A case of steroid responsive pulmonary hyalinising granuloma: complicated by deep venous thrombosis.

A case of pulmonary hyalinising granuloma (PHG) complicated by deep venous thrombosis (DVT) is presented. The DVT was associated with the presence of a lupus anticoagulant. In the past PHG has been linked to various auto-antibodies, but to the best of the authors' knowledge, this is the first case reporting PHG in association with a lupus anticoagulant and clinically significant venous thrombosis. Historically, PHG has been regarded as poorly corticosteroid responsive. However, the patient in this case study responded dramatically to prednisone. This case study suggests that in selected patients with pulmonary hyalinising granuloma experiencing disabling symptoms and worsening pulmonary function, a trial of corticosteroids may be warranted.

Heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia with or without thrombosis has been recognized increasingly as a serious complication of heparin use. This article reviews type II heparin-induced thrombocytopenia, which is mediated by an antibody that in most cases has specificity for a complex between heparin and platelet factor 4, a secreted platelet alpha-granule protein. The antibody-heparin-platelet factor 4 complex can activate platelets and endothelial cells, thereby initiating thrombosis. Clinical thrombosis in this syndrome may be arterial or venous. Treatment of the syndrome requires discontinuation of heparin and institution of an alternative anticoagulant.

Multiple myeloma: a patient with unusual features including intracranial and meningeal involvement, testicular involvement, organomegaly, and plasma cell leukemia.

A 27-year-old Chinese male presented with multiple myeloma. Over his 18-month course he manifested a number of unusual features of his disease including his young age, marked organomegaly, a testicular plasmacytoma, multiple intracranial extraskeletal plasmacytomas and meningeal involvement, and peripheral blood plasmacytosis. The case is described and recent literature on these rare manifestations is reviewed.

Reduced thrombogenicity of type VI collagen as compared to type I collagen.

Type VI collagen has been recently identified as a major constituent of vascular subendothelium where it serves as a binding site for von Willebrand factor. The present study compares the functional characteristics of type VI collagen with those of type I collagen with respect to platelet aggregating and secretory activities. The differences between the two collagens in platelet aggregation and serotonin and beta-thromboglobulin release were found to be highly significant (p < 0.001, p < 0.0007, p < 0.005 respectively). Our results indicate that under in vitro conditions, type VI collagen stimulates a significantly lesser platelet activation and aggregation response than collagen I, suggesting that type VI collagen may play a role in vivo to limit the platelet thrombotic response following injury to the vascular subendothelium.

Inducible release of an endothelial cell-specific protein.

The authors investigated the release of an endothelial cell-specific protein (E92) by cultured porcine aortic endothelial cell cultures. Under normal culture conditions, endothelial cells released little or no E92 into the culture supernatant. Treatment with thrombin (0.01 to 10 units/ml), endotoxin (0.01 to 10 micrograms/ml), or interleukin-1 (0.01 to 3.0 units/ml), however, caused significant, dose-dependent increases in E92 detectable in the culture supernatants. Time-course experiments showed that maximum release of E92 into cellular supernatants occurred 24 hours after stimulation with all mediators. Parallel experiments used 51Cr-loaded endothelial cells as a measure of lethal cellular injury. None of the mediators caused significant injury at the doses observed to induce release of E92. These results suggest that the release of E92 into the supernatants of cultured endothelial cells is an inducible event. The data also support the hypothesis that detection of E92 antigen in sera from patients with rheumatic disease represents a marker of in vivo vascular endothelial cell activity.

Anticoagulant effects of nonionic versus ionic contrast media in angiography syringes.

To determine whether nonionic contrast media present a clotting hazard when plastic or glass injection syringes are contaminated with aspirated blood, we evaluated two nonionic (iohexol and iopamidol) and two ionic (ioxaglate and diatrizoate) contrast agents. We used a blood:contrast media ratio of 2 mL:5 mL and ten normal donors, each studied at 10, 20, 30, and 60 minutes, a parallel study of clotting and fibrinopeptide A (FPA) generation in plastic tubes, and life table analysis to estimate more accurately donor-based early clotting probabilities. While ionic contrast media are stronger anticoagulants, both nonionic and ionic media retard clotting in plastic tubes, and clotting in plastic and glass angiography syringes in comparison to saline controls. A clotting probability of 1% for nonionic agents in plastic syringes was not reached until a time (mean +/- SD) of 21.5 +/- 3.2 minutes. This contrasts with a time of 8.7 +/- 2.5 minutes for saline control. With plastic syringes, no clotting at all was observed at 10 and 20 minutes with either class of agents. Neither class of agents hastened the generation of FPA. We found no evidence, therefore, that nonionic agents either cause clots or are procoagulant.

Detection of circulating endothelial antigen.

In order to determine whether endothelial cell antigens are detectable in serum from patients with rheumatic disease characterized by vascular involvement, we developed an ELISA using a monoclonal antibody directed against a 92,000 molecular weight endothelial cell specific antigen designated E92. Sera were assayed from 191 patients and 34 healthy controls. E92 was undetectable or present in very low concentrations in healthy controls and was elevated in most patients with an active rheumatic disease. Our results indicate that circulating endothelial antigens are present in rheumatic vascular syndromes and may provide a measure of endothelial cell function.

Fibrin and the vessel wall.

Fibrin is a major component of atherosclerotic plaques, and there may also be situations in which intravascular fibrin is formed in contact with the endothelium. The studies to be presented describe the distribution of fibrinogen/fibrin I, fibrin II, and fragments D and D-dimer in normal vessels and atherosclerotic plaques of increasing severity and also describe some functional effects of fibrin on normal endothelium. Immunohistochemical studies using three specific monoclonal antibodies with the avidin-biotin complex immunoperoxidase technique demonstrated that little fibrinogen/fibrin I or fibrin II and no D/D-dimer were detected in normal aortas. In early lesions and in fibrous plaques, fibrinogen/fibrin I and fibrin II were distributed in long threads and around vessel wall cells. D/D-dimer was not seen in early lesions. In advanced plaques all three molecular forms were detected in areas of loose connective tissue, in thrombi, and around cholesterol crystals. Thus increased fibrin formation and degradation may be associated with progression of atherosclerotic disease. Additionally, the presence of fibrin II around vessel wall cells suggests that these cells may be involved in the fbgn to fibrin transition within the vessel wall. The second aspect of the work to be presented concerns effects of fibrin on vascular endothelium. Fibrin formed on the surface of cultured human umbilical vein endothelial cells stimulated production of prostacyclin and tissue plasminogen activator by the cells in a time- and dose-dependent manner. Stimulation of prostacyclin was completely inhibited by indomethacin and partially inhibited by actinomycin D, cycloheximide, and trifluoperazine, while stimulation of t-PA synthesis was completely inhibited by actinomycin D and cycloheximide and partially inhibited by cytochalasin D, vinblastine, and trifluoperazine.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical characterization of fibrin(ogen)-related antigens in human tissues using monoclonal antibodies.

A new approach to study the distribution of fibrin(ogen)-related antigens was investigated using three different monoclonal antibodies (MAbs) and the avidin-biotin complex immunoperoxidase technique. MAb I8C6 recognizes B beta 1-42 peptide and can react with either fibrinogen or fibrin I; MAb T2G1 recognizes B beta 15-42 peptide and detects fibrin II but does not cross-react with fibrinogen; MAb GC4 reacts with Fragments D/DD derived from plasmin degradation of fibrinogen or fibrin but not with intact fibrinogen. The method can be applied to frozen or Bouin's fixed paraffin-embedded tissues obtained at biopsy, surgery, and autopsy. The distribution of the three antigens observed with the three MAbs was compared with that obtained with a polyclonal antiserum to fibrinogen and with the more conventional histochemical stains used in pathology to demonstrate fibrin deposits in tissues (Lendrum and PTAH). The staining observed with the three monoclonals clearly detected three different populations of fibrin(ogen)-related antigen in the tissues examined. The staining with MAb T2G1 specifically detected fibrin II with greater sensitivity than did conventional stains. The results of this study suggest that this method allows the molecular form of fibrin(ogen)-related deposits in tissues to be determined and this information may help to elucidate the role of fibrin in various disease states, such as atherosclerosis and renal disease, and in tumor growth and metastasis.

Effect of fibrin on endothelial cell production of prostacyclin and tissue plasminogen activator.

Fibrin formed on endothelial cells has previously been shown to have deleterious effects on the cells. Additionally, substances that cause endothelial cell damage have been reported to induce cultured endothelial cells to synthesize prostacyclin and tissue plasminogen activator (t-PA). The present studies were undertaken to determine whether fibrin formed on cultured human umbilical vein endothelial cells would alter synthesis of prostacyclin and t-PA by the cells. Fibrin was found to increase synthesis of both prostacyclin and t-PA in a dose and time dependent manner. Stimulation of prostacyclin synthesis was completely inhibited by indomethacin; partially inhibited by actinomycin D, cycloheximide, and trifluoperazine; and not affected by cytochalasin D or vinblastine. In contrast, stimulation of t-PA synthesis was completely inhibited by actinomycin D and cycloheximide; partially inhibited by cytochalasin D, vinblastine, and trifluoperazine; and not affected by indomethacin. Fibrin I, formed with Reptilase, caused only slight stimulation of t-PA production, but virtually no stimulation of prostacyclin synthesis. Neither collagen polymerization on the cells nor thrombin added in concentrations that did not induce fibrin polymer formation stimulated production of either substance. Furthermore, soluble fibrin II generated in the presence of the fibrin polymerization inhibitor gly-pro-arg-pro also failed to stimulate either prostacyclin or t-PA production. The presence of platelets in the plasma from which the fibrin was formed did not affect the amount of stimulation of the cells. Fibrin-induced stimulation of endothelial cell production of prostacyclin and t-PA could act to limit vascular occlusion in vivo by inhibiting platelet function and by stimulating fibrinolysis via t-PA.

Thrombosis in atherogenesis.

This review addresses the question of the involvement of fibrin in the development of atherosclerotic plaques. Numerous studies in the older literature demonstrated the presence of fibrinogen and/or fibrin in plaques, but the techniques that were available (mainly immunochemistry and immunohistochemistry with polyclonal antifibrinogen antibodies) did not clearly distinguish fibrinogen from fibrin or fibrinogen/fibrin degradation products. Some of these studies suggested that the fibrinogen-related protein within lesions resulted from incorporation of thrombi into lesions, while other studies suggested that fibrinogen itself entered the vessel wall. Newer studies by the authors and collaborators used specific antibodies for various fibrinopeptides to quantitate fibrinogen, fibrin I, fibrin II, and fragment X in thrombi and different histologic types of plaques. These studies showed that normal aortas contained fibrinogen and that fatty and fibrous plaques contained fibrinogen, fibrin I, and fibrin II, while complicated plaques contained fibrin II and fragment X, indicating a progression from fibrinogen to fibrin and fibrinogen/fibrin degradation products in parallel with increasing severity of the lesions. Later studies by the authors and collaborators used a sensitive immunohistochemical technique with monoclonal antibodies to demonstrate the distribution of fibrinogen-related antigens. Patterns suggesting incorporation of thrombi were seen, as were patterns suggesting formation of fibrin in association with arterial wall monocyte/macrophages and smooth muscle cells. The data from these various studies suggest the possibility that fibrin formation occurs within the arterial wall and contributes to plaque formation.

Identification and distribution of fibrinogen, fibrin, and fibrin(ogen) degradation products in atherosclerosis. Use of monoclonal antibodies.

Samples of normal and atherosclerotic vessels obtained from vascular and cardiothoracic surgery were examined for the distribution of fibrinogen/fibrin I, fibrin II, and fibrin(ogen) degradation products (Fragment D/DD) by using recently characterized monoclonal antibodies that recognize and distinguish the three molecular forms (MAbs 18C6, T2G1, and GC4, respectively) with the ABC-immunoperoxidase technique. In normal aortas, little fibrinogen/fibrin I or fibrin II was present and no fibrin(ogen) degradation products could be detected. In early lesions and in fibrous plaques, fibrinogen/fibrin I and fibrin II were distributed in long threads and surrounding vessel wall cells and macrophages. Fibrin(ogen) degradation products were not seen in early lesions. In fibrous and advanced plaques, fibrinogen/fibrin I, fibrin II, and fibrin(ogen) degradation products were detected in areas of loose connective tissue, in thrombus, and around cholesterol crystals. The results of this study suggest that increased fibrin formation and degradation may be associated with progression of atherosclerotic disease. The observed distribution of the different molecular forms of fibrinogen also suggests the possibility that the cells present in the lesions actively participate in the fibrinogen-to-fibrin transition within the vessel wall.

Immunochemical characterization of fibrinogen, fibrin I, and fibrin II in human thrombi and atherosclerotic lesions.

Arterial thrombi and atherosclerotic lesions were analyzed immunochemically and examined histologically. The extent of in vivo proteolytic cleavage of the amino-terminal end of fibrinogen by thrombin and plasmin was determined and quantitated by specific radioimmunoassays. The samples were treated with cyanogen bromide (CNBr), and the total amount of fibrinogen and fibrin-derived protein was determined as NDSK, the NH2-terminal disulfide knot of fibrinogen. Thrombin-releasable fibrinopeptides A and B were used to quantitate fibrinogen and fibrin I. Previous plasmin cleavage of the B beta chain was inferred from the amount of B beta 1-42 and B beta 15-42 in undigested NDSK. The results obtained in both acute and organized thrombi indicate that approximately 60% of the total protein (as determined by amino acid analysis) was fibrinogen-derived and that 70% to 80% of the fibrinogen-derived material was fibrin II. These findings support the hypothesis that fibrin II as distinct from fibrin I is the predominant component in a thrombus. In samples from normal and atherosclerotic aortas, fibrinogen-derived protein comprised less than 10% of the total protein. Samples from grossly normal aortas contained only fibrinogen and fibrin I. Fibrinogen concentration decreased and fibrin II concentration increased with increasing severity of the lesions, suggesting that increased fibrin II formation is associated with progression of atheromas.

Blood tests for the detection of thrombosis. Effects of blood flow and location of the sampling site.

Assays are available that allow the careful and wary investigator to use blood samples to derive useful information about hemostatic system activity. In practice the validity of the data will depend in very large part on the care which is taken in sample collection and processing. The particular system being studied profoundly influences the way in which the study should be performed. The details of the interacting issues are not yet resolved, and can only be dealt with as caveats. Finally and most importantly, these assays can not and should not be used to make the diagnosis of thrombosis. We believe that in general their use should be restricted to studies of pathophysiology. They are tools of exquisite sensitivity and specificity that allow us to probe the thrombotic process, and with care and imagination perhaps thrombogenesis itself.

Immunologic identification of the cleavage products from the A alpha- and B beta-chains in the early stages of plasmin digestion of fibrinogen.

Fragment X components (Mr 225,000 to 333,000) were distinguished on sodium dodecyl sulfate polyacrylamide gels. Western blotting with monoclonal antibodies to A alpha-chain segments demonstrated that the A alpha-chains of fibrinogen and the largest fragment X components (Mr 285,000-340,000) contained both A alpha 259-276 and A alpha 540-554. Fragment X components of Mr 270,000-285,000 contained A alpha 259-276 but lacked A alpha 540-554, whereas the smallest fragment X components (Mr 225,000-270,000) contained neither A alpha 540-554 nor A alpha 259-276. Studies of the small peptides generated during fragment X formation complemented the studies of the large molecules, by demonstrating peptides containing both A alpha 259-276 and A alpha 540-554 (Mr 41,600-41,800 and Mr 38,700-38,900), peptides containing A alpha 540-554 but not A alpha 259-276 (Mr 20,500-21,000 and Mr 17,300-17,500) and peptides containing only A alpha 259-276 (Mr 23,600-24,000 and Mr 20,500-21,000). Cleavage of B beta 1-42 from the amino terminal ends of the B beta-chains, measured with a specific radioimmunoassay, was linear until 1.6 moles per mole of fibrinogen had been released, and coincided with loss of the central and carboxy terminal A alpha-chain regions, i. e. A alpha 259-276 and A alpha 540-554. Based on present and previously reported data, a model is proposed for the evolution of the heterogeneous group of fragment X derivatives from fibrinogen with the simultaneous release of small peptides.(ABSTRACT TRUNCATED AT 250 WORDS)