Treatment of whole blood (WB) and red blood cells (RBC) with S-303 inactivates pathogens and retains in vitro quality of stored RBC.

A pathogen inactivation (PI) process has been developed using the frangible anchor linker effector (FRALE) compound S-303. A series of experiments were performed in whole blood (WB) to measure the level of viral and bacterial inactivation. The results showed that 0.2mM S-303 and 2mM glutathione (GSH) inactivated >6.5 logs of HIV, >5.7 logs of Bluetongue virus, >7.0 logs of Yersinia enterocolitica, 4.2 logs of Serratia marcescens, and 7.5 logs of Staphylococcus epidermidis. Recent development for S-303 is focused on optimization of the PI process for red blood cell concentrates (RBC). A series of studies in RBC showed that 0.2mM S-303 and 20mM GSH inactivated approximately 5 logs or greater of Y. enterocolitica, E. coli, S. marcescens, S. aureus, HIV, bovine viral diarrhoea virus, bluetongue virus and human adenovirus 5. In both applications of the S-303 process, in vitro parameters of RBC function and physiology were retained compared to conventional RBC. Results from these studies indicate that S-303 can be applicable for PI of RBC and WB.

In vivo lability of glucose-6-phosphate dehydrogenase in GdA- and GdMediterranean deficiency.

A decreased level of glucose-6-phosphate dehydrogenase might result from decreased rate of synthesis, synthesis of an enzyme of lower catalytic efficiency, increased lability, or a combined mechanism. To test the hypothesis of increased lability, the rate of decline of the enzyme in vivo was measured in three groups of individuals, controls, Gd(-),A-males, and Gd(-), Mediterranean males, by the slope of decline of activity in fractions containing erythrocytes of progressively increasing mean age. These fractions were obtained by ultracentrifugation on a discontinuous density gradient of erythrocyte suspensions free of contaminating platelets and leukocytes. The rate of in vivo decline of pyruvate kinase (another age-dependent enzyme) was also measured and found very similar in the three groups. The in vivo decline of glucose-6-phosphate dehydrogenase was found to follow an exponential rate, with a half-life of 62 days for controls and 13 days for Gd(-),A- erythrocytes. The activity in normal reticulocytes was estimated at 9.7 U and in Gd(-),A- reticulocytes at 8.8 U. These estimates were confirmed by direct measurements in reticulocytes isolated from patients with extreme reticulocytosis. In Gd(-),Mediterranean erythrocytes activity could be demonstrated only in reticulocytes, which were estimated to average 1.4 U. The rate of decline is so extreme that no activity could be detected in mature erythrocytes. These data suggest that the glucose-6-phosphate dehydrogenase deficiency of both the Gd(A-) and the GdMediterranean variant results from different degrees of in vivo instability of the abnormal enzyme.

Biochemical heterogeneity in glutathione synthetase deficiency.

Two different clinical syndromes are associated with glutathione synthetase deficiency, one presenting with hemolytic anemia and 5-oxoprolinuria, the other with isolated hemolysis. We have differentiated these disorders on an enzymatic basis. In 5-oxoprolinuria, all cell types examined have grossly deficient enzyme activity and glutathione content. In contrast, in the nonoxoprolinuric variant, erythrocytes have decreased enzyme activity and glutathione content, whereas nucleated cells maintain substantial levels of both. The enzyme in this disorder is unstable in vitro and has shortened survival in intact erythrocytes. Nucleated cells appear able to maintain sufficient enzyme activity and concentrations of glutathione to suppress overproduction of 5-oxoproline.

The insulin receptor is an age-dependent integral component of the human erythrocyte membrane.

Human erythrocytes were separated according to age to determine whether there is heterogeneity in the specific binding of 125I-insulin to red cells. The mean cell age of erythrocyte isolates was determined from the cumulative distribution frequency of the cells in an isotonic (290 milliosmolar) density gradient and confirmed by assay for pyruvate kinase, an age-dependent red cell enzyme. An IBM 2997 centrifugal cell separator was used to obtain larger quantities of younger erythrocytes from normal subjects. 125I-insulin binding to red cells including reticulocytes was found to decrease exponentially as a function of their mean cell age in 8 normal subjects. A change in receptor number rather than affinity appeared to account for the observed change in 125I-insulin binding. An exponential, age-dependent change in binding of a hormone to its cell membrane bound receptor has not previously been observed. Consistent with these results is the possibility that regulation of the red cell insulin receptor concentration takes place only in the younger red cells.

Human platelet arylsulphatases: identification and capacity to destroy SRS-A.

Arylsulphatases IIA and IIB have been separately identified in human platelets by use of anion exchange chromatography and gel filtration. Arylsulphatase IIA had a molecular weight of 160,000 and a pH optimum of 4.5. Arylsulphatase IIB had a molecular weight of 60,000 and a pH optimum of 5.5. Both arylsulphatases IIA and IIB were inhibited by phosphate and sulphate ions characteristic of this enzyme class. Platelets, upon exposure to ionophore A-23187 or thrombin, discharged arylsulphatase coincident with beta-glcuronidase release. Partially purified platelet arylsulphatase IIB inactivated rat SRS-A.

Derivation of lymphoma cell lines from microscopically normal bone marrow in patients with undifferentiated lymphomas: evidence of occult bone marrow involvement.

Evidence is presented that at least 17% of microscopically normal bone marrow samples obtained from patients with undifferentiated lymphomas contain occult tumor cells. Of 19 microscopically normal bone marrow samples tested, continuous tumor cell lines were obtained from 4. A tumor cell origin was confirmed by the presence of an 8;14 chromosomal translocation in each case, and HLA typing confirmed the patient origin of the cell line. In two other patients, direct cytogenetic examination of microscopically normal bone marrow samples revealed karyotypes containing 8:14 translocations or a 14q+ chromosome. These findings indicate that undifferentiated lymphomas are often more widespread than is clinically appreciated. The presence of submicroscopic marrow involvement is also of significance to the design and analysis of treatment protocols involving autologous marrow infusion.