Lupus anticoagulant IgGs do not depend on serum for the induction of cyclooxygenase-2 by human endothelial cells.
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Biomedical subjects
Publications and source records attributed to A Habib.
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The effect of IgGs from 4 patients with antiphospholipid antibodies and elevated excretion of urinary 11-dehydro-thromboxane B2 was evaluated on the production of prostacyclin by human endothelial cells in culture. After 6 h incubation, there was no change in 6-keto-prostaglandin F1 alpha in the supernatant. However patients' IgGs induced a marked increase in cyclooxygenase (Cox) activity compared to IgGs from 2 normal individuals or a commercial pool of IgGs from normal donors, tested by adding exogenous arachidonic acid. Western blot analysis of the cellular Cox content using antibodies specific for the different forms of the enzymes revealed that patients' IgGs stimulated the synthesis of the newly described inducible Cox-2 without affecting the constitutive Cox-1. This effect was partially neutralized by preincubating the IgGs with phospholipids. The induction was dependent on the amount of IgGs; it was visible at 2 h and persisted up to 24 h. Analysis of mRNA levels showed a pattern of variation in good agreement with the results obtained for protein. The protein kinase inhibitor H-7 or long-term incubation of cells with PMA strongly reduced the induction. These results suggest that antiphospholipid antibodies may not prevent the potential of the vascular cells from generating higher amounts of prostacyclin in response to acute episodes of thrombosis.
Cyclooxygenase (Cox) exists in two forms in human endothelial cells (HUVEC). We have raised antibodies that recognize the sequence of the carboxyl-terminal portion of the human Cox-2 (C)-NASSSRSGLD-DINPTVLLK. Cyclooxygenase activity of HUVEC challenged with interleukin 1 alpha or a phorbol ester increased in parallel with the mass of a protein doublet analyzed by Western blot using antibodies directed against the Cox-2 peptide; a monoclonal antibody directed against Cox-1 showed a small change in protein mass. A 35S-labeled protein doublet with a molecular mass of approximately 70,000 daltons was immunoprecipitated with the anti-Cox-2 antiserum in L-[35S] methionine-labeled cells stimulated with interleukin 1 alpha. This protein was not recovered by pretreating the antiserum with the Cox-2 peptide before immunoprecipitation. A minor variation in 35S-immunoprecipitated protein was obtained with the polyclonal anti-Cox-1 antibody. Both immunoprecipitated Cox-1 and Cox-2 possessed cyclooxygenase activity that was inhibited by flurbiprofen. Endoglycosidase H treatment of immunoprecipitated Cox-2 proteins caused a decline in the apparent molecular size similar to that observed with immunoprecipitated Cox-1 or sheep cyclooxygenase but did not suppress the doublet. These results show by direct protein measurement that HUVEC synthesize the novel Cox-2 under appropriate stimulation, with little changes of Cox-1.
Seven distinct mouse monoclonal antibodies (mAbs) directed against human endothelin-1 (ET-1) have been obtained. On the basis of specificity studies performed with competitive immunoassays and of complementary binding studies, these mAbs were classified in two groups. mAbs of group A (Endo-4, -5, -6 and -10) were shown to be directed against the N terminal loop while those of group B (Endo-2, -8 and -18) recognized the C terminal part of the peptide. A pair of monoclonal antibodies with optimal properties for a two-site immunometric assay were selected and the test was performed in 96-well microtiter plates coated with one mAb (Endo-18), while another mAb (Endo-4) covalently labeled with enzyme acetylcholinesterase was used as tracer. Under optimal conditions, the assay appeared to be very sensitive since concentrations as low as 1 pg/ml could be significantly detected. The precision was also very good with a coefficient of variation below 10% from 3 to 250 pg/ml. The assay was specific for mature endothelin presenting no cross-reactivity with the precursor Big ET-1. On the other hand, strong cross-reactivity was observed with other ET-1-related peptides, including ET-2, ET-3, VIC peptide and sarafotoxin 6-b. The assay permitted specific determination of ET-1 in supernatants of cultured endothelial cells and the validity of the test was demonstrated by HPLC fractionation experiments. In addition, the assay also appeared to be suitable for direct determination of ET-1 in plasma. Studies performed with plasma from healthy subjects revealed that circulating levels of ET-1 are below or close to the detection limit of the method (< 8 pg/ml).
This study was undertaken in Long Evans rat to investigate the effect of a single therapeutic as well as toxic dose of indomethacin on the gastrointestinal mucosa. The effect was studied morphologically six hours after oral administration of the drug. The affected tissue was then examined histologically. The histomorphological evaluation revealed that the drug has induced acute hemorrhagic erosive gastritis in the fasted animals (6 mg/Kg body weight) where as in normally fed (10 mg/kg body weight) rats the small intestinal mucosal inflammation and erosions were predominant.
Leukotriene (LT) A4 metabolism was studied in human platelets and endothelial cells, since both cells could be involved in transcellular formation of LTC4. Upon addition of exogenous LTA4, both cells produced LTC4 as a major metabolite at various incubation times, and no LTB4, LTD4, or LTE4 was detected. Kinetic studies revealed a higher apparent Km for LTA4 in endothelial cells as compared to platelets (5.8 microM for human umbilical vein endothelial cells (HUVEC) versus 1.3 microM for platelets); platelets were more efficient in this reaction with a higher Vmax (174 pmol/mg protein/min) versus 15 pmol/mg protein/min in HUVEC. The formation of LTC4 and corresponding kinetic parameters were not modified when platelets or endothelial cells were stimulated by thrombin prior to or simultaneously with the addition of LTA4. In both cells LTC4 synthase activity was not modified by repeated addition of LTA4 showing that it is not a suicide-inactivated enzyme. Furthermore, in platelets and endothelial cells, the enzyme activity was localized in the membrane fraction and was distinct from cytosolic glutathione-S-transferases. Platelet membrane fractions showed apparent Km values of 31 microM and 1.2 mM for LTA4 and GSH, respectively. Inhibition of LTC4 formation from platelets and endothelial cells preparations by S-substituted glutathione derivatives was correlated to the length of the S-alkyl chain. The same substances inhibited cytosolic glutathione-S-transferases with significantly lower IC50, confirming the distinct nature of the two enzymes. These results show that platelets and HUVEC possess similar enzymes for the production of LTC4 from LTA4; however, platelets seem to have a higher efficiency than HUVEC in performing this reaction.
We studied histology, findings on H-1 magnetic resonance (MR) imaging, and correlations of P-31 MR spectroscopy with microelectrode pH and pO2 measurements in the BA1112 rhabdomyosarcoma in WAG/Rij/Y rats. Intratumoral hemorrhage was a prominent feature on MR images and pathologic specimens. Eosinophilic necrosis could be seen microscopically but was not discernible on images. The peaks seen on P-31 MR spectra were similar to those reported in other tumors. The intratumoral pH was neutral despite low pO2 values and P-31 MR evidence for impaired metabolic status.
Myocarditis may be a serious extrahepatic complication of hepatitis. In this fatal case of serologically documented hepatitis B viral hepatitis, acute myocarditis was present, with histologic features consistent with a viral pathogenesis. Hepatitis B surface antigen was demonstrated by immunoperoxidase methods in small intramyocardial vessels, suggesting that hepatitis B virus infected the heart. The resulting inflammatory heart disease may have been caused either directly, by virus infecting the myocardium, or indirectly, by an immune-mediated mechanism.
The authors describe the occurrence of Hodgkin's disease in a patient suffering from actinic reticuloid since 12 years. The severity of the disease (MED less than 1 mJ/cm2 UVB) imposed corticosteroids and immunosuppressive therapy (chlorambucil). The role of actinic reticuloid's pathogeny (persistent light reaction) and principally use of immunosuppressive drugs are discussed and may explain the occurrence of Hodgkin's disease.
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A case of acute myelogenous leukemia terminating in megakaryocytic myelosis is reported. There was a severe, apparently neoplastic proliferation of megakaryocytes so different from that seen in acute granulocytic leukemia or myelofibrosis that a diagnosis of acute megakaryocytic myelosis was warranted. The clinical and pathologic findings of the case are presented in detail. The literature of this extremely rare hematologic disease is reviewed, and differentiation from chronic and malignant myelosclerosis is discussed.
Flat wart-like lesions of two patients with epidermodysplasia verruciformis (EDV) were associated with pink, tan or depigmented pityriasis-like macules. There was no familial history nor mental retardation. Human papilloma virus (HPV) type 5 was demonstrated in the two patients and was responsible for the pityriasis-like lesions. Malignancies seem to be closely related to HPV-5 infection since bowenoid transformation occurred in the 2 patients. Immunological studies showed an increase of serum IgE in both patients and an important decrease of IgM in one of them. Most of delayed hypersensitivity skin tests were negative. T-cell percentages (E rosettes) were decreased in one patient, normal for the other one. The mitogenic response to PHA and ConA was markedly depressed in the two patients but returned to normal values in one of them after 3 months of aromatic retinoid (Ro 10-9359) treatment. Beside the viral type, the defect of cell mediated immunity could play an important role in the disease and in the malignant conversion of the lesions. A long-term preventive treatment by retinoic acid derivatives could be of interest for patients with HPV type 5 EDV.
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The case of a 33-year-old woman with primary carcinoid of the uterine cervix is reported. Primary carcinoid tumor is well known to occur in organs such as the gastrointestinal tract, lung and gonads. However, its occurrence in the uterine cervix is rare. To our knowledge, primary carcinoid of the uterine cervix has not been reported from the United States, although it has been well documented by non-American authors. Light microscopically, the tumor was characterized by formation of solid nests, trabeculae and glands. The cells therein showed argyrophil granules but were negative for argentaffin reaction. Electron microscopy revealed the presence of numerous neurosecretory granules and microfilaments. On the basis of light microscopic ultrastructural and cytochemical properties, the tumor is believed to arise from the normal argyrophil cell of the cervix and is regarded as an endocrine tumor, a member of the group of neoplasms called apudomas.
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Two vaccinia viruses isolated from patients with vaccinial complications (vaccinial ulcer, postvaccinial seizures) showed qualitative differences from the original parental strain. After intradermal injection of the viruses into the rabbit marked necroses developed, which the original strains did not produce. While the parental virus did not grow on the chorioallantoic membrane at 41 degrees C after 2 days incubation, the vaccinia variant produced typical lesions at that temperature. Also the yield of infectious virus on various cell systems was 1--2.5 logs higher for the virus than for the original vaccine strain. With the plaque technique differences were seen in the appearance and size of plaques between the variant and the parental vaccinia strain. These results indicate that virus of an increased pathogenicity could be isolated from the patients and this might be causally connected with the postvaccinial complications from which they were suffering.
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