red cell distribution width coefficient of variation (14.81% ± 1.54baseline to 16.26% ± 1.37week-5). Biochemically, whereas plasma LDH levels significantly increased in asymptomatic malaria blood donors (319% increase at week 5 compared to baseline), SCT blood donors had the most significant increase in plasma potassium levels at week 5 (382% increase). Sodium ions significantly reduced in SCT/malaria negative and sickle cell trait blood at an average rate of 0.21 mmol/L per day. Moreover, elevations in lymphocytes-to-eosinophils and lymphocytes-to-neutrophils ratios were associated with SCT and malaria positive blood whilst elevation lymphocytes-to-basophils ratio was exclusive to malaria positive blood.ConclusionSevere storage lesions were significant in SCT or malaria positive donor blood units. Proper clinical evaluation must be done in prospective blood donors to ensure deferral of such donors.

At Least 20% Donor Myeloid Chimerism Is Necessary to Reverse the Sickle Phenotype after Allogeneic Hematopoietic Stem Cell Transplantation

Hematopoietic stem cell transplantation (HSCT) is the only curative option for patients with sickle cell disease (SCD). HLA-matched sibling HSCT has become increasingly successful, and haploidentical approaches are being explored to increase the donor pool. Advances have also been made in gene therapy such that clinical trials are now underway for the first time in patients with SCD. The question of what percentage of donor chimerism is necessary to cure SCD is critical in identifying the target percentage of the transferred globin gene needed to reverse SCD. One study showed that a whole blood donor chimerism level as low as 11% was sufficient to render patients free of SCD1; however, lineage specific chimerism was not performed. We have previously demonstrated that peripheral blood myeloid chimerism reflects the HSC compartment. We sought to determine the percentage of donor myeloid chimerism (DMC) necessary to reverse the sickle phenotype in our patients who underwent HSCT.In protocols testing both HLA-matched and haploidentical nonmyeloablative HSCT in over 70 patients with SCD, we identified 3 who despite robust initial engraftment experienced declining DMC levels over prolonged follow up eventually associated with return of SCD. All 3 patients’ donors had sickle cell trait with HbS ranging from 36-40%. Percentages of DMC and HbS were monitored at day 100 and at least every 6-12 months post-transplant thereafter. The 3 patients initially achieved nearly 100% DMC; however, their chimerism levels slowly decreased over time (see Figure 1). As long as mean DMC was >20%, mean %HbS remained less than 50% (similar to their sickle cell trait donors) and the patients stayed free of SCD-related symptoms. However, mean DMC at 2, 3, 4, and 5 years post-transplant fell below 20%, were 18, 12, 11, and 10%, and were associated with mean %HbS levels of 52.5, 51.6, 57.8, and 64.2%. Further, 2 of the patients had return of their recurrent painful crises, and the third patient with moyamoya syndrome developed severe anemia requiring re-initiation of chronic transfusion therapy.We subsequently developed a mathematical model to test the hypothesis that the percentage of necessary DMC is determined solely by the differences between donor and recipient red blood cell (RBC) half-lives. The model incorporates donor and recipient stem cell kinetics (self renewal and differentiation), which are assumed to be identical, as well as donor and recipient RBC survival times, which are known to be different. The model assumes first order kinetics for erythropoiesis and RBC senescence/destruction. It predicts the fraction of donor RBC as a function of DMC and donor/recipient RBC lifetimes. Although we did not directly measure RBC lifetimes, we identified 4 patients with SCD in the literature where reticulocyte percentages ranged from 9.0 to 21.0% and corresponding RBC half-lives determined by radioactive sodium chromate were 1 to 11 days2. We assumed that in erythropoietic homeostasis, RBC lifetime must be proportional to the inverse of the reticulocyte fraction; and we confirmed this relationship using data from these 4 patients with a good coefficient of determination (R2= 0.91). Therefore we reasoned that (at stable hemoglobin concentration) the reticulocyte fraction is a good proxy for RBC lifetime. Figure 1 depicts a comparison between the measured and our model predicted HbS fractions in our 3 patients transplanted with sickle cell trait donors using a mean sickle RBC lifetime of 7 days. As observed in our 3 patients, our model predicts that DMC of at least 20% is necessary to reverse the sickle phenotype.Therefore, our model predicts that genetic strategies aimed at SCD must achieve correction levels of at least 20% to reverse the sickle phenotype. Further, the mechanism explaining why a minority of donor myeloid cells is adequate is due soley to the vast differences in RBC survival between donor and recipient cells. Lastly, recipient RBC survival is lowest in patients with the highest reticulocyte counts suggesting that gene therapy may be most efficacious in subjects with high reticulocyte percentages as our model predicts that lower levels of correction would be sufficient in this setting. Further studies are indicated to validate our findings.1. Walters MC et al. Biol Blood Marrow Transplant. 2001;7(12):665-673.2. Weinstein IM et al. Blood. 1954;9(12):1155-1164. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Collection and Processing Outcomes of G-CSF Mobilized Peripheral Blood Stem Cells (PBSC) Collected from Parental Haploidentical Stem Cell Sickle Cell Trait Donors of Patients Undergoing CD34 Enriched Transplantation for High-Risk Sickle Cell Disease (IND

Collection and Processing Outcomes of G-CSF Mobilized Peripheral Blood Stem Cells (PBSC) Collected from Parental Haploidentical Stem Cell Sickle Cell Trait Donors of Patients Undergoing CD34 Enriched Transplantation for High-Risk Sickle Cell Disease (IND

A Comparative Study of Solubility Based Sickle Cell Screening Versus Genomic Method in Blood Donors

Background:Sickle cell trait (SCT) is more common in minority African-Americans (AA) and Hispanic blood donors. Screening donors for Sickle Hemoglobin (HbS) carrier status (SCT) can be useful, especially in managing the transfusion needs of neonates and patients with sickle cell disease. Currently, the Sickledex test, based on the principle of relative insolubility of HbS in phosphate buffers, is routinely used for HbS donor screening. However, there are several limitations including the inability to perform high-throughput testing by automation as donor centers target larger numbers of minority blood donors. False positives results due to hyperlipidemia, dysglobulinemia and higher peripheral blood hematologic counts have also been reported. Hence alternative methods to identify SCT donors are highly desirable in blood centers. The purpose of this study was to validate testing, and to identify rates of False Positive (FP) and False Negative (FN) sickledex screening in blood donors by verifying the presence or absence of the most common mutation seen with HbS production using molecular methods. The comparison could offer evidence to use molecular methods as a test of record to manage HbS carriers at blood centers.Methods and ResultsBetween Jan 2012 till Dec 2014, 200 sickledex screening tests were done daily, totaling approximately 90,000 donors self-identifying as Caucasians and 60,000 donors identifying as African American or Hispanic tested during the study period. Amongst Caucasians tested, 154 had a positive sickledex screen result. Since the prevalence of SCT is less common in the Caucasian population, we hypothesized several of these Caucasian donors could have a false positive screening test. All 154 sickledex positive Caucasian donor samples were prospectively tested for the HbS mutation detection with molecular methods. BioArray Human Erythrocyte Antigen (HEA) Bead Chip DNA assay with PCR multiplex based technology was used to detect the HbS mutation (β-globin change -173 A>T, more commonly designated 20A>T). Of the 154 sickledex + Caucasians, 108 (70%) donors tested negative (FP Screen) and 46 (30%) tested positive for the HbS mutation. A retrospective review of sickledex results in our electronic database for minority donors (n-3431) who also had HEA BeadChip assay was performed to determine concordance. Of 3431 minority donors tested, only 2 (0.0006%) had a positive sickledex screening and were negative by HEA, with the remaining negative by both the sickledex screen and HEA tests. Among minority donors (n= 254) testing + for HbS by HEA, 3% (8/254) were negative by sickledex screening. To establish concordance of HEA assay with high performance liquid chromatographic (HPLC) assay, two separate cohorts were evaluated. Initially 12 sickle screen and HEA positive donors was subject to HPLC analysis, which confirmed SCT status in all 12 samples. Subsequently sickledex screen positive but HEA negative samples (n= 40), were subject to HPLC. All 40 showed absence of SCT by HPLC. Finally, 58 random sickledex + samples had gene sequencing and HEA performed in parallel. Out of 58 samples, 54 showed a positive HbS mutation by HEA with A>T (SCT) nucleotide change noted with gene sequencing. In the remaining 4 samples, HEA was negative with gene sequencing showing (3 T>T and in 1C>T) change at nucleotide 163 associated with Hemoglobin Okayama.Conclusion:The HEA bead chip assay testing for the HbS mutation showed 100% concordance with HPLC and gene sequencing in samples studied. Molecular methods can process multiple samples in a single setting to detect SCT. FP sickledex screening results in Caucasians needs confirmatory testing for the HbS mutation to avoid unnecessary donor notification and deferral. The association, if any, of false positive sickledex screening in Caucasian donors with the Hgb Okayama mutation is under further investigation. DisclosuresNo relevant conflicts of interest to declare.

Glucose-6-Phosphate Dehydrogenase (G6pd) Deficiency and Sickle Cell Trait among Blood Donors in Nigeria

This study is being aimed to determine the prevalence of glucose-6-phosphate dehydrogenase deficiency and sickle cell trait among blood donors. Blood donation from glucose-6-phosphate dehydrogenase (G6PD)-deficient and sickle cell trait (SCT) donors might alter the quality of the donated blood during processing, storage or in the recipient′s circulatory system. It has been proposed that several biochemical changes and depletion in the antioxidant defense system occur on storage of G6PD-deficient blood. Red blood cells collected from sickle cell trait donors frequently occlude white blood cells reduction filters; the main cause of this filtration failure is haemoglobin polymerization. A total of 314 blood donor samples were collected from prospective and healthy blood donors after proper screening. Glucose-6-Phosphate dehydrogenase was determined using two standard methods; methaemoglobin reduction test and fluorescent spots test; haemoglobin variants were determined using cellulose acetate electrophoresis at alkaline PH 8.6. Out of the 314 blood donors, 80 (25.5%) were G6PD deficient, 104 haemoglobin variants AS, AC and SC were 82 (78.8%), 21 (20.2%) and 1 (1.0%) respectively. Commercial blood donors had highest G6PD deficient and highest haemoglobin variants. Age group 25-34 had highest G6PD deficient prevalence while age group 15-24 had highest haemoglobin variants. Transfusion with G6PD-deficient blood carries a potential risk of hemolytic complications, especially if it is used for exchanged blood transfusion in neonates. On the other hand, the blood donated from SCT donors, apart from its undesired effects if transfused to sickle cell disease patients, also leads to WBC filtration failure.

Assessment of leucoreduction of sickle cell trait blood: quality of the filtered product.

With the implementation of universal leucoreduction of blood components in several industrialised countries, the problems associated with leucocyte filtration of sickle cell trait blood have been reconsidered. In this study, we assessed the use of high performance filters for leucoreduction of packed red blood cells donated from subjects with sickle cell trait and evaluated the incidence and recurrence of altered red blood cell filterability. Twenty-one volunteer donors with HbAS were compared to 21 donors with HbAA selected at random. The main parameters analysed were residual white blood cell count and post-filtration haemolysis. Filtration times, flow, volume and haemoglobin loss of the packed red blood cells were also determined. In all, 33% of HbAS red blood cell units with slow flow and prolonged filtration time had high residual white blood cell counts. In 7.7% of cases, despite flow through the filter, the units were not leucoreduced properly. Haemoglobin and volume loss were significantly greater in the slow filtration group. Significant post-filtration haemolysis was present in half of the units with high residual white blood cell counts. Despite the development of new technology for filtration, the problem of filterability of blood from donors with sickle cell trait is not yet resolved. Altered filterability of blood from sickle cell trait donors cannot be predicted from the donors' characteristics and recurrence of the problem is not observed between donations. Screening blood donors for sickle cell trait to ensure the safety and quality of blood products for transfusion does, therefore, remain a relevant issue.

A Pilot Study of Reduced Toxicity Conditioning (RTC) with Busulfan (Bu), Fludarabine (Flu), and Alemtuzumab Followed by Allogeneic Stem Cell Transplantation (AlloSCT) to Induce Sustained Mixed Donor Chimerism In Patients with Symptomatic Sickle Cell Disease (SCD)

AbstractAbstract 3504The only known curative therapy for patients with SCD is a matched family donor AlloSCT (Walters et al, NEJM, 1996). Complications after myeloablative AlloSCT are numerous and include death, graft versus host disease (GVHD), and infections, making reduced toxicity conditioning regimens more appealing. A major obstacle allowing most SCD patients to undergo SCT is the lack of an unaffected, HLA-matched identical sibling (Bhatia et al, BMT, 2008). Umbilical cord blood (UCB) has been proven to be an excellent alternate donor source that can safely reconstitute hematopoiesis after AlloSCT in pediatric recipients with non-malignant conditions (Cairo et al, BBMT, 2008). In this study, we report the results of a RTC regimen followed by an AlloSCT from matched family and unrelated UCB donors in selected patients with symptomatic SCD. Between August 27, 2004 and March 2, 2010, 18 patients (14M:2F) with symptomatic SCD (HbSS=10, HbSC=4, HbSßThal=4) underwent an AlloSCT. Indications for SCT included: ACS (n=8), CVA (n=2), multiple VOC (n=9), splenic sequestration (n=6) and retinopathy (n=1). Conditioning was Bu (4mg/kg × 4d < 4 yrs and 12.8mg/kg × 4d > 4 yrs), Flu (30mg/m2 × 6d), and Alemtuzumab (2mg/m2 × 1d, 6mg/m2 × 2d, and 20mg/m2 × 2d). Median age was 6.29 yrs(1.3–19.2). Median follow-up was 26 months. Donor sources: 8-6/6 HLA-matched sibling bone marrow (BM), 2–6/6 sibling UCB, 1–6/6, 4–5/6 and 3–4/6 unrelated UCB. Donors were HbAA (n=13), HbAC (n=1), HbAS(n=3), or ßThal trait (n=1). Sibling BM recipients received a median total nucleated cell (TNC) dose/kg of 7.16 × 108 (range 2.27–11.31) and a median CD34 cell dose/kg of 5.38 × 106 (range 1.50–6.83). Recipients of related or unrelated UCB received a median TNC dose/kg of 4.35 × 107 (range 3.40–9.10) and a median CD34 cell dose/kg of 2.21 × 105 (range 0.60–7.94). All received tacrolimus and mycophenolate mofetil as GVHD prophylaxis (Bhatia/Cairo et al, BBMT, 2009) and phenytoin or keppra as seizure prophylaxis for 180 days post SCT. Patients receiving sibling donor AlloSCT had a median time to neutrophil engraftment of 16 days (range 13–41). Among evaluable unrelated UCBT recipients, median time to neutrophil engraftment was 34 days (range 27–47). Four patients (all unrelated UCBT) experienced primary graft failure at day +60 post-SCT secondary to CMV (n=2), adenovirus, and low Bu Css levels with early withdrawal of MMF. Patients with sibling donors had a median time to platelet engraftment of 26 days (range 17–75). Of evaluable unrelated UCB recipients, median time to platelet engraftment was 54 days (range 43–70). Patients achieved mean whole blood donor chimerism of 71.0, 79.6, 85.7, 92.9, 90.7% and 93.2% at days 30, 60, 100, 180, 365 and 730 post-transplant, respectively. Mean erythroid (CD71) donor chimerism at these time points was 78.1, 81.1, 86.6, 90.5, 87.9% and 94.0%, respectively. Patients with non-sickle cell trait donors had median HbS levels of 0% at 1 yr (n=8) and 2 yrs (n=3) post-SCT; those with sickle cell trait donors had median HbS levels of 38.3% at 1 yr (n=3) and 39% at 2yrs (n=3) post-SCT. Among evaluable patients, the Kaplan-Meier probability of grade II-IV acute GVHD is 33.3% and of chronic GVHD is 8.3%. Among the ten donor-recipient pairs in which at least one member of the pair had CMV positivity, four experienced CMV reactivation on days +13, +26, +30 and +32 (BM: n=2, unrelated UCB: n=2). Of the four patients with primary graft failure (all unrelated UCBT recipients), one resumed chronic transfusions, two died of complications from disseminated CMV and adenoviral infections, and one received a second myeloablative SCT 1 yr after initial SCT and died of fungal sepsis. Among all patients, the Kaplan-Meier probability of OS is 81.8% (100% with sibling donors, 62.5% with UCB donors) and of EFS is 77.8% (100% with sibling donors, 50% with UCB donors) with none of these patients showing any evidence of SCD symptomatology post-SCT. The longest follow-up is 2134 days. In summary, we report the largest experience of RTC and matched family AlloSCT in selected children with symptomatic SCD with persistent long-term donor chimerism and absence of SCD symptoms or progression of disease. However, with the high incidence of viral infections and toxic deaths following unrelated UCBT, it is too premature to extend this treatment option to those SCD patients lacking a sibling donor. Disclosures:No relevant conflicts of interest to declare.

Cryopreserving and deglycerolizing sickle cell trait red blood cell components using an automated cell-processing system

Abstract RBC components with rare phenotypes are sometimes required for patients with sickle cell disease, and these rare components can often be found among donors with sickle cell trait. Cryopreserving RBC components from sickle cell trait donors requires a modified deglycerolization method to preserve the integrity of the RBCs. This study evaluated the feasibility of using an automated cell-processing system to cryopreserve and deglycerolize sickle cell trait donor RBC components. CP2D/AS-3 RBC components were collected from three donors with sickle cell trait. Each component was processed with an automated cell-processing system (ACP 215, Haemonetics Corp., Braintree, MA) and cryopreserved within 6 days of collection. The components were stored at –65°C or less for at least 2 days and were deglycerolized using the automated cell-processing system’s standard procedure. Before cryopreservation and after deglycerolization, several variables were measured. Deglycerolization resulted in recovery of 43.0, 76.5, and 67.5 percent of RBCs from the three sickle-cell-trait donor components compared with 80 percent or greater for all six control components. A small, dark red, jelly-like mass was noted in the bowl of the disposable set after deglycerolization of each of the three RBC sickle cell trait components. The osmolalities of all three sickle cell trait components were less than 400 mOsm/kg, but only one of the three was acceptable for a 14-day outdate. Freezing and deglycerolization of sickle cell trait donor RBC components with the automated cell-processing system resulted in recovery of some RBCs, but a decrease in RBC recovery was problematic. Modifications of the procedure are needed for processing sickle cell trait donor RBC components. Immunohematology 2008:24:107–111.

Leukoreduction of sickle cell trait blood: an unresolved issue

With the increasing demand for leukoreduction of blood components and the implementation of universal leukoreduction in several countries, the problems associated with leukocyte filtration of sickle cell trait blood have been revisited. Currently, there is no unified standard practice for sickle cell trait donors. Different blood centers adopt different policies. While some defer these donors from red cell component donation, some do not. Some screen all ethnic African donors for hemoglobin S (Hb S), others do not. Furthermore, there are differences in views of whether sickle cell trait red cells should be considered as equivalent to non-sickle cell trait red cells. Some blood centers do not give red cells from a sickle cell trait donor to the newborn or patients undergoing general anesthesia as a preventative measure. In this presentation, we discuss the epidemiology of the sickle gene, the sickling process, problems associated with leukoreduction of sickle cell trait whole blood and red cells, and some unresolved issues concerning donor referral and the usage of sickle cell trait blood.

BetaS Chain turnover in reticulocytes of sickle trait individuals with high or low concentrations of haemoglobin S.

Reticulocytes, isolated from the blood of sickle cell trait donors with either low (25-30%) or high (40-42%) haemoglobin S(Hb S) concentrations, were incubated with [3H]leucine for various times from 1.25 to 60 min. Samples of the total soluble fractions of the cells were denatured with urea and mercaptoethanol. The mixtures were analysed by electrophoresis on cellulose acetate strips. The specific radioactivities (dpm/mg) of the separated betaS and betaA globin chains were determined. The betaS/betaA ratios of globin chain specific radio activities in the reticulocytes of the 'low Hb S' donors decreased gradually from initial values higher than 1.30 to values near unity. These data suggested that faster turnover of some of the soluble, newly synthesized betaS chains compared to the newly synthesized betaA chains could explain part, but not all, of the disparity in concentrations of Hbs S and A in these people. When reticulocytes from 'high Hb S' donors were 3H-labelled for times longer than 5 min, the betaS/betaA specific radioactivity ratios remained at or near unity. This result suggested that newly synthesized betaS chains were not turning over selectively in these cells. Instead, there was a relative decrease in betaS chain synthesis proportional to the difference in blood concentrations of Hb S and Hb A. Additional calculations suggested that the more rapid turnover of newly synthesized betaS chains in the 'low Hb S' reticulocytes could explain the difference in Hb S concentrations between 'high and low Hb S' people. These results are consistent with previous reports that an alpha-thalassaemia gene, present in 'low Hb S' but absent in 'high Hb S' donors, may be responsible for the selective turnover of betaS chains.

Patterns of hemoglobin assembly in reticulocytes of sickle cell trait individuals.

Venous blood was obtained from five sickle cell trait donors with relatively high hemoglobin S concentrations (40% of total hemoglobin) and five donors with unusually low hemoglobin S concentrations (25 to 30%). A fraction of cells with 15 to 20% reticulocytes was isolated from the blood and incubated with [3H]leucine in a medium supporting protein synthesis for various times from 1.25 to 60 min. Previous studies showed an imbalance in globin chain synthesis in reticulocytes of "low hemoglobin S" donors which suggested the presence of an alpha-thalassemia gene; reticulocytes of "high hemoglobin S" donors had balanced globin chain synthesis (DeSimone, J., Kleve, L., Longley, M.A., and Shaeffer, J. (1974) Biochem. Biophys. Res. Commun. 59, 564-569). In the present study the soluble phase of the 3H-labeled reticulocytes was examined by electrophoresis on strips of cellulose acetate. The tetramer hemoglobins A and S were separated from each other and from a small pool of free, newly synthesized alpha and beta chains. Kinetics of labeling studies showed that the free alpha and beta chains were intermediates in tetramer hemoglobin assembly. The distribution of radioactivity between the alpha and beta chains of each of the electrophoretically isolated components were determined by separation of their globin chains on CM-cellulose columns. After 5 min of 3H-labeling of the reticulocytes from donors with 40% hemoglobin S the ratio of newly synthesized alpha chains to beta chains in the tetramer hemoglobins A and S ranged from 0.37 to 0.58. This ratio increased with longer labeling times. Almost all of the radioactivity of the free chain intermediates was in the alpha chain. These results confirmed the presence of a significant pool of newly synthesized alpha chains and a normal pattern of hemoglobin assembly in which initially unlabeled alpha chains combined with labeled beta chains when the cells were exposed to [3H]leucine. Conversely, in the reticulocytes of donors with 25 to 30% hemoglobin S the ratio of newly synthesized alpha chains to beta chains in the completed hemoglobins A and S ranged from 0.96 to 1.37 and remained unchanged throughout the 3H-labelling period. The radioactivity of the free alpha chain pool was substantially less that the total radioactivity of the betaA and betaS chain pools. These results confirmed the existence of a decreased pool size of soluble alpha chain intermediates and a pattern of hemoglobin assembly consistent with the presence of the alpha-thalassemia gene.

The influence of storage on erythrocyte survival in blood obtained from donors with sickle cell trait.

Abstract Blood collected from donors with sickle cell trait was stored under routine blood bank conditions. Erythrocyte survival studies, begun 1, 8, 15, and 17 days following collection, revealed no significant reduction over what might be expected in stored blood from donors with the hemoglobin genotype AA. Following 20 to 21 days of storage of AS erythrocytes, there may be 2 populations of cells present; a smaller relatively fragile population and a considerably larger normally resistant population. Blood obtained from sickle cell trait donors provides no additional risk of hemolysis to the recipient and may, therefore, be treated in a routine fashion in the blood bank.