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Biomedical subjects

M T Aye

Publications and source records attributed to M T Aye.

At least 19 recordsLinked to original sources
PubMed1998

Prevention of cytokine accumulation in platelets obtained with the COBE spectra apheresis system.

BACKGROUND AND OBJECTIVES: Febrile nonhemolytic transfusion reactions frequently accompany platelet transfusions and may be due to accumulation of cytokines mediating inflammation during storage of platelet concentrates (PCs). We wished to determine whether PCs collected using the COBE(R) SpectraTM Apheresis System (Version 4) were sufficiently leukocyte reduced (LR) to limit cytokine accumulation during storage. MATERIALS AND METHODS: Cytokine accumulation - interleukin (IL)-1beta, IL-6, IL-8, tumor necrosis factor-alpha (TNF-alpha) - and release of platelet alpha-granule - P-selectin, transforming growth factor-beta1 (TGF-beta1), platelet-derived growth factor AB (PDGF-AB), von Willebrand factor (vWf) - or dense granule (serotonin) markers were investigated during a 7-day storage period comparing apheresis-collected, LR PCs (LR PCs) and random donor platelets prepared from whole blood (WB). RESULTS: Leukocyte counts were reduced 99.95% comparing LR PCs (5.7 x 10(5)/l) and WB PCs (1.09 x 10(9)/l). Little or no accumulation of leukocyte-derived cytokines was observed in LR PCs during storage in contrast to WB PCs. A reduction in the release of platelet alpha-granule proteins, such as P-selectin, TGF-beta1 and PDGF-AB, was observed on day 0 for LR PCs compared to WB PCs with little or no difference observed from day 3 to 7. Plasma vWf levels were higher in LR PCs compared to WB PCs on days 0-7. CONCLUSION: Leukocyte levels in PCs collected with the COBE Spectra Apheresis System are sufficiently low to limit cytokine production during 7 days of storage.

Blood Preservation↗
PubMed1996

Flow cytometric immunofluorescence assay for detection of antibodies to human immunodeficiency virus type 1 using insoluble precursor forms of recombinant polyproteins as carriers and antigens.

A new serological assay, the recombinant flow cytometric immunofluorescence assay (r-FIFA), was developed for the early detection of human immunodeficiency virus type 1 (HIV-1) antibodies by using recombinant insoluble forms of HIV-1 Gag-p45, Gag-gp41 chimeric protein, gp160, Po197 polyprotein as antigens and autologous carriers through flow cytometry. These recombinant proteins were expressed in insect cells by a baculovirus expression system. Eight anti-HIV-1 seroconversion panels, a low-titer anti-HIV-1 panel from Boston Biomedica Inc. (BBI), and three HIV-1 seroconversion specimens from the Provincial Health Laboratory of Ontario, Toronto, Ontario, Canada (PHL), were tested and analyzed by r-FIFA. In sensitivity comparisons between r-FIFA and tests licensed by the U.S. Food and Drug Administration, which were used to test all of the HIV-1 panels from BBI, detection of HIV-1 antibody by r-FIFA was on average greater than 20 days earlier than that by enzyme immunoassay. The sensitivity of r-FIFA has permitted the detection of HIV-1-specific immunoglobulin G (IgG), IgM, and IgA antibodies during seroconversion. A kinetic analysis of HIV-1 antibody production of r-FIFA has shown that either IgG or IgM, or both, can be detected, depending on the phase and type of the immune response in the HIV-1-infected individual. Both primary and secondary immune responses were observed during this period. The r-FIFA results suggest that implementation of r-FIFA may significantly reduce the "window" period from the time of infection to the time of seroconversion, with earlier detection of antibodies after initial infection. This would also make it possible for us to understand the immune response and the precise mechanisms of immunopathogenesis in the early period of HIV-1 infection.

AIDS Serodiagnosis↗
PubMed1995

Effect of filtration of platelet concentrates on the accumulation of cytokines and platelet release factors during storage.

BACKGROUND: Platelet transfusions are frequently accompanied by febrile nonhemolytic transfusion reactions. These may be due, in part, to the release of cytokines interleukin 1 beta (IL-1 beta), interleukin 6 (IL-6), interleukin 8 (IL-8), and tumor necrosis factor alpha (TNF-alpha) by white cells (WBCs) into the plasma during storage of platelet concentrates (PCs). Acting as endogenous pyrogens, these agents may induce inflammatory responses. STUDY DESIGN AND METHODS: This study proposed to determine if WBC reduction in PCs by filtration significantly reduced the levels of cytokines normally generated during storage of unfiltered PCs up to 5 days. Serotonin, platelet-derived growth factor (PDGF-AB), and von Willebrand factor levels were also assessed to establish whether or not filtration or storage elicited significant platelet activation and granule release. RESULTS: Filtration significantly reduced total WBC counts by 99.1 percent before storage (p < 0.001) without affecting total platelet counts. Compared to unfiltered PCs, filtration prevented a rise in the levels of each cytokine by Day 3 for IL-1 beta (27.7 vs. 0.6 pg/mL; p < 0.05), IL-6 (114.2 vs. 0.4 pg/mL; p < 0.001), and IL-8 (4.2 vs. 0.02 ng/mL; p < 0.001). By Day 5, further increases in the levels of all cytokines were noted in unfiltered PCs, but Day 0 levels remained in filtered PCs (IL-1 beta: 105.4 vs. 0.4 pg/mL, p < 0.001; TNF-alpha: 42.2 vs. 7.5 pg/mL, p < 0.025; IL-6: 268.8 vs. 0.4 pg/mL, p < 0.001; and IL-8: 7.6 vs. 0.02 ng/mL, p < 0.001). From Day 0 to Day 5, there were significant increases in serotonin (21.3 vs. 6.3 ng/mL, p < 0.05), PDGF-AB (72.6 vs. 25.8 ng/mL, p < 0.001), and von Willebrand factor (4.7 vs. 2.7 IU/mL, p < 0.05) in unfiltered PCs, with similar increased levels being observed in filtered PCs during storage. CONCLUSION: These data indicate that the accumulation of high levels of cytokines in stored PCs could be prevented by WBC-reduction filtration of PCs without the induction of significant platelet activation or granule release. As cytokines have the potential to induce febrile nonhemolytic transfusion reactions in patients, the transfusion of WBC-reduced PCs would be expected to reduce the frequency and severity of such reactions.

Blood Platelets↗
PubMed1995

DNA typing for bone marrow engraftment follow-up after allogeneic transplant: a comparative study of current technologies.

DNA typing is widely used to document engraftment after allogeneic bone marrow transplantation (BMT). Most DNA typing procedures discriminate allogeneic engraftment on the basis of DNA length polymorphisms or sequence variations found in variable number of tandem repeat (VNTR) loci, or the presence of Y chromosome-specific DNA We have compared 3 types of VNTR analysis, their respective mode of allele detection and Y chromosome DNA detection in order to assess the strengths and limitations of each approach. Chimerism was assessed in 8 recipients after allogeneic BMT. Samples were subjected to 6 restriction fragment length polymorphism (RFLP) loci-analysis using radioactivity, 2 amplified fragment length polymorphism (AmpFLP) loci-analysis using a silver-stain mode of detection, 12 short tandem repeat (STR) loci-analysis using fluorescence detection and Y chromosome analysis. We evaluated each procedure for its ability to (1) discriminate sibling donor-recipient pairs in our samples; (2) generate a concordant chimerism diagnosis; and (3) detect and assess the contribution of minority components in mixed-chimera situations. In sex-mismatched BMTs with a female graft donor, Y chromosome probing has proven most efficient. In all other cases, AmpFLPs proved to be a rapid and efficient procedure with sufficient discriminating capability and sensitivity to warrant their use in clinical settings. STRs are rapid as well but require a larger loci complement to discriminate efficiently and they do not currently detect, under our conditions, all mixed chimeras. RFLPs are clearly superior at discriminating siblings but are time-consuming and serve best in cases where AmpFLP and STR analyses fail.

Alleles↗
PubMed1994

Spontaneous remission of polycythemia vera: clinical and cell culture characteristics.

A 20-year-old woman presented with polycythemia vera and was treated with phlebotomy alone for eleven years, following which all clinical manifestations of the disease disappeared. The clinical remission with normal physical findings and normal peripheral blood counts has persisted for a further 11 years. Erythroid colony culture results have paralleled the clinical state. Initial bone marrow cultures revealed spontaneous growth of erythroid burst-forming units (BFU-E). Subsequent cultures of peripheral blood cells throughout most of the period of spontaneous clinical remission have revealed little or no spontaneous growth of BFU-E. This suggests a suppression of the abnormal stem cell clone during the period of remission.

Adult↗
PubMed1994

Adenosine nucleotides and serotonin stimulate von Willebrand factor release from cultured human endothelial cells.

Endothelial cells (ECs) synthesize and release von Willebrand factor (vWf) either constitutively or from Weibel-Palade bodies by a regulated pathway. Although stimulated release of vWf from ECs occurs following exposure to thrombin, histamine, interleukin, tissue necrosis factor and fibrin in vitro, these agents are unlikely to be present in physiologically relevant concentrations during the initial stages of primary hemostasis. Alternatively, agents known to be released from the dense granules of activated platelets at the sites of vascular injury may provide the initial physiological stimuli for vWf release from ECs in vivo. We have examined the effects of the platelet secretagogues ADP, AMP, ATP and serotonin on the release of vWf from ECs and demonstrated enhanced release in all cases. The extent and time at which optimum vWf release was observed depended on the agonist and its concentration. At 3 nM, optimum release occurred after 4 hours with ADP (330%/ml) or 1 h with AMP (153%/ml) or ATP (450%/ml). At 30 nM, optimum release was seen after 1 hour with ADP (315%/ml) or AMP (595%/ml) and after 15 min with ATP (938%/ml). With serotonin, optimal release was seen by 30 min at 0.3 microM (1034%/ml) and after 1 h at 1 microM (745%/ml) although the response after 15 min was nearly equivalent (667%/ml). The doses giving 50% of maximal response (ED50) after 1 h were 6.5 nM (ADP), 15.2 nM (AMP) and 2.4 nM (ATP) and 20 nM for ATP or 75 nM for serotonin after 15 or 30 min respectively. ADP also enhanced PGI2 release from ECs in a dose- and time-dependent manner.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenine Nucleotides↗
PubMed1993

Platelet-activating factor secreted by DDAVP-treated monocytes mediates von Willebrand factor release from endothelial cells.

We have previously shown that although DDAVP (1-deamino-8-D-arginine vasopressin), a synthetic analogue of the natural hormone arginine vasopressin, does not directly promote release of vWf from human umbilical vein endothelial cells (ECs), enhanced release does occur when ECs were exposed to either monocytes or to supernatants recovered from DDAVP-treated monocytes. In the present study, we have found that exposure of monocytes to DDAVP did not increase secretion of interleukins (IL)-1 beta, IL-6, IL-8, tumor necrosis factor (TNF-alpha), growth factors G-CSF (granulocyte-), GM-CSF (granulocyte, monocyte-colony stimulating factor), prostaglandins (PG) E2, PGF2 alpha, or PGI2 or purine nucleotides such as ATP and ADP. However, increased levels of platelet-activating factor (PAF) were secreted by DDAVP-treated monocytes in a time- and dose-dependent manner that positively correlated with the enhancement in vWf release from ECs. Moreover, this effect could also be elicited when lipid extracts of these supernatants or purified PAF were added directly to ECs. This response could be inhibited with (+/-)-trans-2,5-Bis(3,4,5-trimethoxyphenyl)-1,3-dioxolane, a specific PAF receptor antagonist, when the ECs were exposed to supernatants from DDAVP-treated monocytes or to pure PAF. The present data indicate that enhanced secretion of PAF from monocytes is one mechanism whereby DDAVP can provoke release of vWf from ECs.

Adenosine Diphosphate↗
PubMed1992

Anti-hepatitis C virus (HCV) screening at a Canadian Red Cross center: significance of a positive c100 HCV enzyme-linked immunosorbent assay.

The c100 hepatitis C virus (HCV) enzyme-linked immunosorbent assay (ELISA) has been used to screen blood donors to prevent transfusion-associated non-A,non-B hepatitis. This test is not specific, and only about 25 percent of c100 HCV ELISA-positive blood samples appear to transmit hepatitis C. However, the intensity of the ELISA (sample/cutoff ratio [S/C], greater than 2) could identify a subpopulation of donors that are at high risk for transmitting hepatitis. Blood samples from 20,186 volunteer blood donors at a Canadian Red Cross blood transfusion center were screened for antibodies to HCV using the c100 HCV ELISA. Fifty-nine (0.3%) of these donors were repeatably reactive on ELISA. When their samples were tested with the c100 recombinant immunoblot assay (RIBA) and second-generation RIBA (RIBA-2), 26 (44%) and 31 (52%) samples, respectively, were found to be positive. Thirty-three of the 59 ELISA-reactive donors had an S/C greater than 2. Of these 33 donors, 30 (91%) had elevated alanine aminotransferase (ALT), 27 (82%) were RIBA-2 positive, and 22 (67%) had risk factors for hepatitis. In contrast, of the 26 ELISA-reactive donors with S/C less than 2, only 7 (27%) had elevated ALT, and 4 (15%) were RIBA-2 positive and also had high risk factors for hepatitis. Thus, while the HCV ELISA may lack specificity, its intensity can serve to identify a subgroup of donors that are at high risk for transmitting hepatitis.

Adult↗
PubMed1992

Expression of stem cell factor and c-kit mRNA in cultured endothelial cells, monocytes and cloned human bone marrow stromal cells (CFU-RF).

Previously, we have shown that conditioned medium from a subpopulation of human marrow stromal cells (CFU-RF) contain an activity able to stimulate the growth of macroscopic epo-dependent erythroid colonies. The ligand for the product of the c-kit proto-oncogene (also known as stem cell factor or SCF), among other activities, has been reported to have similar effects on erythroid colony growth. We have also presented data showing that SCF together with phytohemagglutinin-stimulated leukocyte conditioned medium can stimulate erythroid colony growth in the presence of antibodies to erythropoietin. Using the human SCF cDNA probe (K. Zsebo, Amgen Inc.) we now show that cells derived from CFU-RF colonies express SCF but not c-kit. Human umbilical vein endothelial cells were also found to express SCF and this expression was increased by addition of monocyte supernatant, IL-1 beta or thrombin. Cells of the human erythroleukemia cell line HEL were found to express c-kit but not SCF. Neither c-kit nor SCF mRNA were detected in phytohemagglutinin-stimulated lymphocytes. Together, these data support the view that the behaviour of proliferating erythroid stem cells in the marrow, which may express c-kit, could be regulated by membrane-bound SCF present on surrounding stromal cells.

Base Sequence↗
PubMed1992

Rearrangement of BCR genes in malignant lymphoma.

The Philadelphia chromosome, originally thought to be associated solely with chronic myelogenous leukemia (CML), has since been identified in acute leukemias and in some cases of lymphoma. The Philadelphia chromosome results from reciprocal translocation of genetic material between chromosome 9 and 22 involving the c-abl and BCR genes respectively. Southern blot analysis of the BCR genes was carried out on biopsy specimens from 49 patients presenting with malignant lymphoma without a previously documented CML phase. In two patients, BCR gene rearrangements were detected in the malignant lymph nodes but not in the bone marrow samples. A third patient showed BCR gene rearrangements in the bone marrow but not in the lymph node. From this limited study, it seems that the overall incidence of BCR gene rearrangement in malignant lymphoma is similar to that observed in adult AML.

Blotting, Southern↗
PubMed1991

Identification of an endothelial cell surface protein that binds plasminogen.

To identify and characterize endothelial cell surface components that bind plasminogen, we used ligand-blotting to study binding of plasminogen to sodium dodecyl sulphate solubilized extracts of human umbilical vein endothelial cells. It was observed that glu-plasminogen bound predominantly to a 45 kDa endothelial cell polypeptide. The interaction of labelled glu-plasminogen with this polypeptide was reversible and specific as the binding could be inhibited by both excess cold lysine and unlabelled glu-plasminogen but not by unrelated proteins. Binding of glu-plasminogen to cell extracts prepared from endothelial cells that had been pretreated with proteinase K was significantly reduced indicating that the 45 kDa polypeptide is a cell-surface protein. The cell-surface localization of the 45 kDa polypeptide was also indicated by the positive interaction of glu-plasminogen with membrane fractions of endothelial cells. Lys-plasminogen also interacted with the 45 kDa polypeptide in a specific manner and reversibility experiments indicated that lys-plasminogen could also displace the bound glu-plasminogen. Since binding of plasminogen to the 45 kDa endothelial cell surface polypeptide was very similar to plasminogen binding to intact endothelial cells, we propose that the 45 kDa protein represents one of the major receptors for plasminogen on human endothelial cells.

Cells, Cultured↗
PubMed1991

Characterization of novel platelet and endothelial cell target antigens in a family with genetic susceptibility to autoimmunity.

This report describes a French Canadian family whose members exhibit a high incidence of allo- and autoantibodies to antigens present on both platelets and endothelial cells. This is correlated with various HLA specificities known to be associated with autoimmunity, such as A1, B8, DR3, and, in some cases, with clinical disorders, including nephritis, hypertension, and thrombocytopenia. Immunoblot analysis using platelet and endothelial cell lysates showed serum antibodies to a 75 kDa endothelial cell surface polypeptide and to polypeptides with apparent mass of 115 kDa and 26 kDa found on both platelets and endothelial cells. This 115 kDa internal platelet protein was also found in a variety of other cell types, such as mononuclear cells, and increased following cell activation. Monoclonal antibody immunobilization assays were used to characterize the 26 kDa polypeptide; in three of the four patients tested, an antibody to leukocyte differentiation antigen CD9 was identified. The asymptomatic child of the propositus also exhibited an autoantibody against an 80 kDa platelet protein which was sensitive to thrombin digestion, suggesting that this polypeptide may be platelet glycoprotein V. In addition, P1A1 alloantibody was identified in one sister who had given birth to a severely thrombocytopenic boy and who herself had a severe vascular rejection to a cadaver kidney 2 years prior to this study. The propositus also developed hypertensive renal disease following a pregnancy and became dialysis dependent. Thus, members of this family have developed a variety of antibodies, particularly to platelet and endothelial cell antigens. Some subjects have remained asymptomatic in spite of having autoantibodies. However, others have been seriously ill, and their immune response to these antigens is believed to have played a role in the pathogenesis of their neonatal alloimmune thrombocytopenic purpura, hypertensive renal disease, renal graft rejection, and thrombocytopenia.

Antigens, CD↗
PubMed1991

Erythroid lineage-specific activity in conditioned medium derived from cloned human marrow stromal cells (CFU-RF).

Using long-term culture techniques, it has been shown that stromal cells in the marrow microenvironment are essential for the continued production and self-renewal of hematopoietic stem cells. We previously reported the development of a methylcellulose colony assay for a population of marrow stromal progenitors called CFU-RF. In this paper, a method is described for subculturing cells from individual CFU-RF-derived colonies to allow conditioned medium production (StCM). StCM, prepared in this way, was found to possess an erythroid lineage-specific activity that stimulated the formation of macroscopic erythroid colonies in cultures containing erythropoietin (epo). Using dose-response curves, the KG1 colony assay, and antibody neutralization, it was shown that the activity could not be attributed to interleukin 3 (IL3) or granulocyte-macrophage colony-stimulating factor (GM-CSF). However, it was further shown that a monolayer of stromal cells, which had earlier been producing the erythroid activity, could be stimulated by IL1 to produce granulocytic colony-stimulating activity, but only as long as IL1 was present in the culture medium. These findings indicate a mechanism whereby the same stromal population could be modulated to promote growth and differentiation of different hematopoietic lineages.

Bone Marrow↗