Quantitation of CD62, soluble CD62, and lysosome-associated membrane proteins 1 and 2 for evaluation of the quality of stored platelet concentrates

BACKGROUND: Platelets become activated during storage, which results in secretion of granules, vesiculation of microparticles, secretion of protein, and a number of other biochemical and morphologic processes that decrease the utility of platelet concentrates stored for transfusion. STUDY DESIGN AND METHODS: To evaluate the quality of stored platelet concentrates, the cell surface expression of specific activation-dependent antigens (CD62 and lysosome-associated membrane proteins 1 and 2 [LAMP-1, LAMP-2]) on platelets stored in a hospital blood bank over a 7-day period was examined. Relative microparticle counts and the expression of CD62 by microparticles, as well as platelet concentrate supernatant levels of soluble CD62, were determined. RESULTS: The percentage of platelets expressing CD62 increased significantly from Day 1 to Day 5 (p < 0.05) of storage; the mean fluorescence values for CD62 did not. In contrast, the mean fluorescence values of LAMP-1 and LAMP-2 rose significantly (p < 0.01 and p < 0.05, respectively) between Days 1 and 5. Significant declines in CD62, LAMP-1, and LAMP-2 percent expression and mean fluorescence were seen on Day 6 of storage (p < 0.001). Microparticle numbers increased significantly during storage and correlated with levels of CD62 protein (free and membrane-bound) (r = 0.95 vs. Day 2, p < 0.05; r = 0.88 vs. Day 5, p < 0.05). CONCLUSION: Flow cytometric evaluations of the expression of cell surface CD62, LAMP-1, and LAMP-2 are complementary tests that, especially when used in conjunction with the quantitation of CD62 protein, provided a simple and effective means of evaluating the quality of platelet concentrates stored for transfusion.     

Multivalent Pneumococcal Protein Vaccines Comprising Pneumolysoid with Epitopes/Fragments of CbpA and/or PspA Elicit Strong and Broad Protection

Immunization with the pneumococcal proteins pneumolysin (Ply), choline binding protein A (CbpA), or pneumococcal surface protein A (PspA) elicits protective responses against invasive pneumococcal disease in animal models. In this study, we used different mouse models to test the efficacy of a variety of multivalent protein-based vaccines that comprised various combinations of full-length or peptide regions of the immunogens Ply, CbpA, or PspA: Ply toxoid with the L460D substitution (referred to herein as L460D); L460D fused with protective peptide epitopes from CbpA (YPT-L460D-NEEK [YLN]); L460D fused with the CD2 peptide containing the proline-rich region (PRR) of PspA (CD2-L460D); a combination of L460D and H70 (L460D+H70), a slightly larger PspA-derived peptide containing the PRR and the SM1 region; H70+YLN; and other combinations. Each mouse was immunized either intraperitoneally (i.p.) or subcutaneously (s.c.) with three doses (at 2-week intervals) of the various antigen combinations in alum adjuvant and then challenged in mouse models featuring different infection routes with multiple Streptococcus pneumoniae strains. In the i.p. infection sepsis model, H70+YLN consistently provided significant protection against three different challenge strains (serotypes 1, 2, and 6A); the CD2+YLN and H70+L460D combinations also elicited significant protection. Protection against intravenous (i.v.) sepsis (type 3 and 6A challenge strains) was largely dependent on PspA-derived antigen components, and the most protection was elicited by H70 with or without L460D or YLN. In a type 4 intratracheal (i.t.) challenge model that results in progression to meningitis, antigen combinations that contained YLN elicited the strongest protection. Thus, the trivalent antigen combination of H70+YLN elicited the strongest and broadest protection in diverse pneumococcal challenge models.     

Evidence for functional endothelial cell damage in early diabetic retinopathy

Summary Two aspects of endothelial cell function were examined in two matched groups of male insulin-dependent diabetics, six with background retinopathy and seven without retinopathy. Leakage of fluorescein from the retinal capillaries was estimated by vitreous fluorophotometry. In addition, factors VIII/von Willebrand (vWF) and VIII-related antigen (VIII-RAG), which are synthesized by the endothelial cells, were measured, together with VIII-antihaemophilic factor (VIII-AHF). The patients without retinopathy had normal leakage of fluorescein in the macula (mean ± SEM: 1.10±0.10 g × 10^-8/ml) and the posterior vitreous (0.45±0.11 g × 10^-8/ml), and normal circulating levels of vWF (123% of a normal reference plasma ± 18%), VIII-RAG (137±14%) and VIII-AHF (112±18%). In contrast, the patients with background retinopathy showed higher leakage of fluorescein in the macula (6.34±1.74 g ×10^-8/ml; p<0.01), and the posterior vitreous (3.09±0.94 g ×10^-8/ml; p<0.02), as well as increased levels of vWF (177±16%; p<0.05). There was a trend towards increased VIII-RAG (195±24%; p<0.1), but not VIII-AHF (126 ±13%). Alterations of endothelial cell function thus accompany the development of retinopathy. It cannot be said from the present study whether these alterations also precede the appearance of retinopathy.