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Recent technologic advancement enables us to prepare leukocyte-depleted or UVB-irradiated platelet concentrates for possible prevention of primary HLA alloimmunization. However, it is yet not known which of these two approaches is more... more
Recent technologic advancement enables us to prepare leukocyte-depleted or UVB-irradiated platelet concentrates for possible prevention of primary HLA alloimmunization. However, it is yet not known which of these two approaches is more efficacious. Because well-controlled studies cannot be easily conducted in human subjects to answer this question, a series of experiments were performed using a mouse transfusion model. The results showed that 100% of CBA mice with H2k haplotype developed antibody to donor H2d major histocompatibility complex (MHC) antigens after two weekly transfusions of platelet concentrates containing 2000 leukocytes/microL. In contrast, only 50% of the mice became alloimmunized after receiving platelets containing < or = 2 leukocytes/microL. More impressively, none developed anti-H2d antibodies after receiving two platelet concentrates irradiated with 1,200 mJ/cm2 UVB. UVB irradiation was found to be equally effective in reducing the alloantigenicity of plate...
Research Interests: Immunology, Biology, Medicine, Platelet, Mice, and 13 moreHaplotypes, Animals, Blood, Clinical Sciences, Time Factors, Major histocompatibility complex, Blood Transfusion, Leukocytes, Antigen, Ultraviolet Rays, Blood platelets, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
Transfusions (Tx) of Ultraviolet B (UVB)-irradiated peripheral blood mononuclear leukocytes (MNL) have been shown to induce humoral immune tolerance to major histocompatability complex (MHC) antigens (Blood 88:4375, 1996). To determine... more
Transfusions (Tx) of Ultraviolet B (UVB)-irradiated peripheral blood mononuclear leukocytes (MNL) have been shown to induce humoral immune tolerance to major histocompatability complex (MHC) antigens (Blood 88:4375, 1996). To determine whether cellular immune tolerance to MHC antigens can be induced by the same approach, transplantation of bone marrow and spleen cells from tolerant donors across the H-2 barrier was conducted to study its effect on prevention of graft-versus-host disease (GVHD). After immune tolerance induction by four weekly Tx of UVB-irradiated BALB/c (H-2d) peripheral blood MNL into CBA/HT6 (H-2k) mice, bone marrow cells (BMC) and spleen MNL from tolerant or naive CBA mice were transplanted into lethally irradiated BALB/c mice. The transplanted mice were followed by measuring body weight, peripheral leukocyte counts, GVHD, survival, and cytokine response. All BALB/c recipient mice were fully engrafted with H-2k CBA donor cells after transplantation. The severity o...
Research Interests: Immunology, Biology, Medicine, Spleen, Mice, and 15 moreAnimals, Blood, Immunosuppression, Bone marrow, Clinical Sciences, Immune system, Bone Marrow Transplantation, Body Weight, Immune Tolerance, Leukocytes, Interleukin, Interferon gamma, Cardiovascular medicine and haematology, Paediatrics and reproductive medicine, and leukocyte Count
Recent studies on platelet HLA indicate that greater than 50% of platelet HLA antigens are adsorbed on the platelet surface and may be derived from plasma. It has been speculated that platelet HLA may be directly proportional to plasma... more
Recent studies on platelet HLA indicate that greater than 50% of platelet HLA antigens are adsorbed on the platelet surface and may be derived from plasma. It has been speculated that platelet HLA may be directly proportional to plasma HLA concentration. To determine the quantitative correlation between plasma and platelet HLA, a precise competitive enzyme-linked immunoassay (ELISA) for measurements of soluble and cellular HLA antigens was developed by using purified HLA antigens and W6/32 anti-HLA monoclonal antibody. The useful range of the standard curve for the assay was 0.01 to 5.0 micrograms/mL. The intraassay and interassay variations were 7% and 14%, respectively. The plasma HLA concentrations measured in 61 healthy adults ranged from 0.25 to 4.1 micrograms/mL, and the mean plasma HLA concentration was 1.47 +/- 0.87 micrograms/mL (+/- SD). Platelet HLA concentrations determined in the same 61 persons ranged from 4.7 to 17.33 fg/platelet, and the mean concentration was 9.3 +/...
Research Interests: Chemistry, Medicine, Platelet, Humans, Blood, and 13 morePlasma, Monoclonal Antibodies, Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis, Clinical Sciences, Immunoassay, Monoclonal Antibody, Reference Values, Enzyme Linked Immunosorbent Assay, Sodium Dodecyl Sulfate, Blood platelets, Human leukocyte antigen, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
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Research Interests: Immunology, Treatment Outcome, Medicine, Humans, Female, and 13 moreMale, Regression Analysis, Total Hip Replacement, Clinical Sciences, Aged, Middle Aged, Knee replacement, Blood Transfusion, University Hospitals, Logistic Models, Transfusion, Erythrocyte transfusion, and Cardiovascular medicine and haematology
To determine whether chloroquine can specifically elute HLA antigens and beta 2-microglobulin (beta 2-M) from the platelet surface, quantitative immunofluorescence flow cytometry and monoclonal antibodies were used to show that HLA... more
To determine whether chloroquine can specifically elute HLA antigens and beta 2-microglobulin (beta 2-M) from the platelet surface, quantitative immunofluorescence flow cytometry and monoclonal antibodies were used to show that HLA antigens and beta 2-M were proportionally eluted from the platelet surface without affecting the membrane glycoproteins IIb and IIIa. Second, an autoradiogram of electrophoresed I125-labeled platelets showed that only beta 2-M but not other I125-labeled membrane proteins could be eluted. Although HLA antigens were poorly labeled by I125 and could not be detected on the autoradiogram, the eluted HLA antigens could be detected by anti-HLA monoclonal antibody and immunoblotting techniques. No loss of plasma membrane integrity was observed by transmission electron microscopy after chloroquine treatment of platelets. The results indicate that chloroquine selectively elutes HLA antigens and their noncovalently associated beta 2-M without affecting other integral platelet membrane proteins.
Research Interests: Chemistry, Immunology, Flow Cytometry, Transmission Electron Microscopy, Medicine, and 15 morePlatelet, Humans, Beta decay, P-glycoprotein, Clinical Sciences, Fluorescent Antibody Technique, Monoclonal Antibody, Membrane Protein, Chloroquine, Transfusion, Antigen, Blood platelets, Elution, Human leukocyte antigen, and Cardiovascular medicine and haematology
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To understand the complexity of plasma HLA antigens, the distribution of different molecular weight forms of class I HLA in plasma was investigated in 44 HLA-phenotyped and unrelated individuals. Plasma class I HLA were immunoprecipitated... more
To understand the complexity of plasma HLA antigens, the distribution of different molecular weight forms of class I HLA in plasma was investigated in 44 HLA-phenotyped and unrelated individuals. Plasma class I HLA were immunoprecipitated by using the W6/32 anti-HLA monoclonal antibody, separated by SDS-polyacrylamide gel electrophoresis and characterized by immunoblotting with the HC-10 monoclonal antibody. Four different forms of HLA heavy chains (HLA-HC) with relative molecular masses of 44, 39, 36, and 34 kd were detected. Plasma samples from all individuals contained 44 and 36 kd HLA-HC, but varied as to the presence of 39 and 34 kd HLA-HC. Eighteen percent of the individuals did not have any detectable class I HLA with 39-kd heavy chains in their plasma and 61% did not have plasma class I HLA with 34-kd heavy chains. Thus, four different distribution patterns were identified for plasma class I HLA among all individuals included in our study. The distribution patterns in four different individuals were evaluated quarterly and remained unchanged during 1 year follow-up. A significant association of absence of 39-kd plasma class I HLA-HC with female gender (p less than 0.05) and HLA-B7 phenotype (p less than 0.00015) was also found. Further pedigree analyses of four families of HLA-B7-positive and 39-kd HLA-HC-negative probands indicated that genetic factor(s) other than those associated with HLA-B7 allele and female gender is involved in regulating the expression of the plasma class I HLA with 39-kd heavy chains.
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We describe a patient who developed a markedly prolonged PT, PTT, and thrombin time 13 days after repeat exposure to fibrin sealant during coronary artery bypass grafting and aortic valve replacement. Evaluation revealed an inhibitor to... more
We describe a patient who developed a markedly prolonged PT, PTT, and thrombin time 13 days after repeat exposure to fibrin sealant during coronary artery bypass grafting and aortic valve replacement. Evaluation revealed an inhibitor to bovine thrombin that cross-reacted with human thrombin. In addition an inhibitor to human coagulation factor V was identified. Despite coagulation abnormalities there was no evidence of bleeding. Nevertheless, effective anticoagulation was required to minimize the thrombotic complications associated with the patient&#39;s prosthetic valve. We elected to take a conservative approach and not utilize pharmacologic anticoagulation until there was diminution in the effect of the acquired inhibitors. We report on our patient&#39;s course and review the available literature addressing the management of patients demonstrating inhibitors to blood coagulation factors after repeat exposure to fibrin sealants.
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Research Interests: Immunology, Gastroenterology, Adolescent, Humans, Child, and 15 moreInternal Medicine, Female, Infant, American, Incidence, Antiphospholipid antibodies, Clinical Sciences, Longitudinal data, Enzyme Linked Immunosorbent Assay, Immunoglobulin, Hemolytic Anemia, Lupus Anticoagulant, False Positive, Child preschool, and Antiphospholipid Syndrome
Very little information is available concerning the relationship between metallothionein and disease in humans. Recently, investigators have used the Cd-heme method to measure metallothionein levels in human liver samples obtained from... more
Very little information is available concerning the relationship between metallothionein and disease in humans. Recently, investigators have used the Cd-heme method to measure metallothionein levels in human liver samples obtained from autopsy. This assay, however, is not sensitive enough to measure metallothionein in small tissue biopsy specimens. As an alternative, we report the development of a human metallothionein enzyme-linked immunosorbent assay (ELISA). This assay used high-performance liquid chromatography-purified human metallothionein-1 and purified sheep anti-human metallothionein-1 IgG. A limiting antigen coating concentration of 100 ng/ml and a minimal antibody dilution of 1:4000 were chosen. The sensitivity of the ELISA extended to 300 ng/ml (15 ng). The coefficients of interassay and intraassay variation were 15.4% and 4.2%, respectively. Human livers obtained from autopsy were assayed by this method and the values compared with values obtained by the Cd-heme method....
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SummaryThe use of plasma thrombospondin (TSP) concentration was investigated as an indicator of intravascular platelet activation. Patients (n = 20) with diseases that have known vasculitis were included in the study. The range and the... more
SummaryThe use of plasma thrombospondin (TSP) concentration was investigated as an indicator of intravascular platelet activation. Patients (n = 20) with diseases that have known vasculitis were included in the study. The range and the mean of plasma TSP concentrations of patients with vasculitis were 117 ng/ml to 6500 ng/ml and 791±1412 ng/ml (mean ± SD); the range and the mean of plasma TSP concentrations of control individuals (n = 33) were 13 ng/ml to 137 ng/ml and 59±29 ng/ml. When plasma TSP concentrations were correlated with plasma concentrations of another platelet activation marker, β-thromboglobulin (P-TG), it was found that the TSP concentration inei eased exponentially as the plasma β-TG level rose. A positive correlation between plasma levels of plasma TSP and serum fibrin degradation products was also observed. The results suggest that platelets are the primary source of plasma TSP in patients with various vasculitis and that plasma TSP can be a better indicator than ...
Research Interests: Chemistry, Medicine, Platelet, Fibrin, Humans, and 12 moreChild, Internal Medicine, Vasculitis, Haemostasis and Thrombosis, Glycoproteins, Platelet activation mechanism, Clinical Sciences, Thrombospondins, Blood platelets, Thrombospondin, Platelet-Poor Plasma, and Cardiovascular medicine and haematology
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Female B6C3F1 mice were exposed to ammonium metavanadate (NH4VO3) by intraperitoneal injection every 3 d at 2.5, 5.0, or 10 mg V/kg for 3, 6, or 9 w and were then assayed for alterations in immunoresponsiveness. Resistance to Escherichia... more
Female B6C3F1 mice were exposed to ammonium metavanadate (NH4VO3) by intraperitoneal injection every 3 d at 2.5, 5.0, or 10 mg V/kg for 3, 6, or 9 w and were then assayed for alterations in immunoresponsiveness. Resistance to Escherichia coli endotoxin lethality increased in a dose-dependent manner up to 6 w of exposure, while resistance to viable gram-positive Listeria lethality was depressed in a dose-dependent manner. Comparison of LD20 values indicated a 250-fold decrease in resistance to Listeria at the lowest vanadium exposure and a 40% increase in resistance to endotoxin after the highest vanadium exposure. Peritoneal macrophage phagocytic capacities were decreased in a dose-dependent manner, but viabilities remained unaffected. Rosetting capacity of splenic lymphocytes was increased following vanadium exposure. Liver and splenic enlargement was observed, and examination of splenic tissue indicated enhanced formation of megakaryocytes and red blood cell precursors. Subchronic exposure to vanadium may thus disrupt the normal function of the immune system.
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Research Interests: Immunology, Medicine, Humans, Smoking, Female, and 15 moreMale, Plasma, Artery, Clinical Sciences, Myocardial Infarction, Odds ratio, Age Factors, Hematocrit, Reoperation, Sex Characteristics, Blood coagulation disorders, Platelet transfusion, Erythrocyte transfusion, Cardiovascular medicine and haematology, and Coronary Artery Bypass
Previous studies suggest that recombinant thrombopoietin (rTPO) will increase platelet production in thrombocytopenic neonates. However, the target populations of neonates most likely to benefit should be defined. Studies suggest that... more
Previous studies suggest that recombinant thrombopoietin (rTPO) will increase platelet production in thrombocytopenic neonates. However, the target populations of neonates most likely to benefit should be defined. Studies suggest that rTPO will not elevate the platelet count until 5 days after the start of treatment. Therefore, the neonates who might benefit from rTPO are those who will require multiple platelet transfusions for more than 5 days. This study was designed to find means of prospectively identifying these patients. A historic cohort study of all patients in the neonatal intensive care unit (NICU) at the University of Florida who received platelet transfusions from January 1, 1997, through December 31, 1998, was conducted. Of the 1389 patients admitted to the NICU during the study period, 131 (9.4%) received platelet transfusions. Seventeen were treated with extracorporeal membrane oxygenation and were excluded from further analysis. Of the remaining 114 patients, 55 (48%) received one transfusion and 59 (52%) received more than one transfusion (21 had &amp;amp;amp;amp;amp;amp;gt;4). None of the demographic factors examined predicted multiple platelet transfusions. However, two clinical conditions did; liver disease and renal insufficiency. Neonates who received one platelet transfusion had a relative risk of death 10.4 times that in neonates who received none (p = 0.0001). Neonates who received &amp;amp;amp;amp;amp;amp;gt;4 platelet transfusions had a risk of death 29.9 times that in those who received no transfusions (p = 0.0001). NICU patients with liver disease or renal insufficiency who receive one platelet transfusion are likely to receive additional transfusions. Therefore, these patients constitute a possible study population for a Phase I/II rTPO trial.
Research Interests: Immunology, Evolution, Medicine, Platelet, Humans, and 15 moreRisk factors, Clinical Sciences, Newborn Infant, Intensive Care Unit, Multiple, Time Factors, Thrombopoietin, Risk Factors, Neonatal Intensive Care Unit, Transfusion, Predictive value of tests, Cohort Studies, Extracorporeal Membrane Oxygenation, Platelet transfusion, and Cardiovascular medicine and haematology
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Ristocetin cofactor activity is the test of choice for diagnosing von Willebrand disease. There is however no simple and precise quantitative method for its rapid determination in the clinical laboratory. We describe here a new approach... more
Ristocetin cofactor activity is the test of choice for diagnosing von Willebrand disease. There is however no simple and precise quantitative method for its rapid determination in the clinical laboratory. We describe here a new approach for measuring ristocetin cofactor activity using an ELISA plate reader. Paraformaldehyde-fixed platelets, diluted plasma samples and ristocetin were mixed and incubated in the wells of a microtiter plate. Platelet agglutination was measured by light absorbance at 405 nm. Measurements of platelet agglutination at 1x to 64x dilution of a normal pooled plasma were used to construct a standard curve. Further characterization showed a significant linear correlation between results determined by the new method and by an aggregometer assay (p &amp;amp;lt; 0.0001), and that the intra- and inter-assay variations were 6% and 17%, respectively. Therefore, the newly developed assay provides a simple and rapid way for precise quantitation of plasma ristocetin cofactor activity.
Research Interests: Chemistry, Chromatography, Medicine, Platelet, Humans, and 15 morevon Willebrand factor, Female, Male, Spectrophotometry, Clinical Sciences, Chemical Precipitation, Time Factors, Thrombosis, Reproducibility of Results, Absorbance, Enzyme Linked Immunosorbent Assay, Platelet Count, Blood Proteins, Titrimetry, and Standard Curve
ABSTRACT
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Thermally stimulated discharge (TSD) current studies on negatively‐corona‐charged low‐density polyethylene reveal a deep surface trap distribution centered at 95 °C and a shallow surface and bulk distribution centered at 55 °C. TSD also... more
Thermally stimulated discharge (TSD) current studies on negatively‐corona‐charged low‐density polyethylene reveal a deep surface trap distribution centered at 95 °C and a shallow surface and bulk distribution centered at 55 °C. TSD also shows that corona‐generated neutral excited gas molecules in the surrounding air free the shallow surface trapped charge which moves into the bulk under its own field, and can be retrapped or transit through the sample. Crossover in surface potential decay curves is caused by these molecules, and can be eliminated by removing them from the corona discharge.
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Research Interests: Endocrinology, Biology, Medicine, Biological Sciences, Progesterone, and 15 moreInternal Medicine, Testosterone, Female, Animals, Ovarian Follicle, Estrogens, Chickens, Cell communication, Granulosa Cells, Luteinizing Hormone, Theca, Follicle stimulating hormone, Biology Reproduction, Follicular phase, and Medical and Health Sciences
Research Interests: Algorithms, Incidence Geometry, Adolescent, Medicine, Platelet, and 15 moreHumans, Child, Female, Male, Central Nervous System, Incidence, Clinical Sciences, Aged, Middle Aged, Bone Marrow Transplantation, Adult, Hemorrhage, Platelet Count, Hematopoietic Stem Cell Transplantation, and Child preschool
Transfusions of UV-B–irradiated peripheral blood mononuclear cells (UV-B–PBMCs) from BALB/c (H-2d) mice into CBA (H-2k) mice can induce humoral immune tolerance to H-2d antigens, and the induced tolerance is partially mediated by negative... more
Transfusions of UV-B–irradiated peripheral blood mononuclear cells (UV-B–PBMCs) from BALB/c (H-2d) mice into CBA (H-2k) mice can induce humoral immune tolerance to H-2d antigens, and the induced tolerance is partially mediated by negative regulatory PBMCs. To further identify which subset of spleen mononuclear leukocytes (MNLs) in the tolerant CBA mice is responsible for the negative regulatory activity, adoptive transfer experiments were conducted using spleen MNLs from the tolerant CBA mice. Results showed that only CD4+ T cells could transfer the negative regulatory activity in a dose-dependent manner. This negative regulatory activity was significantly reduced when CD25+ helper T cells were removed. Further study suggested that inhibition of IL-12 production by UV-B–irradiated PBMCs played a role in the induction of immune tolerance. In vitro study of the cytokine production profile by CBA CD4+ T cells, after stimulation with gamma-irradiated BALB/c spleen cells, revealed an enh...
Research Interests: Immunology, Biology, Cytokines, Gamma Rays, Medicine, and 15 moreSpleen, Mice, Animals, Blood, Clinical Sciences, Cytokine, Immune system, Immune Tolerance, Clinical Application, Interleukin, Gamma radiation, Ultraviolet Rays, Adoptive Cell Transfer, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
Interhospital differences in blood transfusion practice during coronary artery bypass graft (CABG) surgery have been noted, but the underlying issues have not been identified. Records of 3217 consecutive CABG cases in five university... more
Interhospital differences in blood transfusion practice during coronary artery bypass graft (CABG) surgery have been noted, but the underlying issues have not been identified. Records of 3217 consecutive CABG cases in five university teaching hospitals in 1992 and 1993 were stratified by hospital, type of revascularization conduit, patients&#39; sex, and other factors. Statistical methods were used to compare patient characteristics, transfusion outcomes, and hospital outcomes. Forward two-step logistic regression using patient likelihood of red cell transfusion factors in the first step and the specific hospital in the second step revealed a significant effect of hospital on the delta odds ratios for red cell transfusion. This finding was confirmed by analyses of a highly stratified subset of cases, males in diagnosis-related group 107 (primary cases of coronary bypass without coronary catheterization) who underwent revascularization with venous and internal mammary artery grafts, revealing variations among hospitals from 109 to 457 units of red cells transfused per hundred cases. Corresponding variations in transfusions of all blood components were from 324 to 1019 units by hospital. Variation in red cell transfusion practice among surgeons in the same hospital was not responsible for these interhospital differences. The effect of the specific hospital on transfusion practice is attributed to institutional differences that, through reasons of training or hierarchy, become ingrained in hospitals.
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Research Interests: Immunology, Clinical Trial, Filtration, Medicine, Platelet, and 12 moreHumans, Clinical Sciences, Reproducibility of Results, Erythrocytes, Leukocytes, Transfusion, Plateletpheresis, Ultraviolet Rays, Blood platelets, Platelet transfusion, Cardiovascular medicine and haematology, and leukocyte Count
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Research Interests: Medicine, Probability, Humans, Heparin, Female, and 15 moreBlood sampling, Male, Clinical Sciences, Newborn Infant, Perinatology, Adverse Event, Thrombosis, Study design, Sensitivity and Specificity, Equipment Design, Factor Xa, Thrombolytic Therapy, Test Methods, Equipment safety, and Paediatrics and reproductive medicine
The use of 8-methoxypsoralen (8-MOP) and UV-A irradiation to inactivate contaminating donor leukocytes in platelet concentrates and to prevent primary alloimmunization against donor class I major histocompatibility (MHC) antigens in mice... more
The use of 8-methoxypsoralen (8-MOP) and UV-A irradiation to inactivate contaminating donor leukocytes in platelet concentrates and to prevent primary alloimmunization against donor class I major histocompatibility (MHC) antigens in mice was investigated. CBA/CaH-T6J mice with the H2k haplotype and BALB/cByJ mice with the H2d haplotype were used as donors and recipients, respectively. The mixed leukocyte reaction between these two strains of mice showed that treatment of spleen cells with 500 ng/mL 8-MOP and 5J/cm2 UV-A inhibited 99% of responder and 92% of stimulator function. There was no measurable loss of platelet aggregating activity after the treatment. After two weekly transfusions of platelets without any treatment, 93% of control mice (n = 15) developed anti-H2k antibody. In contrast, only 33% of mice (n = 15) receiving platelets treated with 8-MOP and UV-A became alloimmunized. After six weekly platelet transfusions, all mice became alloimmunized. Nevertheless, the mean ti...
Research Interests: Medicine, Platelet aggregation, Mice, Haplotypes, Beta decay, and 12 moreAnimals, Blood, Clinical Sciences, Major histocompatibility complex, Blood Transfusion, Leukocytes, Enzyme Linked Immunosorbent Assay, Control Group, Blood platelets, Cardiovascular medicine and haematology, Paediatrics and reproductive medicine, and In Vitro Techniques
SummaryThrombospondin (TSP) is a major platelet secretory glycoprotein. Earlier studies of various investigators demonstrated that TSP is the endogenous platelet lectin and is responsible for the hemagglutinating activity expressed on... more
SummaryThrombospondin (TSP) is a major platelet secretory glycoprotein. Earlier studies of various investigators demonstrated that TSP is the endogenous platelet lectin and is responsible for the hemagglutinating activity expressed on formaldehyde-fixed thrombin-treated platelets. The direct effect of highly purified TSP on thrombin-induced platelet aggregation was studied. It was observed that aggregation of gel-filtered platelets induced by low concentrations of thrombin (≤0.05 U/ml) was progressively inhibited by increasing concentrations of exogenous TSP (≥60 μg/ml). However, inhibition of platelet aggregation by TSP was not observed when higher than 0.1 U/ml thrombin was used to activate platelets. To exclude the possibility that TSP inhibits platelet aggregation by affecting thrombin activation of platelets, three different approaches were utilized. First, by using a chromogenic substrate assay it was shown that TSP does not inhibit the proteolytic activity of thrombin. Second...
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In order to provide a simple and specific assay for the detection and quantitation of IgG and IgM anti-HLA antibodies in sera, HLA antigens purified from a pool of 240 random donor platelets were used to develop a solid-phase... more
In order to provide a simple and specific assay for the detection and quantitation of IgG and IgM anti-HLA antibodies in sera, HLA antigens purified from a pool of 240 random donor platelets were used to develop a solid-phase enzyme-linked immunoassay (EIA). The reference values for identifying the presence of IgG or IgM anti-HLA antibodies were determined by assaying sera from 39 healthy individuals without prior HLA alloimmunization. The assay was evaluated by studying sera from 122 patients who had been characterized previously for panel reactive antibodies by the lymphocytotoxicity assay (LCA). A significant linear correlation between two assays was noted (r = 0.8, P = 0.0001). Further analyses of the data demonstrated that the newly developed EIA has 100% specificity and 95.3% sensitivity as compared with the LCA. Additional studies revealed that patients whose PRA increased or decreased over time were in parallel with antibody levels measured by EIA. When the EIA was used to measure anti-HLA antibody titers, it was more sensitive than the LCA. Since the EIA is sensitive, specific, and technically less demanding, it should provide an useful alternative to reduce the number of the more laborious panel studies for monitoring anti-HLA antibody status in candidates for organ transplantation and recipients of blood transfusions.
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Research Interests: Biology, Cell Adhesion, Medicine, Spleen, Mice, and 15 moreAnimals, Blood, Immunosuppression, Clinical Sciences, Major histocompatibility complex, Blood Transfusion, Immune Tolerance, Leukocytes, Clinical Application, Adoptive Transfer, Cell Membrane, Antibody Response, Platelet transfusion, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
Our recent study suggested the reverse relationship between the production of interleukin-10 (IL-10) and IL-12 in dendritic cells (DCs) activated by lipopolysaccharide (LPS) or LPS plus interferon (IFN)-gamma. In the present study, a... more
Our recent study suggested the reverse relationship between the production of interleukin-10 (IL-10) and IL-12 in dendritic cells (DCs) activated by lipopolysaccharide (LPS) or LPS plus interferon (IFN)-gamma. In the present study, a series of experiments were performed to investigate the mechanisms responsible for this reverse relationship. Our results showed that neutralization of the secreted IL-10 by antibody could enhance the production of IL-12. Neutralization of IL-12 by antibody did not affect the IL-10 production. Addition of exogenous IL-10 suppressed the production of IL-12 by activated DCs, and addition of exogenous IL-12 did not affect IL-10 production. TaqMan real-time reverse transcriptase-polymerase chain reaction supported the fact that the observed effects occurred at mRNA transcription level. We also found that LPS or LPS plus IFN-gamma significantly enhanced the phosphorylation of extracellular signal-regulated protein kinase (ERK) and p38 mitogen-activated protein kinase. In addition, inhibition of ERK by PD98059 significantly suppressed IL-10 and increased the IL-12 production. Exogenous IL-10 reversed the upregulated production of IL-12 induced by PD98059. The above findings suggest a unidirectional negative autocrine regulation of IL-12 by IL-10 in activated DCs and that activation of ERK involves the differential production of IL-10 and IL-12 by activated DCs. Thus, the regulation of differential production of IL-10 and IL-12 may play an important role for DCs in priming T helper 1 (Th1) or Th2 in the immune responses.
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Research Interests: Protein Folding, Biology, Biological Chemistry, Transcription Factors, Medicine, and 14 moreBiological Sciences, Humans, Mice, Animals, C2H2 zinc fingers, Biological, Phosphorylation, CHEMICAL SCIENCES, Amino Acid Sequence, Base Sequence, DNA binding proteins, Molecular Sequence Data, Thymus Gland, and Medical and Health Sciences
ABSTRACT Pancreatic islets of Langerhans exhibit an architecture and cellular organization ideal for rapid, yet finely controlled, responses to changes in blood glucose levels. In type I, insulin-dependent diabetes (IDD), this... more
ABSTRACT Pancreatic islets of Langerhans exhibit an architecture and cellular organization ideal for rapid, yet finely controlled, responses to changes in blood glucose levels. In type I, insulin-dependent diabetes (IDD), this organization is lost as a result of the progressive autoimmune response which selectively destroys the insulin-producing pancreatic beta cells. Since beta cells are perceived as end-stage differentiated cells having limited capacity for regeneration in situ, individuals with IDD resulting from beta cell loss or dysfunction require life-long insulin therapy. Efforts to produce islet neogenesis or initiate islet growth in vitro from either fetal or adult tissue have had minimal success. We now report that pancreatic-derived, pluripotent islet-producing stem cells (IPSCs), isolated from prediabetic mice, can be grown in long-term cultures and differentiated into immature functional islet-like structures containing cells which express low levels of insulin, glucagon and/or somatostatin. When such in vitro grown islets were implanted into clinically diabetic NOD mice, the implanted mice were successfully weaned from insulin long-term (&gt;50 days) without ill effects. The implanted mice maintained blood glucose levels just above euglycemic (180-220 mg/dl) and showed no signs of disease. Thus, this technical breakthrough provides new therapeutic approaches to diabetes as an alternative to insulin therapy.
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The tests currently used to monitor atopic dermatitis in children--serum IgE level and eosinophil count--are not sensitive enough to accurately track the course of the disease. Because previous studies have shown that atopic dermatitis is... more
The tests currently used to monitor atopic dermatitis in children--serum IgE level and eosinophil count--are not sensitive enough to accurately track the course of the disease. Because previous studies have shown that atopic dermatitis is an inflammatory disease and because our previous work has shown that plasma thrombospondin level correlates well with the course of other inflammatory diseases, we conducted this study to determine the relationship between plasma thrombospondin level and the severity of skin inflammation in children with atopic dermatitis. Eosinophil count, serum IgE level, and plasma thrombospondin level were measured in 48 children with atopic dermatitis at onset of flare-up, 2 weeks after treatment started, and 2 months after treatment started. The results of all three tests were better after 2 months of therapy than they had been at the initial visit, but only plasma thrombospondin level showed a statistically significant decrease, which coincided with clinical improvement. Plasma thrombospondin level seems to correlate with the clinical course of patients with atopic dermatitis better than do serum IgE level or eosinophil count.
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Research Interests: Treatment Outcome, Medicine, Platelet, Humans, Female, and 12 moreBlood, Male, Thrombocytopenia, Clinical Sciences, Middle Aged, Time Factors, Platelet Count, Mean platelet volume, Blood platelets, Platelet transfusion, Cardiovascular medicine and haematology, and Paediatrics and reproductive medicine
Half of 30 human polyclonal IgM rheumatoid factors (RF) showed positive ELISA reactions with affinity-isolated human class I molecules (A2 and B7). Positive RF reactivity with isolated class I heavy chains indicated that anti-class I RF... more
Half of 30 human polyclonal IgM rheumatoid factors (RF) showed positive ELISA reactions with affinity-isolated human class I molecules (A2 and B7). Positive RF reactivity with isolated class I heavy chains indicated that anti-class I RF interaction did not merely reflect RF anti-beta 2m specificity. Cross-reactions between antigenic determinants on human IgG, class I molecules, and beta 2m reacting with RF could be demonstrated by ELISA inhibition. When gene products of A2 alpha 2 exons 2, 3, and 4 were synthesized as overlapping heptamers on polypropylene pins, six RF-reactive epitopes within solvent accessible class I HLA regions were identified: EPRAPWI, QEGPEYW, QTHRVDL (second A2 exon), EQLRAYL, GTCVEWL, and WLRRYLE (third A2 exon). Glycine-alanine substitution for each residue within these RF-reactive sites identified R48, W51, E55, Y107, R108, W147, Q155, and E172 as immunodominant residues for RF reactivity. Many of these RF-reactive class I regions also showed positive ELIS...