Changes in the Phosphate Compounds of the Human Red Blood Cell During Blood Bank Storage*

Nakao and his associates have demonstrated that when blood, which had previously been stored at 40 C for 8 to 10 weeks in acid-citratedextrose (ACD), was subsequently incubated at 370 C with both adenine and inosine, regeneration of ATP and restoration of post-transfusion viability took place (1-3). ATP regeneration did not occur when the incubation medium contained either adenine or inosine alone. Moreover, supplementation of the ACD preservative with both adenine and inosine at the beginning of storage resulted in better maintenance of ATP and posttransfusion viability than in samples stored with inosine alone or not supplemented at all (3-5).Although these studies indicated that both inosine and adenine were necessary for rejuvenation of long-stored erythrocytes, it seemed possible that adenine could act without inosine in preventing loss of red cell viability when it was present from the beginning of storage, since at that time, unlike later in storage, glucose utilization for energy production is still intact. Moreover, eliminating inosine would have the further advantage of avoiding toxicity from uric acid overload in the recipient. Accordingly, we investigated the

Section: Discussionmentioning

confidence: 65%

The Mechanism of Action of Adenine in Red Cell Preservation*

Sugita¹,

Simon²

1965

“…Figure 5 summarizes the results of the 15 survival studies carried out on 9 U stored blood (mean length of storage, 30.5 days). When whole blood (control) was administered, 71 Bromelin-treated blood after transfusion. Fresh Cr51-labeled red cells were exposed to bromelin for 15 minutes, then washed four times in 100 vol saline.…”

Section: In Vivo Studies Of Erythrocyte Changesmentioning

confidence: 99%

Agglutination of Stored Erythrocytes by a Human Serum. Characterization of the Serum Factor and Erythrocyte Changes*

Ozer¹,

Chaplin²

1963

There have been four previous reports of human sera causing agglutination of stored but not of fresh red cells (1-4). The sera are of special interest for two reasons. Since the agglutinins are active against the patients' own stored cells as well as against cells from other donors, one must consider whether such patients represent examples of autoimmune disease. Of more immediate practical importance, the sera provide a new means for studying the red-cell storage defect, since the agglutinins detect in the erythrocyte membrane a hitherto unrecognized alteration that progresses during in vitro storage concurrently with deterioration in cellular metabolism.Characterization of the previously reported stored-cell agglutinins has been limited. The present communication describes detailed studies of a serum causing strong agglutination of stored human red cells. The serum factor responsible for the agglutination was shown to be a gamma macroglobulin. Studies of red cells during storage in vitro indicate that development of agglutinability is closely related to the alterations in erythrocyte glucose metabolism occurring during storage. Also included are the results of sixteen studies of erythrocyte survival in vivo that are designed to examine the relationship of agglutinability to nonviability of the stored cells. METHODSThe patient was a 71-year-old woman whose chief complaints were abdominal swelling and progressive weakness. The principal positive physical findings were pallor and hepatosplenomegaly. Laboratory findings of special interest were: hemoglobin, 9.7 g per 100 ml; reticulocytes, 4.2%o; serum bilirubin concentration, nor-* Work supported by U. S. Public Health Service research grant G-2918 (C5) from the National Cancer Institute, Bethesda, Md. mal; total serum protein concentration, 7 g per 100 ml (albumin, 2.7 g, globulin, 4.3 g). Survival in vivo of Cr51-labeled, f resh, compatible, red cells of normal donors was moderately severely reduced, judged by redcell life-span (Cr51 tt, about 8 days) based on daily blood sampling for 1 week. Exploratory laparotomy revealed moderate hepatic enlargement, diffuse mesenteric and retroperitoneal lymph-node enlargement, and massive splenomegaly. Splenectomy was performed, and the patient's postoperative course was uneventful. Two units of blood were transfused before and two additional units during surgery. Although the patient was improved at the time of discharge and required no further transfusions, she died at home 7 months later of undetermined cause; necroscopy was not performed. The spleen removed at operation weighed 2,050 g and showed hyperplasia of the white pulp, with impaired follicle formation; each Malpighian corpuscle was surrounded by a margin of hyperplastic lymphoid cells. Lymph nodes revealed intact architecture, but were filled out with lymphoid elements without follicle formation. A liver sample from biopsy was not remarkable, and aspirated bone marrow revealed only generalized hyperplasia. A specific pathological diagnosis was not made. Stndies...

“…As had been noted previously, diphosphoglycerate proved quite labile to the conditions of storage (19,24). Its concentration fell rapidly in ACD so that after 2 weeks only 18 per cent of the initial level of 8.75 Mmoles (all data are presented as umoles of phosphorus per ml of erythrocytes) remained, and none was present at 6 We have reported previously that fructose diphosphate disappeared immediately after the addition of rabbit or human blood to ACD and that it was absent throughout storage (19,24).…”

Section: Methodsmentioning

confidence: 53%

Phosphorylated Carbohydrate Intermediates of the Human Erythrocyte During Storage in Acid Citrate Dextrose. I. Effect of the Addition of Inosine at the Beginning of Storage*

Shafer¹,

Bartlett²

1961

Self Cite

The post-transfusion survival of stored human or rabbit erythrocytes can be influenced favorably by the addition of certain purine nucleosides to the preserving solution (1-5). It had been noted earlier that the water-soluble organic phosphates of the erythrocyte disappeared to a large extent during storage (6), but nucleosides have been found to retard this change and, if added late in storage followed by a short incubation at 370 C, they bring about appreciable resynthesis of the organic phosphate (7-10).Early studies on the nature and changes of the phosphate compounds of the erythrocyte during storage were performed for the most part by using hydrolysis rates and paper chromatography (1, 2, 7-9). Recently improved technics for the isolation and identification of the metabolic intermediates, which use columns of ion exchange resin and more specific chemical and enzyme analytical methods, have greatly extended our knowledge of the normal carbohydrate intermediates of the human erythrocyte and make possible a more detailed analysis of changes occurring during storage (11-18).We have previously reported on the phosphorylated carbohydrate intermediates of the rabbit erythrocyte during storage with and without inosine ( 19). The present paper describes a similar study in which the glycolytic intermediates and nucleotides of human erythrocytes were analyzed at biweekly intervals during 8 weeks of storage at 40 C in acid citrate dextrose (ACD) and in ACD plus inosine (ACDI).* Supported by the US Army Medical Research and Development Command, Office of the Surgeon General, and by the National Heart Institute. METHODSFrom each of 2 healthy male donors who had normal blood counts, 480 ml of blood was drawn by vacuum into standard blood storage bottles containing 120 ml of ACD (NIH formula B). Inosine which had been autoclaved in normal saline in a concentration of 40 mg per ml was added to one sample in the amount of 4 mg (15 ,gmoles) per ml of blood; 100 ml of the ACD-blood mixture was removed aseptically from each sample for assay immediately and after 2, 4, 6, and 8 weeks of storage at 40 C.The blood samples were centrifuged, plasma and buffy coat removed, and the erythrocytes washed once with normal saline. The erythrocytes were mixed with 2 vol of 10 per cent trichloroacetic acid (TCA), the mixture centrifuged, and the precipitate re-extracted with 5 per cent TCA. The above procedures were carried out in the cold. The TCA was removed with ether.The erythrocyte extracts were neutralized, diluted 4-fold, and run through 1 X 15 cm columns of Dowex 1 X 8-chloride (60 to 80 wet mesh). The columns were eluted at a rate of 2.5 ml per minute with solutions of HCl and NH4C1, as shown in Figures 1 and 2