摘要:

AbstractThe endothelial cell line ECV304 is a spontaneously transformed cell line established from human umbilical vein. The characterization of tissue factor (TF) expression by ECV304 cells has been accomplished in this study. ECV304 cells expressed both TF mRNA and antigen (TFag) constitutively. In ECV304 cell lysates, the levels of TFag(1.4 ± 0.3 ng of TFag/106cells) were considerably higher than in THP‐1 monocytoid cells (0.07 ± 0.03 ng of TFag/106cells). TFagwas also detected on the ECV304 cell surface by flow cytometric studies. In binding analyses, 3.5 ± 0.7 × 104molecules of TF per cell were estimated, similar to the amounts found in ECV304 cell lysates (2.9 ± 0.6 × 104molecules/cell), suggesting that all TFagwas translocated to the cell surface. Phorbol myristate acetate (PMA) stimulation of ECV304 cells resulted in an increase of TF mRNA levels, which was abrogated when gene transcription was impaired, suggesting a transcriptional regulation of the TF gene by PMA. In contrast, TFagwas not elevated by PMA‐stimulation, indicating the existence of additional posttranscriptional mechanisms. Thus, ECV304 cells constitute a singular endothelial cell model for exploring the regulation of TF expression. Am. J. Hematol. 56:71–78, 1997. © 1997 Wil

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<71::AID-AJH1>3.0.CO;2-Y

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractThe AML14.3D10 human myeloid leukemic cell line expresses receptors for granulocyte‐macrophage colony stimulating factor (GM‐CSF) and interleukin‐5 (IL‐5), but not IL‐3. We have found that this cell line produces GM‐CSF in amounts up to 113 pg/ml in culture supernatants. Deprivation of endogenous GM‐CSF by addition of neutralizing anti‐GM‐CSF antibody strongly inhibits proliferation of the cells, suggesting a GM‐CSF autocrine growth mechanism. To examine whether endogenously produced GM‐CSF activates intracellular GM‐CSF/IL‐3/IL‐5‐related signal transduction pathways, we performed anti‐phosphotyrosine immunoblotting of cell lysates of AML14.3D10 cells before and after deprivation of endogenous GM‐CSF. We found constitutive tyrosine‐phosphorylation of a number of proteins in AML14.3D10 that could not be detectably increased by the addition of exogenous GM‐CSF, IL‐3, or IL‐5. However, GM‐CSF‐deprived cells demonstrated a marked increase in phosphorylation of proteins of identical molecular mass following addition of GM‐CSF and IL‐5, but not IL‐3, consistent with the receptor expression of the cells and the known use of the same signaling pathways by the three cytokines. This suggests that AML14.3D10 cells use endogenously produced GM‐CSF to activate signal transduction pathways, interfering with activation by exogenous cytokine until the endogenous stimulation is removed. We then assessed the activation of the β‐subunit common to the GM‐CSF/IL‐3/IL‐5 receptors (βc), JAK2 and p53/56lyn,known to be involved in the common signaling pathways of the three cytokines. We found that phosphorylation of βc and JAK2 in response to GM‐CSF and IL‐5 could be markedly enhanced by depriving cells of endogenous GM‐CSF. Constitutive hyperphosphorylation oflynwas found in AML14.3D10 cells, and no further activation oflynin response to cytokine was demonstrable in GM‐CSF‐deprived cells, suggesting thatlynis activated in this cell line by a mechanism other than GM‐CSF. These studies represent the first demonstration of autocrine activation of intracellular cytokine signaling pathways by malignant hematopoietic cells. Because the addition of anti‐GM‐CSF to cell cultures improved responsiveness of intracellular signal transducing molecules to exogenous GM‐CSF and IL‐5, it can be inferred that endogenously produced GM‐CSF exerts its effects by secretion and binding to surface GM‐CSF receptors, although an intracellular component to signaling cannot be excluded. These observations provide further information regarding an autocrine contribution to leukemic cell growth, and establish a new

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<79::AID-AJH2>3.0.CO;2-Y

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractMale (NZW × BXSB)F1 (W/BF1) mice develop a systemic lupus‐like syndrome characterized by thrombocytopenia, coronary vascular disease, nephritis, and anticardiolipin antibodies. Three stable hybridoma cell lines secreting monoclonal anticardiolipin antibodies were developed from these mice by fusing their splenic lymphocytes with nonsecreting myeloma cell line, NS‐1. Monoclonal antibody A1.17 reacted with cardiolipin in a β2‐Glycoprotein I‐dependent manner. The epitope for this antibody consisted of β2‐glycoprotein I bound to cardiolipin or immobilized on plastic plates. Other anionic phospholipid‐binding proteins, such as prothrombin or annexin V, had no significant effect in the reactivity of these antibodies. The specificity is similar to the autoimmune anticardiolipin antibodies described in patients with systemic lupus erythematosus and other infectious diseases. In contrast, monoclonal antibodies A1.72 and A1.84 reacted with cardiolipin in the absence of β2‐glycoprotein I. β2‐glycoprotein I, either in the fluid phase or bound to cardiolipin, inhibited the binding of these antibodies. The specificity of the latter two antibodies was similar to that described in patients with syphilis and allied disorders. Both types of antibodies had lupus anticoagulant properties. Thus lupus‐prone male (NZW × BXSB)F1 (W/BF1) mice develop both β2‐glycoprotein I‐dependent and β2‐glycoprotein I‐independent anticardiolipin antibodies. Am. J. Hematol. 56:86

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<86::AID-AJH3>3.0.CO;2-X

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractVascular complications are the main cause of morbidity in diabetes mellitus. To evaluate lipoprotein and hemostatic parameters and their relationship with clinically detectable microangiopathy, we studied 58 insulin‐dependent diabetes mellitus patients and 60 controls matched for age, sex, and body mass index. Thirteen patients presented clinically detectable microangiopathy (8 retinopathy and 5 both retinopathy and microalbuminuria). A cross‐sectional study of lipid profile, coagulation parameters, and a flow‐cytometric evaluation of tissue factor expression in normal monocytes induced by patient plasma were performed. Patients were re‐evaluated for microangiopathy in a 3‐year median follow‐up. Patients showed triglyceride enrichment in low (P= 0.00002) and high density lipoproteins (P= 0.004) and increased levels of D‐dimer (P<0.00001), prothrombin fragment 1 + 2 (P<0.00001), and thrombin‐antithrombin III complex (P= 0.0001). Patients with clinically detectable microangiopathy had increased type 1 plasminogen activator inhibitor (P= 0.00001), thrombomodulin (P= 0.02), and induced monocyte tissue factor expression (P<0.00001). Nine patients developed clinically detectable microangiopathy in the follow‐up and the only predictive variable was increased induced tissue factor expression. In conclusion, in these patients elevated thrombin and fibrin generation reflects a hypercoagulable state but clinically detectable microangiopathy seems related to endothelial cell injury markers and to increased induced tissue factor expression on monocytes. Am. J. Hematol. 56:93–99, 1997. ©

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<93::AID-AJH4>3.0.CO;2-W

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractWe have tested the hypothesis that dense cell formation in sickle cell disease is associated with increased binding of calpromotin to the membrane, an event that occurs during the activation of calcium‐dependent potassium transport. By SDS polyacrylamide gel electrophoresis, we found that sickle cell membranes contained more calpromotin than did normal membranes when stained with Coomassie brilliant blue or when transferred to nitrocellulose paper and immunostained with horseradish peroxidase. Also, the membranes from dense sickle cells contained significantly (P= 0.00055) higher levels of calpromotin, 2.62 ± 1.59 μg/mg membrane protein, compared to light sickle cells, 1.40 ± 0.70 μg/mg membrane protein, when measured by an enzyme‐linked immunosorbent assay. The ratio of calpromotin associated with dense cell membranes to light cell membranes was significantly greater than 1.0 (P<0.00005). Transmission electron micrographs of immunogold‐labelled membranes supported the increase in calpromotin binding in dense sickle cell membranes. In addition, the immunogold probe demonstrated clustering, which was not observed in light sickle cell membranes nor in normal membranes. Finally, we incubated HbSS cells in vitro using a repetitive deoxygenation/reoxygenation procedure to produce dense cells and then measured the levels of calpromotin associated with their membranes. As expected, the levels of calpromotin bound to the membrane doubled during the procedure relative to the basal levels at the beginning of the incubation. The correlation coefficient, calculated between the increase in dense cell formation and the increase in calpromotin associated with the membrane, was statistically significant (P= 0.038). The results demonstrate that an increase in calpromotin binding to the membrane is associated with dense cell formation presumably through the activation of the calcium‐dependent potassium channel. Am. J. Hematol. 56:100–106, 1997. © 1997 W

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<100::AID-AJH5>3.0.CO;2-2

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractDefects of α spectrin have been identified in many cases of hereditary elliptocytosis (HE) and hereditary pyropoikilocytosis (HPP). To aid in the genetic analysis of families with these disorders, the locations of three α‐spectrin gene polymorphisms were mapped, the genetic basis of these polymorphisms identified, and PCR‐based assays designed for their identification. The frequencies of these polymorphisms were determined in two populations and in patients with αI/50a HE and HPP. These studies identified two distinct haplotypes and provided evidence that two HE/HPP mutations associated with the αI/50a protein phenotype, L207P and L260P, arose on separate chromosomal backgrounds. Am. J. Hematol. 56:107–111, 1997. © 1997 Wiley

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<107::AID-AJH6>3.0.CO;2-2

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractThe effectiveness of continuous infusion porcine factor VIII (PFVIII) has been evaluated in the treatment of 7 consecutive patients with factor VIII(FVIII) inhibitors. Two patients had hemophilia A and five were nonhemophiliacs with acquired FVIII inhibitors. The median pretreatment anti‐porcine FVIII titre was 0.2 (range: 0–15.0) Bethesda units (BU), and the anti‐human FVIII titer was 12.0 BU (range: 2.4–50.0). All patients presented with major bleeding. Patients were given a bolus dose of PFVIII followed by continuous infusion. Six patients also received immunosuppressive therapy. Therapeutic FVIII levels (>0.5 U/ml) were achieved in 6 of 7 patients at a median time of 12.5 hr, and then maintained with continuous infusion PFVIII. Six patients were treated for more than 7 days, and in four of these there was a decline in FVIII recovery between days 7 to 11, presumably related to a rising antibody response to PFVIII. These four patients were plasmapheresed and the three patients with autoantibodies recovered therapeutic FVIII levels but this did not occur in the patient with hemophilia. Thrombocytopenia developed in 4 patients at days 18 to 24, with the platelet count falling to 11 to 87 × 109/L, and the PFVIII was discontinued in 3 patients. All patients recovered from the acute bleeding events. With prolonged immunosuppressive therapy, the FVIII inhibitor disappeared in all patients with autoantibodies and there have been no relapses after a median follow‐up period of 581 days. This study demonstrates that continuous infusion PFVIII is an effective therapy for patients with FVIII inhibitors, but that prolonged treatment is associated with the development of inhibitors to porcine FVIII and severe thrombocytopenia, which readily corrects with discontinuation of PFVIII. Am. J. Hematol. 56:112–118, 1997. © 1997 Wi

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<112::AID-AJH7>3.0.CO;2-1

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractImmune thrombocytopenia due to passive transfer of anti‐PIA1alloantibody has been noted as a rare but potentially dangerous complication of plasma transfusions. We report a patient with a preoperative platelet count of 241 × 109/l who developed severe thrombocytopenia within 2 hr following transfusion of 2 U of fresh frozen plasma. The plasma donor was found to be a PIA1‐negative woman. The platelet count of the PIA1‐positive patient recovered within 7 days to normal values. In the frozen plasma, excessive antibody binding to GPIIb‐IIIa on the recipient's platelets was detected. The antibody was shown to have anti‐PIA1‐specificity. Only 40 min after transfusion of the frozen plasma, no antibody was detected in the plasma of the recipient. This case suggests that passively administered anti‐PIA1alloantibody is immediately adsorbed onto the recipient's platelets and thus removed from circulation. Am. J. Hematol. 56:119–121, 1997. © 199

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<119::AID-AJH8>3.0.CO;2-1

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractWe compared the expression of the BCL‐2 oncoprotein (p26) on B cells from 24 patients with splenic lymphoma with circulating villous lymphocytes (SLVL) with that observed on normal, mature B lymphocytes and on neoplastic B cells from 91 patients with other chronic B‐cell malignancies. SLVL B cells showed levels of p26 intermediate between those found on normal B lymphocytes and on neoplastic B cells from patients with other chronic B‐cell lymphoproliferative disorders. Am. J. Hematol. 56:122–125, 1997. © 1997 Wiley

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<122::AID-AJH9>3.0.CO;2-0

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY

摘要:

AbstractWe periodically analyzed bone‐marrow cytogenetic features in 8 patients belonging to a series of 38 subjects with polycythemia vera (PV), all treated with recombinant interferon‐alpha 2a (rIFN‐alpha) at a weekly dose of 9,000,000 IU. Six out of these 8 patients never showed any chromosome alterations, while 2 displayed at diagnosis the presence of trisomy 8 in all bone‐marrow metaphases. Interestingly enough, in these 2 patients rIFN‐alpha treatment was able to induce not only complete hematological response but also the disappearance of trisomy 8, as shown by conventional cytogenetic investigation and fluorescence in situ hybridization performed on bone‐marrow cells after 1 year of treatment. This finding indicates that, as previously shown in chronic myeloid leukemia, in PV rIFN‐alpha can also eradicate the malignant clone by means of a selective effect on bone‐marrow transformed cells. Am. J. Hematol. 56:126–128, 1997. © 19

ISSN:0361-8609    

DOI:10.1002/(SICI)1096-8652(199710)56:2<126::AID-AJH10>3.0.CO;2-A

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1997

数据来源: WILEY