These results show that PS expression increases after preparation of PCs from platelet-rich plasma and rises progressively during platelet storage under blood bank conditions. Furthermore, the greater PS expression is associated with increased platelet- dependent thrombin-generating capacity.

No abstract

Acute hemolytic anemia associated with red blood cell (RBC) glucose-6-phosphate dehydrogenase (G6PD) deficiency is commonly encountered in the Mediterranean basin. Nevertheless, concomitant clinical evidence of white blood cell G6PD deficiency is extremely rare in Israel. This study sought to assess simultaneously levels of G6PD activity in polymorphonuclear leukocytes (PMN) and in red blood cells (RBC) of patients with G6PD deficiency, including full-term newborn infants. In PMN, the correlation between G6PD activity, hexose monophosphate shunt activity, and superoxide anion release was evaluated. In G6PD-deficient patients, a parallel and significantly decreased G6PD activity was found in neutrophils (range of activity 0 -4.5 IU/10 6 PMN) and erythrocytes (range of activity 0 -1.8 IU/g Hb), compared with healthy controls (5-23 IU/10 6 PMN and 2.4 -6.4 IU/g Hb, respectively). A positive correlation was found in PMN between the levels of G6PD activity, hexose monophosphate (HMP) shunt activity, and superoxide anion release (p Ͻ 0.01). Nevertheless, all patients' bactericidal activity of neutrophils remained in the range of healthy controls. Although many episodes of acute hemolytic anemia were recorded, no increased incidence of pyogenic infections was observed in any group of patients investigated. Neutrophil and erythrocyte G6PD levels were re-assessed in some of these patients several times a day. A significant diurnal fluctuation of the enzyme activity was found. It is speculated that the patients produce fluctuating daily quantities of NADPH, sufficient to initiate the neutrophil respiratory burst and to achieve normal bactericidal activity, necessary to prevent the development of microbial infections. (Pediatr Res 55: 807-813, 2004) Abbreviations G6PD, glucose-6-phosphate dehydrogenase CGD, chronic granulomatous disease HMP, hexose monophosphate RBC, red blood cell WBC, white blood cell PMN, polymorphonuclear Erythrocyte G6PD deficiency, mainly manifested by hemolytic anemia induced by oxidative stress to the RBC, is a relatively common worldwide disorder (1). The G6PD gene is located on the X chromosome, leading to the clinical manifestations of the disease being usually confined to hemizygous men, although female carriers with marked expression of the aberrant allele may also suffer (2). This entity is encountered among various ethnic groups in Israel as well (3-5).G6PD catalyzes the reduction of NADP to NADPH, the first step of the HMP pathway. Phagocytic leukocytes use NADPH as a substrate for the NADPH-oxidase enzyme, which contributes to the killing of ingested microorganisms by subjecting them to highly toxic reactive oxygen species (ROS) and bringing about myeloperoxidase-catalyzed halogenation of proteins. Recently, it was reported that the killing activity of neutrophils is enhanced in yet another way by the NADPH oxidase (6).

Recently, the asymmetric distribution of phospholipids in eukaryotic cell membranes has been appreciated and been found to be dependent on the activity of a number of enzymes. The expression of phosphatidylserine (PS), a negatively charged phospholipid, on the platelets of patients with polycythemia vera (P vera) and essential thrombocythemia (ET) was compared to that in normal individuals. The effect of platelet aggregation on PS expression was determined. Exposure of PS on platelets obtained from patients with P vera and ET and from age- and sex-matched healthy volunteers was measured by fluorescein-labeled Annexin V binding to platelets and by the platelets' thrombin-generating capacity determined by the prothrombinase assay. PLatelet prothrombinase activity (mean +/- standard deviation [SD]), as measured by thrombin generation, was 2.32+/-2.2 micro/mL in the P vera group and 1.55+/-1.0 micro/mL in the control group (p=0.3). PS expression as measured by Annexin V binding (mean +/- SD) was 2.6+/-2.4 % in the P vera group versus 1.55+/-1.2% among controls (p=0.03). In the ET group, prothrombinase activity (mean +/- SD) was 1.0+/-0.6 micro/mL and 2.1+/-0.9 micro/mL in the control group (p=0.006). Annexin V binding (mean +/- SD) was 4.8+/-4.2% in the ET group and 2.77+/-2.1% among control subjects (p=0.09). When the prothrombinase assay was performed after addition of adenosine diphosphate (ADP) to the platelets, there was a significant increase in thrombin generation in the myeloproliferative disorder (MPD) group (3.1+/-2.0 micro/mL) compared to the thrombin generated by unstimulated myeloproliferative disorder platelets (2.07+/-1.69 micro/mL) (p=0.0006). An increase in thrombin generation was seen in the ADP-stimulated platelet samples in all ten paired samples studied. Likewise, the addition of ADP to control platelets increased thrombin generation from 2.0+/-1.0 micro/mL in unstimulated platelets to 4.3+/-1.6 micro/mL in ADP-treated platelets (p=0.0006). Thrombin generation increased in all of the ADP-stimulated platelet samples compared to the untreated platelets. There was however, no difference in the increased thrombin generation when ADP-stimulated platelets from MPD patient and control subjects were compared (p=0.3). Results indicate that some patients with MPDs may show increased PS expression on platelet surface. When analyzed overall, there was a tendency toward greater PS expression in the P vera and ET patient groups; however, the increase did not reach statistical significance. This increase was noted in both the prothrombinase assay the Annexin V binding assay. We have also shown that stimulation of platelets by addition of the agonist ADP results in enhanced PS expression, which appears increase the thrombogenic potential of the platelets as demonstrated by the enhanced thrombin generation demonstrated by these platelets in the prothrombinase assay. There was no difference in the degree of PS expression in response to ADP stimulation between MPD and control platelets. Results show that PS express...