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Biomedical subjects

E N Harris

Publications and source records attributed to E N Harris.

At least 73 records · Page 4Linked to original sources
PubMed1994

Antiphospholipid antibodies in an in vivo thrombosis model in mice.

The mouse model described in this study offers a unique method of determining the characteristics and mechanism(s) of action of aCL antibodies in thrombosis in vivo. In addition, this animal model enables the study of the kinetics of formation and dissolution of thrombus, as well as clot area, to be studied in a dynamic fashion. Other models for evaluation of thrombus formation rely on measurements of thrombus size and weight in ligated vessel segments where flow may be interrupted artificially. In addition, two important findings can be extracted from the study. (1) The size of the thrombi were significantly larger in mice that were passively immunized with IgG-APS (four patient samples examined) and with IgM-APS (two patient samples examined) compared with mice injected with saline or with immunoglobulin from control patients. (2) The clot persisted significantly for longer periods of time (total time) in animals injected with IgG-APS or IgM-APS when compared with control animals. Based on in vitro experiments, it is possible that these antibodies may inhibit protein C activation, neutralize the inhibitory activity action of beta 2 glycoprotein I (beta 2GPI), or activate platelets at the site of the femoral vein injury. Because this model enables to study the dynamics of thrombus formation, it is possible that these hypotheses and other mechanisms by which aPL antibodies are thrombogenic be investigated. Future studies will also include the effects of different levels of antibodies, as well as effects of affinity purified and monoclonal aPL antibodies on thrombus formation.

Animals↗
PubMed1994

Effect of human IgG antiphospholipid antibodies on an in vivo thrombosis model in mice.

High levels of IgG antiphospholipid antibodies (aPL) have been associated with clinical thrombosis. It is uncertain however whether these antibodies play a direct role in thrombosis or are merely epiphenomena. To investigate whether antiphospholipid antibodies might play a role in thrombosis, we utilized a novel mouse model in which the dynamics of in vivo thrombosis can be studied. CD1 mice (26-30 g) were passively immunized with 25 mg of human IgG from a patient with the Antiphospholipid Syndrome (IgG-APS) (n = 17), IgG from normal pooled sera (IgG-NHS) (n = 9), or saline solution (n = 12), followed by 40 mg of the same preparations at 48 h. At 72 h, levels of human aPL antibodies, detected using the anticardiolipin ELISA test (aCL ELISA test), in mice immunized with IgG-APS, were 50-100 GPL units. Each animal was anesthetized, femoral vein minimally mobilized and subjected to a standardized "pinch" injury to induce thrombosis. The vessel was transilluminated using acrylic optical fibers connected to a light source, and clot formation and dissolution were visualized by a standard surgical microscope equipped with a video camera, video recorder, and computer assisted analysis system. Results showed that average clot size was significantly larger in mice immunized with IgG-APS compared to those treated with saline (p < 0.037). In addition, the thrombus persisted longer in a significantly higher number of mice immunized with IgG-APS (10/17) compared to mice immunized with IgG-NHS (1/9) or saline (2/12) (p < 0.02).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

A comparison of cardiac valvular involvement in the primary antiphospholipid syndrome versus anticardiolipin-negative systemic lupus erythematosus.

Recurrent thrombosis and pregnancy loss are well-recognized features of the antiphospholipid syndrome. Uncertainty exists, however, as to whether other reported features of the antiphospholipid syndrome such as cardiac valvular vegetations are truly part of this disorder or more reflective of associated systemic lupus erythematosus (SLE). Several recent studies have concluded that patients with antiphospholipid antibodies have a higher risk of developing Libman-Sacks endocarditis. This study was performed to determine whether antiphospholipid antibodies are the only risk factors for cardiac valvular disease in patients with primary antiphospholipid syndrome (PAPS) or SLE. Ten patients with PAPS were matched with 20 patients with SLE and 20 healthy control subjects by sex. All participants were tested for anticardiolipin (aCL) antibodies by means of a standardized enzyme-linked immunosorbent assay technique, and all underwent two-dimensional and color-flow Doppler echocardiography. The echocardiograms were interpreted by two cardiologists blinded to the patients' underlying disease. Sixty percent of the PAPS group had cardiac valvular involvement compared with 40% of the SLE group (p = NS). We conclude that cardiac valvular vegetations are common both in aCL-negative patients with SLE and in patients with PAPS. This suggests that aCL antibodies either play no causative role or are not the only risk factors in the development of cardiac valvular vegetations.

Adult↗
PubMed1993

Are immunoglobulins with lupus anticoagulant activity specific for phospholipids?

Recent studies have suggested that the lupus anticoagulant (LA) may be specific for prothrombin, prothrombin-phospholipid complexes, or beta 2 glycoprotein 1 (beta 2GP1) rather than phospholipids. We performed a series of experiments to determine whether LA is indeed phospholipid specific. IgG was purified from sera of six patients with the antiphospholipid syndrome (APS) and 10 healthy controls. The six IgG-APS preparations had both LA and anticardiolipin (aCL) activity. Incubation of the six IgG-APS preparations with cardiolipin (CL), phosphatidylserine (PS), phosphatidylcholine (PC), or PS/PC (20:80) liposomes in Tris-buffered saline, resulted in loss of LA activity from the supernatant. We postulated that loss of activity might have resulted from absorption of IgG LA antibodies by phospholipids, a dilutional effect, or the presence of phospholipids in the supernatant causing 'by-pass' of IgG LA inhibitory activity. To distinguish between these possibilities, we re-isolated IgG from the supernatants and re-tested them for LA activity. IgG re-isolated from the PS. CL and PS/PC supernatants had no LA activity, but LA activity remained in the PC supernatant. This suggested that IgG with LA activity was absorbed by negatively charged but not by zwitterionic phospholipids. In like manner, PS, CL and PS/PC, but not PC liposomes, absorbed IgG with aCL activity. Mixtures of the phospholipid liposomes with beta 2GP1 did not modify the absorption of IgG with LA or aCL activity. Finally, we demonstrated that IgG eluted from immunoglobulin-cardiolipin liposome complexes had LA activity. Based on these findings, we conclude that at least one population of antibodies with LA activity is phospholipid specific.

Antibodies, Anticardiolipin↗
PubMed1993

Induction of phospholipid-binding antibodies in mice and rabbits by immunization with human beta 2 glycoprotein 1 or anticardiolipin antibodies alone.

Anticardiolipin (aCL) antibodies are autoantibodies present in high concentrations in patients with the antiphospholipid syndrome (APS), a disorder of recurrent thrombosis and pregnancy loss. What induces aCL antibodies is uncertain, but a recent report suggested that immunization of mice with beta 2 glycoprotein 1 (beta 2 GP1) in Freund's complete adjuvant (FCA) resulted in aCL antibody production in the recipient mice. Since this observation might explain how autoantibodies might be induced by poor immunogens, such as phospholipids, we decided to explore the question further. In our first series of experiments, we found that aCL antibodies were induced in mice by beta 2GP1 mixed with adjuvants that did not contain lipids (Adju-Prime or aluminium hydroxide). This excluded the possibility that antibody induction occurred because beta 2GP1 formed complexes with lipids in FCA. We also found that aCL antibodies always appeared before anti-beta 2GP1 antibodies, excluding the possibility that aCL antibodies were directed to beta 2GP1 or were induced by formation of anti-idiotypic antibodies (to anti-beta 2GP1). In experiments, we found that immunization of mice with human IgG antibodies from patients with the APS (IgG-APS), also induced aCL antibodies. Immunization with pure bovine serum albumin (BSA) did not induce aCL antibodies. We propose that aCL antibodies are induced by proteins with high avidity for phospholipids. These proteins may be bound to phospholipids when introduced, or may bind circulating phospholipids, so transforming phospholipid molecules into immunogens. Similar mechanisms might explain autoantibody induction to other poor immunogens.

Animals↗
PubMed1993

Do patients with antiphospholipid syndrome have autoantibodies to beta 2-glycoprotein I?

High positive anticardiolipin antibody tests have been associated with recurrent thrombosis and pregnancy loss. Although these antibodies were believed to bind negatively charged phospholipids, recent reports have suggested that a serum protein, beta 2-glycoprotein I (beta 2-GPI), may be the true antigen for these antibodies. To resolve this issue, we compared binding of 75 anticardiolipin-positive and 71 anticardiolipin-negative serum samples from patients with rheumatic diseases to beta 2-GPI by using an enzyme-linked immunosorbent assay (ELISA). Serum samples from 30 healthy blood donors and 10 laboratory personnel were used as normal controls. We found no difference in binding between the three groups of serum samples. In addition, when binding to beta 2-GPI coated plates was compared with binding to ELISA plates without beta 2-GPI (blank), no difference was observed. Finally, binding of anticardiolipin-positive serum samples to plates coated with cardiolipin-beta 2-GPI mixture varied directly with the cardiolipin concentrations. Based on these findings, we conclude that anticardiolipin-positive serum samples do not bind beta 2-GPI.

Antiphospholipid Syndrome↗
PubMed1992

Beta 2-glycoprotein 1 (beta 2GP1) enhances cardiolipin binding activity but is not the antigen for antiphospholipid antibodies.

Some investigators have reported that a serum protein, beta 2-glycoprotein 1 (beta 2GP1), either alone or in combination with negatively charged phospholipid, may be the antigen for anticardiolipin (aCL) antibodies. To examine these reports further, ELISA tests, inhibition experiments, Ouchterlony and Western blot techniques were used to examine anticardiolipin binding to beta 2GP1. Sera from patients with the antiphospholipid syndrome (APS) and syphilis were studied, as well as whole IgG immunoglobulin and affinity purified (a.p.) IgG aCL antibodies. Results showed no binding of aCL antibodies to beta 2GP1 in the absence of cardiolipin. beta 2GP1 caused enhanced binding of aCL antibodies to cardiolipin, but this enhancement was not observed in inhibition experiments. Binding to cardiolipin occurred in the absence of beta 2GP1. Enhancement of cardiolipin binding activity by beta 2GP1 was observed for APS, but not for syphilis. We conclude that beta 2GP1 is not the antigen for aCL antibodies, nor is it likely that the antibody recognizes shared beta 2GP1-cardiolipin epitopes. Instead, this protein may make cardiolipin more available for aCL binding on solid surfaces by some yet undefined mechanism. This effect may not extend to aqueous suspensions.

Antibodies, Anticardiolipin↗
PubMed1992

Antiphospholipid antibodies in homozygous sickle cell disease.

Serum samples from 108 unselected Jamaican patients with homozygous sickle cell disease and 116 control subjects with normal haemoglobin were screened for the presence of antiphospholipid antibodies. Slightly increased levels of IgG antiphospholipid antibodies were found in nine patients with sickle cell disease and in none of the control subjects. Serial control samples confirmed the increased levels of antiphospholipid antibodies. A comparison of the haematological and clinical features of patients with positive and negative antiphospholipid sickle cell disease did not highlight any differences between the groups.

Adolescent↗
PubMed1992

The antiphospholipid syndrome: the clot thickens.

The "Antiphospholipid Syndrome" is a disorder that mainly affects young people. Recurrent venous or arterial thrombosis, recurrent fetal loss, and thrombocytopenia are the primary features. The diagnosis is established by the presence of the anticardiolipin (aCL) antibody or the lupus anticoagulant (LA). Treatment is symptomatic.

Autoantibodies↗
PubMed1992

Serological detection of antiphospholipid antibodies.

Antiphospholipid antibodies are heterogeneous. A subset of these antibodies is associated with arterial and venous thrombosis and recurrent pregnancy loss. This subset is best diagnosed by the lupus anticoagulant and solid-phase anticardiolipin tests. Solid-phase anticardiolipin tests use an enzyme-linked immunosorbent assay employing cardiolipin or other negatively charged phospholipids as the antigen. Patients with antiphospholipid antibodies who have experienced thrombotic disorders often have an IgG isotype, persistent at high levels for prolonged periods. The anticardiolipin test has been standardized so that antibody isotype and level can be determined reproducibly. Standardized serological tests to measure antiphospholipid antibodies will enable uniform identification of patients at risk for thrombosis and will help elucidate the origin and mechanisms of action of these antibodies.

Antibodies↗
PubMed1991

Should anticardiolipin tests be performed in otherwise healthy pregnant women?

Positive anticardiolipin and lupus anticoagulant tests are not confined to patients with the antiphospholipid syndrome, and the usefulness of these tests in healthy pregnant women is uncertain. This study sought to determine the prevalence of anticardiolipin antibodies and correlation with pregnancy outcome in 1449 pregnant women. Results were compared with 40 patients with the antiphospholipid syndrome. Persistence of positive anticardiolipin antibody tests was also ascertained. Twenty-six of 1449 sera (1.79%) were immunoglobulin G anticardiolipin positive, and 63 (4.3%) were immunoglobulin M anticardiolipin positive. Twenty-three of 26 positive for immunoglobulin G anticardiolipin and 55 of 63 positive for immunoglobulin M anticardiolipin results were low. Anticardiolipin positivity did not correlate with complications or outcome. Immunoglobulin G isotype and level distinguished patients with antiphospholipid syndrome from otherwise healthy women with positive anticardiolipin tests. In healthy pregnant women positive anticardiolipin tests occur infrequently, at low levels, and are rarely associated with adverse pregnancy outcome. This test should be requested only when the antiphospholipid syndrome is suspected.

Adult↗
PubMed1990

Comparison of test methods for the lupus anticoagulant: international survey on lupus anticoagulants-I (ISLA-1).

Six lyophilized plasma samples were sent to 20 "expert" laboratories for assessment of lupus anticoagulant (LA). Four samples contained pooled LA of graded potency mixed with aged normal plasma. One contained LA plus cephalin phospholipid and one contained a nonspecific venom anticoagulant. Sixteen methods were used overall with some participants using up to 8 methods. Results were scored in regard to the known potencies of LA in the samples and other known induced defects. Activated partial thromboplastin time (APTT) tests used by most participants for preliminary screening were relatively sensitive, but non-specific. Platelet or phospholipid neutralization procedures (PNP) appeared to be sensitive and specific but showed a non-linear response to increased LA content. Kaolin clotting time (KCT) tests showed the most sensitive response to increased LA content but the weaker LA were not scored as abnormal by most laboratories as the samples may have contained platelet fragments. Other commonly used tests such as the tissue thromboplastin inhibition (TTI) test and the dilute Russell's viper venom test (DRVVT) were carried out somewhat inconsistently. The variability in performance of tests in different laboratories indicates that standardization of methodology is urgently required. Generally it seemed that most clotting tests were "bypassed" by the addition of phospholipid to a known LA-positive sample in apparently direct proportion to their sensitivity. Sample preparation, especially prevention of contamination with activated platelets is a vital preliminary part in the assay of LA.

Autoantibodies↗