Blood preservation XVI packed red cell storage in CPD-adenine

Interest has been renewed in CPD-adenine as a long-term liquid blood preservative. The question of whether the metabolic product of adenine, 2,8-dioxyadenine was toxic to humans has apparently been resolved by extensive animal and human studies in favor of there being no potential toxicity in the amounts used in blood preservation. Sweden is adopting CPD-adenine (0.25 mM) as its national blood preservative after ten years of clinical experience in trials. They have shown that each additional week of storage time beyond the current three weeks with CPD results in a 50 per cent reduction of wasteage caused by outdating. They are adopting the 35-day time for regular use with 42 days for an emergency reserve supply. However, many units of blood in the U.S. are stored as packed red blood cells and the question has been raised as to whether there is sufficient glucose in the preservative to maintain red blood cell metabolism in the packed cell unit. The present investigation indicates that there is sufficient glucose for 35 days of packed cell storage in CPD-adenine (0.25 mM) but in some units this might be marginal at 42 days of storage.     

Blood preservation. XXVIII. Galactose and maltose maintain red blood cell 2,3-DPG and ATP

Because there may be inadequate dextrose in the newly licensed CPD- adenine for five or six weeks storage of high hematocrit red blood cells, this laboratory has examined some alternate sugars for their ability to maintain red blood cell metabolism during storage. In the current study, dextrose and fructose were studied as model or prototype nutrients. A third six carbon monosacharide, galactose, three dissacharides, lactose, maltose, and sucrose were studied in the same experiment. Of these, fructose best maintained ATP and 2,3-DPG during the fourth to sixth week of whole blood storage at 4 C. Dextrose was next best during this time and was nearly equivalent to fructose in the first three weeks of storage. Galactose and maltose both maintained ATP and 2,3-DPG, but not nearly so well as did fructose and dextrose. Sucrose and lactose were associated with the most rapid deterioration of ATP and DPG levels and they failed to maintain the progressive fall in pH which is usually associated with continuing, useful metabolism.     

Five-week red cell storage with preservation of 2,3 DPG

The 2,3 diphosphoglycerate (2,3 DPG) content of red cells stored in current anticoagulant-preservative products decreases rapidly after the first few days of storage, and by 3 weeks the red cells are essentially depleted of 2,3 DPG. Because ascorbic acid and ascorbate-2-phosphate (A-2-P) are effective in maintaining erythrocyte 2,3 DPG during liquid preservation, ascorbate was stabilized through autoclaving and subsequent storage by adding it as the trisodium salt of A-2-P to a phosphate-adenine-saline solution at a pH of 8.5 to 9.0. Red cell concentrates prepared from blood drawn into citrate-phosphate-double-dextrose were supplemented with the A-2-P additive solution (AS-4) and studied in vitro and in vivo. Mean 2,3 DPG values for 22 units were 147.6, 113.5, and 82.3 percent of initial value after storage for 3, 4, and 5 weeks, respectively. Maintenance of 2,3 DPG was at the expense of adenosine triphosphate (ATP), which fell to as low as 22.2 percent of initial value after 5 weeks. Despite the low ATP values, the 24 hour 51Cr-labeled red cell recoveries averaged 80.8 and 74.1 percent after 4 and 5 weeks of storage, respectively. The AS-4 system provides a red cell product with acceptable viability and improved oxygen off-loading function.     

The preservation of red cell antigens at low ionic strength

Low-ionic-strength saline (LISS) techniques permit a safe and substantial reduction in incubation time and have therefore become the method of choice for antibody detection and compatibility testing in many transfusion laboratories. Consequently, the supply of reagent red cells (RBCs) in a low-ionic-strength preservative solution would remove the daily need for laboratories to wash and resuspend cells in LISS before use. However, the storage of fresh RBCs at low ionic strength in the presence of aminoglycoside antibiotics can cause a rapid loss of certain antigens, possibly as a result of the release of proteolytic enzymes from contaminating white cells. This article describes a low-ionic-strength solution that achieves preservation of antigens on liquid nitrogen-frozen-thawed RBCs for 21 days' storage at 4 degrees C.     

Blood preservation. XLIII. Studies on the ascorbate mechanisms of maintaining red cell 2,3-DPG

Our previous experiments on the mechanisms of ascorbate's effect on the red blood cell failed to show an effect of iodoacetate (IA), a sulfydryl inhibitor. In this study, in contrast to the previous, iodoacetate (85 micromolar) was seen to prevent continued red blood cell metabolism. During the first weeks there was an absence of a continual fall in pH; ATP levels were depressed below half normal; and 2,3-DPG levels fell to very low values within the first week. ATP was best maintained in the control preservative and next best maintained, at adequate levels, with ascorbate, 5 mM, with and without glutathione, 5 mM. 2,3-DPG levels were well maintained with ascorbate and ascorbate with glutathione. Poor ATP maintenance and rapid decreases in 2,3-DPG were observed with iodoacetate, IA plus ascorbate, and IA plus ascorbate and glutathione.     

Blood preservation. XXIX. Pyruvate maintains normal red cell 2,3-DPG for six weeks of storage in CPD-adenine

Pyruvate was placed in experimental CPD-adenine (0.25 mM) blood preservative mixtures in four concentrations ranging from 40 to 320 mM. In the 320 mM pyruvate preservative, 2,3-DPG levels were elevated above normal for six weeks of whole blood storage at 4 C. The lower pyruvate concentrations maintained elevated or normal 2,3-DPG levels for less time: four weeks with 160 mM, two weeks with 80 mM, and one week or less with 40 mM or the control. ATP values were best maintained in the control. The higher pyruvate concentrations resulted in the most rapid decreases at ATP. However, even the 320 mM pyruvate did not cause ATP to fall below 2 microM/gm of Hb. The higher pyruvate concentrations produced and maintained a higher pH during storage. On the other hand, 2,3-DPG levels increased with pyruvate during the first week of storage when the pH was decreasing rapidly. This could be the result of its oxidation of NADH to NAD. The high pyruvate concentration which maintained elevated 2,3-DPG levels throughout the six weeks might be simulating the effect reported in pyruvate kinase-deficient red blood cells, in which blockage of glycolysis at that step is preventing 2,3- DPG catabolism.     

The effect of hypotonicity, glutamine, and glycine on red cell preservation

BACKGROUND : Red cells (RBCs) stored in hypo-os-molar additive solutions with the same concentrations of adenine, dextrose, mannitol, and sodium chloride and varied amounts of ammonium, phosphate, glycerol, and glutamine were better preserved than RBCs in the standard additive solution (Adsol). Cell swelling occurred in all the experimental additives. This observation prompted the evaluation of glutamine and glycine alone, as well as a combination of glutamine and glycine, all of which have been described as producing swelling of rat liver cells. STUDY DESIGN AND METHODS : Aliquots of RBCs were stored at 4°C in Adsol or experimental additive solutions (EASs) all containing adenine, 2 mM; dextrose, 110 mM; mannitol, 55 mM; and sodium chloride, 50 mM. EAS 42 had, in addition, glutamine, 10 mM; glycine 5 mM; and phosphate, 20 mM. EAS 43 had glutamine, 10 mM; glycine, 10 mM; and phosphate 20 mM. EAS 44 had glutamine, 10 mM; EAS 45 had glutamine, 10 mM, and phosphate, 20 mM; and EAS 46 had only glycine, 10 mM. At intervals, measurements were made of mean corpuscular volume, mean corpuscular hemoglobin concentration, morphology, ATP, hemolysis, supernatant potassium, ammonia, pH, and microvesicles shed. RESULTS : The initial mean corpuscular volumes were larger in all EASs than in Adsol, but the greatest difference was between EASs 44 and 46 (108 fL) and Adsol (86 fL) (p<0.001). The morphology scores were significantly better in all the EASs (p<0.04). The ATPs were significantly greater in all the EASs (p<0.001), and highest in those with phosphate. Potassium leakage and hemolysis were less in the EASs (p<0.001). The ammonia levels were higher in all the EASs than in Adsol, with the exception of EAS 46. During storage, the extracorpuscular and intracorpuscular pH levels were essentially identical. The shedding of microvesicles was greatly reduced in all the EASs. CONCLUSION : Cell swelling induced in RBCs after collection appears to improve preservation. Ammonia and phosphate enhance RBC ATP maintenance. Glycine decreases the formation of ammonia by RBCs stored in a hypotonic medium.     

器官保存液中聚乙二醇对人红细胞聚集性和血液流变学的影响

背景:聚乙二醇作为一种非渗透性大分子物质,在器官保存液中可发挥保护细胞膜、维护细胞骨架完整性、防止细胞水肿、抗脂质过氧化和免疫调节的作用.目的:探讨器官保存液中的聚乙二醇对人红细胞聚集性和血液流变学的影响.设计、时间及地点:对比观察实验,于2008 10在解放军第二军医大学附属长征医院器官移植科完成.材料;抽取接受体格检查的6名健康志愿者的肘前静脉血.方法:在抽取的新鲜血液中按5:1的稀释比分别加入生理盐水、器官保存液及含不同相对分子质量、不同浓度聚乙二醇的多器官保存液,按加入液体的不同分为:生理盐水组、器官保存液组、不添加聚乙二醇的保存液组、20聚乙二醇1,10,30 g/L和35聚乙二醇1,10,30 g/L组.主要观察指标:室温下通过魏氏法检测红细胞沉降率、自动血液流变仪检测血液流变学指标、光镜观察红细胞聚集的形态学改变,分析聚乙二醇对人红细胞聚集性和血液流变学的影响.结果:不含胶体的保存液对红细胞聚集无影响,含低浓度聚乙二醇的保存液对红细胞聚集性和血液流变学的影响较小,器官保存液组、20聚乙二醇30 g/L,35聚乙二醇10 g/L和35聚乙二醇30 g/L的保存液则可显著加快红细胞沉降率,降低红细胞变形能力,引起红细胞明显聚集.结论:器官保存液中的聚乙二醇可引起红细胞聚集,降低红细胞变形能力,其相对分子质量越大,浓度越高,促进红细胞聚集的作用越明显,对血液流变学的影响越大.     

Clinical trials in dermatology. The phase I trials]

Application of the new topical product on the diseased skin should be preceded by its safety evaluation on the healthy skin in human volunteers. We propose here guidelines for the evaluation of the irritation, sensitization, phototoxicity and photoallergy potentials for topical products. The methods for evaluation of percutaneous absorption are also discussed. The studies presented here are not the object of any regulations. Therefore, we propose here an approach for the safety evaluation of topical products in human volunteers.     

Conservative treatment of erectile dysfunction. 2: Clinical trials

This is the second of a three-part paper addressing whether physiotherapy involving pelvic floor muscle exercises (PFMEs) is efficacious as a first-line treatment for erectile dysfunction (ED). The first part (Vol 9(11): 691-4) highlighted the prevalence of ED, associated risk factors, the anatomy of the penis and the physiology of erection. This part concentrates on the computer-aided and manual search for published clinical trials investigating the treatment and prevention of ED. The methodological quality of the trials was assessed using criteria based on generally accepted principles of interventional research. The literature search revealed 14 trials which met the broad inclusion criteria. Of these, eight trials used PFMEs with or without biofeedback or electrical stimulation.     

Feline glucose-6-phosphate dehydrogenase cellular mosaicism. Application to the study of retrovirus-induced pure red cell aplasia.

Neoplasms result from the uncontrolled proliferation of abnormal or transformed cells. The early stages of this process are difficult to study because of the lack of sensitive and specific markers of clonal evolution in an experimental system. We have developed a cat model using cellular mosaicism for glucose-6-phosphate dehydrogenase (G-6-PD). Our findings confirm that the structural locus for feline G-6-PD is on the X-chromosome and demonstrate that it is randomly inactivated in somatic cells. Heterozygous cats have balanced ratios of G-6-PD enzyme types in peripheral blood cells and hematopoietic progenitors that remain stable over time. In our initial studies, we used the model to analyze the events surrounding marrow failure experimentally induced by selected strains of feline leukemia virus (FeLV). Two G-6-PD heterozygous cats, one F1 male hybrid and one domestic cat were infected with FeLV (C or KT) and developed pure red cell aplasia (PRCA). Colonies arising from the more mature erythroid colony-forming cell were not detected in marrow culture of anemic animals although erythroid bursts persisted, suggesting that the differentiation of early erythroid progenitors (BFU-E) was inhibited in vivo. The ratio of G-6-PD types in hematopoietic progenitors and peripheral blood cells from the heterozygous cats did not change when the animals developed PRCA. Thus, the anemia did not result from the clonal expansion of a transformed myeloid stem cell. With this experimental approach, one may prospectively assess clonal evolution and cellular interactions in other FeLV-induced diseases.