Levels Of Anti-endothelial Cell Antibodies Research Articles

P4494Renal insufficiency mitigates the protective effects of elevated anti endothelial cell antibodies on survival in patients undergoing coronary angiography

Abstract Background We previously demonstrated that elevated levels of antiendothelial cell antibodies are associated with improved survival of patients undergoing coronary angiography. However, renal insufficiency, which is associated with decreased survival, leads to increased level of the antiendothelial cell antibodies. The purpose of this study was to evaluate the correlation of the levels of these antibodies with survival in patients with normal versus reduced renal function. Patients and methods This was a single center prospective study. Eight hundred thirty eight consecutive patients undergoing coronary angiography with detectable antibody levels were enrolled. The levels of antiendothelial antibodies were determined by ELISA and measured in optical density units. Renal insufficiency was defined as adjusted GFR<60 ml/min. The mean follow up was 30 months. Results Of total 838 patients, 481 (57%) had normal and 357 (42.6%) had reduced renal function. Total mortality was 13.6%, being significantly higher in patients with reduced (23.2%) versus normal (6.4%) renal function (p<0.0001). Antiendothelial cell antibody levels were higher in patients with normal versus reduced renal function (4.76±7.05 versus 3.84±6.09 OD units, p=0.042). Antiendothelial cell antibody levels were significantly higher in survived versus deceased patients (4.56±6.82 versus 3.14±5.50 OD units, p=0.014). This association with survival was even more pronounced in patients with normal renal function (4.98±7.19 versus 11.69±3.29 OD units, p<0.0001). In patients with reduced renal function, there was no significant difference between survived and deceased patients (3.89±6.12 versus 3.68±6.05 OD units, p=0.788). Conclusions Renal insufficiency leads to mild decrease in the antiendothelial cell antibodies' levels. Higher levels of antiendothelial cell antibodies are associated with improved survival in patients undergoing coronary angiography. This association is significant in patients with normal but not reduced renal function. Thus, the effect of the higher levels of these antibodies on survival cannot be explained by its association with renal dysfunction.

Pattern of antiendothelial cell antibodies in patients with chronic obstructive pulmonary disease

BackgroundAutoimmune mechanisms have been recently recognized as being partly involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Circulating autoantibodies have been detected in patients with COPD.ObjectivesThe aim of this study was to estimate the level of antiendothelial cell antibodies (AECA) in COPD patients in the various GOLD stages.Patients and methodsA comparative study assessed the level of AECA in three groups. COPD patients without cor pulmonale (18), with cor pulmonale (12), and the control group (10), from Beni-Suef University Hospital. Each group underwent background questionnaires and BMI measures. Enzyme-linked immunosorbent assay was used to measure the level of AECA in serum (ng/ml). Right ventricular systolic pressure (mm Hg) was measured in COPD patients with cor pulmonale.ResultsThe studied groups consisted of male patients (age: 50–80 years). BMI was significantly lower (20.36) in the COPD group with cor pulmonale. Spirometry forced expiratory volume and forced expiratory volume/forced vital capacity ratios showed significantly lower levels among the COPD patients without cor pulmonale. AECA showed a lower level in the control group (26.25) compared with the COPD without cor pulmonale (57.87) and much lowered when compared to cor pulmonale group (71.47). The level of AECD was higher in the patients in third (74.78) and fourth stages (79.10) compared with those in the second stage (58.75).ConclusionThere is a much higher level of AECA in COPD patients with cor pulmonale and in advanced stages in comparison with patients without cor pulmonale and at early stages, and a significant positive correlation was found between AECA levels and right ventricular systolic pressure.

Serum anti-endothelial cell antibodies in patients with age-related macular degeneration treated with intravitreal bevacizumab.

To analyse the prevalence and changes in circulating anti-endothelial cell antibodies (AECA) during anti-vascular endothelial growth factor (anti-VEGF) therapy. Ninety-eight patients with exudative age-related macular degeneration (AMD) were treated with intravitreal bevacizumab. Fifty sex- and age-matched healthy subjects were used as controls. Serum AECA were detected using indirect immunofluorescence on primate skeletal muscle and cultivated human umbilical vein endothelial cells (HUVEC). These investigations were repeated at 4-week intervals within 8months of follow-up. At baseline examination, 30 of the 98 patients (30.6%) were positive for AECA. The titres of AECA ranged from 1:10 to 1:320. In the control group, AECA were present in only nine sera (18%) with titres ranging between 1:20 and 1:80 (p=0.0000). The greatest rates of reduction of AECA titres were observed during the 'loading' phase of therapy. During the 'maintenance' phase, the rates of changes in serum AECA levels were less significant and remained constant. In follow-up period in 13 patients (13.3%), serum AECA were detected de novo in titres of 1:10 to 1:80. Statistical analysis did not show any significant correlation between the presence of AECA and activity of the disease. There is growing evidence that AMD is an immune-mediated disease, and thus it cannot be excluded that AECA may be involved in its pathogenesis and progression. We also speculate that AECA develop in response to retinal damage and anti-VEGF therapy.

Association of Nail Dystrophy With Accrued Damage and Capillaroscopic Abnormalities in Systemic Lupus Erythematosus.

There are some nail abnormalities described in systemic lupus erythematosus (SLE). The aim of this study was to evaluate the association between nail dystrophy (ND) and disease activity, accrued organ damage, capillaroscopic abnormalities, autoantibodies, and some markers of endothelial cell activation in patients with SLE. This was a cross-sectional study of SLE patients from a rheumatology clinic in a tertiary care hospital. Patients were allocated in groups, according to the presence or absence of ND. Demographics, clinical data, disease activity, accrued damage, serology, nailfold capillaroscopy characteristics, serum levels of anti-endothelial cell antibodies, and plasma levels of endothelin 1 were compared between groups. Disease activity was assessed by the Systemic Lupus Erythematosus Disease Activity Index 2000 index and accrued organ damage by the Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index. Sixty-one patients were included; 50 patients (82%) were female. Thirty-two patients (52.5%) showed ND, and 29 did not. Besides a more frequent use of cyclophosphamide (46.9% vs 20.7%; P = 0.03) in the ND group, clinical features were similar. A greater organ damage was found in patients with ND (median Systemic Lupus International Collaborating Clinics/American College of Rheumatology index = 0.5, minimum = 0, maximum = 6) than in patients without ND (0, 0, 3, respectively; P = 0.04); specifically, only the skin domain was associated with ND (P = 0.04). Onycholysis (40.6%) and longitudinal ridging (25%) were the most frequent nail changes. Nailfold capillaroscopy changes were more frequent in ND patients (40.6%) than in control subjects (13.8%) (P = 0.02). The most frequent nailfold capillaroscopy findings in the ND group were enlarged capillaries (40.6%) and microhemorrhages (12.5%). There was no association between ND and the autoantibody profile, plasma endothelin 1, or serum anti-endothelial cell antibodies. Nail dystrophy was associated with higher accrued organ damage and the presence of capillaroscopic abnormalities. This suggests that ND might be related to chronic microvascular damage.

Endothelial antibody levels in the sera of children with autism spectrum disorders

BackgroundThe neurobiological basis of autism remains poorly understood. We hypothesized that endothelial antibodies may be associated with the pathophysiology of autism and may predict intellectual/social developmental abnormalities. MethodsPlasma levels of antiendothelial cell antibodies (AECAs) were measured by enzyme-linked immunosorbent assay in autistic children (n = 55) and age-matched healthy controls (n = 25). ResultsThe serum level of AECAs in children with autism {n = 55, 306.4 ± 45.6 pg/mL [mean ± standard error of the mean (SEM)]} was higher (two-tailed Student t test: p = 0.05) than that of healthy controls [n = 25, 209.6 ± 24.6 pg/mL (mean ± SEM)]. Children with severe autism exhibited significantly higher AECAs than healthy controls (diagnoses of autistic children based on the Childhood Autism Rating Scale score, >40) [n = 20, 369.6 ± 65.6 pg/mL (mean ± SEM)] (p = 0.03). Disease severity and the Childhood Autism Rating Scale score, which represent stereotyped patterns of behavior in children with autism, were positively correlated (r2 = 0.27, p = 0.05). ConclusionElevated AECA serum levels may be implicated in the pathogenesis of autism. However, these data should be interpreted with caution until further investigations are performed using larger sample sizes to determine whether the increase in serum AECA levels is a mere consequence of autism or it plays a pathogenic role in the disease.

Etiopathogenesis, clinical diagnosis and treatment of thromboangiitis obliterans - current practices.

Thromboangiitis obliterans (TAO) is a segmental inflammatory occlusive disorder that affects small- and medium-sized arteries, and arm and leg veins of young smokers. Several different diagnostic criteria have been offered for the diagnosis of TAO. Clinically, it manifests as migratory thrombophlebitis or signs of arterial insufficiency in the extremities. It is characterized by highly cellular and inflammatory occlusive thrombi, primarily of the distal extremities. Thromboses are often occlusive and sometimes display moderate, nonspecific inflammatory infiltrate, consisting mostly of polymorphonuclear leukocytes, mononuclear cells and rare multinuclear giant cells. The immune system appears to play a critical role in the etiology of TAO. However, knowledge about immunological aspects involved in the progression of vascular tissue inflammation, and consequently, the evolution of this disease, is still limited. There are several studies that suggest the involvement of genetic factors and results have shown increasing levels of antiendothelial cell antibodies in patients with active disease. Vasodilation is impaired in patients with TAO. TAO disorder may actually be an autoimmune disorder, probably initiated by an unknown antigen in the vascular endothelium, possibly a component of nicotine. There are various therapies available for treatment of TAO, but the major and indispensable measure is smoking cessation. Except for discontinuation of tobacco use, no forms of therapy are definitive. Sympathectomy, cilostazol and prostaglandin analogues (prostacyclin or prostaglandin E) have been used in specific conditions. Recently, therapeutic angiogenesis with autologous transplantation of bone marrow mononuclear cells has been studied in patients with critical limb ischemia.

Presence of antibodies against endothelial cells in the sera of patients with episodic angioedema and hypereosinophilia

We reported three additional cases of a newly described syndrome called episodic angioedema with hypereosinophilia. In order to investigate its pathophysiological mechanisms, four parameters were concurrently investigated, including blood eosinophil density, serum chemoattractant activity, serum major basic protein (MBP) levels and the presence of anti-endothelial cell antibodies. Distribution of eosinophils through a metrizamide density gradient showed a preferential sedimentation of blood eosinophils in intermediate layers, clearly different from the hypodense cells (low-density layers) identified in a group of seven patients with idiopathic hypereosinophilic syndrome (HES). In two of the three patients with cyclic angioedema, a chemotactic activity towards eosinophils was detected in the serum (30 +/- 6 and 42 +/- 12 eosinophils per high-power field; P less than 0.05 compared with a control group). Serum MBP levels were at 1524, 619 and 1200 pg/ml. All three patients had circulating anti-endothelial cell antibodies, predominantly of the IgG isotype, in contrast to controls (P less than 0.01) or to patients with HES (P less than 0.01). Specificity of the antibody for endothelial cells was demonstrated in the three patients studied by the absence of binding to various blood cells, including monocytes, lymphocytes, eosinophils and platelets. In one case (patient 2), the levels of anti-endothelial cell antibodies, as well as the serum chemoattractant activity to eosinophils varied according to the successive acute phases of the disease. Although further investigations are needed to clarify the exact pathophysiology of this syndrome, and especially the possible participation of the anti-endothelial cell antibodies in the cutaneous lesions, these data suggest that angioedema observed in this syndrome could result from the combined effects of activated eosinophils and of immunologically induced endothelial lesions.

Soluble intercellular adhesion molecule-1 (sICAM-1) as a marker of disease relapse in idiopathic uveoretinitis.

This study reports the results of a point prevalence study of markers of endothelial dysfunction in the serum of patients with idiopathic uveoretinitis. sICAM-1, soluble endothelial leucocyte adhesion molecule (sELAM), anti-endothelial cell antibodies (AECA) and von Willebrand factor (vWF) levels were measured in 32 patients with isolated idiopathic uveoretinitis and seven with uveitis in association with systemic disease, using commercial and in-house ELISAs. Raised levels of AECA were found in 31% of patients with isolated uveitis, vWF in 28%, sELAM in 15.6% and sICAM-1 in 31%. Further analysis revealed that raised sICAM-1 levels were closely associated with recent relapse of disease (P = 0.00003). Patients with accompanying systemic disease were found to have a similar prevalence of these serum abnormalities to those with isolated ocular disease. In conclusion, vascular endothelial dysfunction may contribute to pathogenesis in uveoretinitis, and in particular sICAM-1 may prove a marker of disease relapse in this condition.

Antibodies to endothelial cells in systemic lupus erythematosus: a potential marker for nephritis and vasculitis

Using an ELISA, anti-endothelial cell antibodies (AECA) have been found in sera obtained at the time of renal biopsy in 46 out of 57 patients (81%) with systemic lupus erythematosus (SLE) and nephritis (mean binding index (BI) = 84% +/- 52.8) compared with 22 out of 50 SLE patients (44%) without nephritis (mean BI = 45% +/- 35.9). Seventy normal human sera had a mean BI of 10% +/- 9.8. The highest levels were seen in patients with diffuse proliferative glomerulonephritis (WHO grade IV) and in patients with proteinuria and nephrotic syndrome. When the biopsies were assessed for activity and chronicity scores, AECA were associated with active renal lesions (P less than 0.001). AECA levels correlated with low complement levels but not with anti-DNA antibodies to extractable nuclear antigens (ENA), anti-cardiolipin or anti-neutrophil cytoplasmic antibodies. The presence of AECA conferred a positive predictive value of 0.68 for the presence of nephritis. Twenty-five patients had active vasculitis at the time of assay and the highest AECA values were seen in patients with both nephritis and vasculitis. No correlation was seen with serum immunoglobulin levels and immune complexes did not bind significantly to the endothelial surface. The possible role of these antibodies as a marker in lupus nephritis is discussed.

Anti-Endothelial Cell Antibodies in Patients With Sarcoidosis

Anti-endothelial cell antibodies (AECA) are circulating antibodies that bind to endothelial antigens and induce endothelial cell damage. These antibodies have been detected in patients with collagen vascular disease and systemic vasculitis. Sarcoidosis is a multiple granulomatous disorder of unknown etiology, and its clinical presentation, organ involvement, and prognosis are highly diverse. In this study, we aimed to investigate the expression of AECA in patients with sarcoidosis. We also examined whether these antibodies could serve as a marker for the activity, severity, and prognosis of sarcoidosis. Forty sarcoidosis patients, whose diagnosis was established by clinicoradiologic findings and histologic confirmation of noncaseating epithelioid cell granulomas, were studied. Serum and BAL samples were examined for AECA by cellular enzyme-linked immunosorbent assay (ELISA) using human umbilical vein endothelial cells. The findings were expressed in terms of an ELISA ratio (ER). Fifty-seven subjects without any clinical, radiologic, or serologic evidence of pulmonary disease or autoimmune disorders served as the reference population to judge the positivity of AECA. Patients with sarcoidosis had a significantly higher positivity rate for and levels of AECA in both serum and BAL fluid than the reference population. In addition, the ER of AECA was significantly elevated in patients with multiple lesions or who required corticosteroid therapy compared with that in patients without multiple lesions or who did not need corticosteroid therapy. Expression of AECA might be a useful marker to predict the course of sarcoidosis.

Antiendothelial cell antibody levels in patients with masked hypertension

Recent evidence demonstrate that masked hypertension (MH) is a significant predictor of cardiovascular disease, while, elevated levels of circulating antibodies against endothelial cell surface antigens (antiendothelial cell antibodies — AECA) seem to play an important role at the early stages of atherosclerosis process and of borderline hypertension as well. Aim of this study was to investigate the presence of AECA in patients (pts) with MH and to compare the AECA title among pts with MH and healthy normotensives (HN), matched for age, sex and body mass index. Methods One hundred-thirty (60 M, 70 F) healthy subjects mean age 45 ± 12 yrs who had clinic blood pressure < 140/90 mm Hg were studied. The whole study population underwent 24 hour ambulatory blood pressure monitoring (ABPM). According to the ABPM recordings, 24 individuals (8 M, 16 F) had MH (daytime systolic blood pressure ≥ 135 mm Hg or daytime diastolic blood pressure ≥ 85 mm Hg — group A) and the remainder 106 subjects (52 M, 54 F) had normal ABPM recordings, group B. IgG and IgM AECA levels were determined by ELISA method. AECA levels were expressed as mean value ± SD. None of the study population had a history of connective tissue disease or any metabolic disorder. Results Significantly increased titles of AECA class IgG were found in 8/24 pts of group A (30%) vs. 5/106 (4.6%) of group B ( p < 0,001). Significantly increased titles of AECA class IgM were also found in 6/24 pts of group A (25%) vs. 3/80 (3.8%) of group B ( p < 0,001). Conclusions Our results suggest that patients with MH have significantly higher AECA levels of both classes (IgG, IgM) compared to healthy normotensives. These findings may indicate a possible explanation of the increased cardiovascular risk in MH. The possibility that high AECA levels may be a driving mechanism for the development of MH needs further investigation.

Induction of Endothelial Cell Apoptosis by IgG Antibodies from SLE Patients with Nephropathy

Systemic lupus erythematosus (SLE) is a prototype of an autoimmune disease with vasculopathy as demonstrated by the presence of vascular immune-complex deposition, inflammation, and thrombosis. A pivotal role in the initiation of vasculopathy is ascribed to vascular endothelium. In this respect, anti-endothelial cell antibodies (AECA), which are highly associated with SLE, are putative candidates for the initiation of SLE vasculopathy. In addition to the potency of AECA to induce a proinflammatory endothelial cell phenotype, AECA have also been described to trigger endothelial cell apoptosis. However, in SLE data are not uniform on the potentials of AECA to induce endothelial cell apoptosis in vitro. We have addressed this question in a cohort of SLE patients with nephropathy. AECA levels, and the apoptosis-inducing potentials of serum IgG were measured at the time of renal complication and biopsy. Also serum antibody reactivity with various SLE-related autoantigens including HSP60 was determined in patients. The results show that the SLE patient group has increased AECA levels as well as increased levels of induction of endothelial cell apoptosis by serum IgG. AECA and apoptosis values largely varied among the patients. Our data show that antibodies other than anti-HSP60 are also involved in apoptosis induction. The results are discussed in the context of recent findings on the role of AECA in endothelial cell apoptosis and renal vasculopathy in SLE.

Anti-endothelial cell antibodies are prevalent in juvenile idiopathic arthritis: implications for clinical disease course and pathogenesis

To determine the prevalence of anti-endothelial cell antibodies (AECA) in children with juvenile idiopathic arthritis (JIA) versus healthy control children. Twenty-eight children with active JIA were studied (ten each with polyarticular and oligoarticular disease, and eight with systemic onset disease). AECA were determined by a cell-based ELISA from samples obtained every 3 months over a 2 year period in each subject. These levels were compared against previously determined levels of von Willebrand factor antigen, fibrin d-dimer, and soluble forms of ICAM-1 and E-selectin, as well as clinical measures of disease activity. AECA were detected in 5/10 oligoarticular, 6/10 polyarticular, and 7/8 systemic JIA subjects and 0/14 controls. Mean levels of AECA were significantly higher in subjects with oligoarticular, and especially systemic disease as compared to polyarticular and control groups when analyzed by ANOVA. AECA are prevalent in JIA and are present more often and at higher levels in systemic disease.

Circulating IgA from acute stage of childhood Henoch-Schönlein purpura can enhance endothelial interleukin (IL)-8 production through MEK/ERK signalling pathway

Recently, sera from children with active Henoch-Schönlein purpura (HSP) have been found to enhance interleukin (IL)-8 production by human umbilical venous endothelial cells (HUVEC). To further determine the possible factor with the ability to enhance endothelial IL-8 production in sera from acute stage of HSP, 10 children with HSP at the acute stage and 10 healthy controls were enrolled. IgA antiendothelial cell antibodies (AECA) were detected by cell-based ELISA. Active sera with or without pretreatment with anti-human IgA antibody, sera of controls, and immunoglobulin A (IgA) derived from sera were used to stimulate the HUVEC. The ability of these factors to enhance endothelial IL-8 production was evaluated. Furthermore, signalling pathways were also assayed by different inhibitors, and confirmed by immunoblotting. Serum levels of IgA AECA in HPS patients at the acute stage were significantly higher than in controls (P < 0.001). The active sera could enhance endothelial IL-8 production (P = 0.004, compared with control sera), and the ability of these sera was mostly abolished when pretreated with fixed anti-human IgA antibody. The supernatant IL-8 levels of endothelial cells stimulated by IgA derived from acute stage of HSP were statistically higher than controls (P < 0.001). PD98059, an inhibitor of ERK phosphorylation, significantly reduced IgA AECA-stimulated endothelial IL-8. IgA AECA also enhanced the phosphorylation of ERK1 with a time-dependent manner. Together with these findings, it is concluded that IgA AECA derived from acute stage of HSP may bind to endothelial and enhance endothelial cells to produce IL-8 via MEK/REK signalling pathway.

Thromboangiitis obliterans (Buergerʼs disease)

Thromboangiitis obliterans is a nonatherosclerotic segmental inflammatory disease that affects the small and medium-sized arteries and veins in the upper and lower extremities. This review will help to familiarize physicians with this vasculitis that is completely different from every other type of vasculitis encountered. While tobacco is central to the initiation and continuance of Buerger's disease activity, two interesting pathophysiologic observations have been made. There is endothelial dysfunction in arteries not yet clinically or angiographically involved in thromboangiitis obliterans. There are elevated levels of anti-endothelial cell antibodies and measurement of these antibody titers may be useful in following disease activity in Buerger's disease patients. The only therapy clearly shown to prevent amputation is the complete abstinence of tobacco. There is exciting work under way on the use of selective cannabinoid receptor antagonists to help patients stop smoking. In addition, preliminary results on use of therapeutic angiogenesis in patients with Buerger's disease has demonstrated excellent collateral blood vessels formation and clinical improvement. The difficulty in studying rare diseases such as thromboangiitis obliterans is that there are no significant research dollars available and even the most active centers only see a few patients per year. Therefore, there has been little progress in understanding the pathogenesis of the disease. There are new therapeutic modalities that help patients with this disease, however, and patients can be assured that if they are able to discontinue tobacco use completely, amputation will not occur if critical limb ischemia is not already present.

Antiendothelial Cell Antibody Levels in Healthy Normotensives with High Normal Blood Pressure

High normal blood pressure (HNBP) seems to be related to an increased cardiovascular risk in healthy normotensive subjects. According to the literature, elevated levels of antibodies against endothelial cell surface antigen antiendothelial cell antibodies (AECA) play an important role in the early stages of atherosclerosis process and in borderline hypertension. The aim of this study was to compare AECA levels of healthy normotensives with HNBP to those of healthy normotensives with normal blood pressure (NBP), matched for age, sex, and body mass index (BMI).Methods. Ninety healthy normotensives with HNBP (43M, 47F; mean age, 48 ± 2.6 yrs; BMI 23.6 ± 1.5 Kg/m2) (Group A) and 80 healthy normotensives with NBP (41M, 39F; mean age, 46 ± 3 yrs; BMI 24 ± 1.8 Kg/m2) (Group B) were studied. Both group subjects were matched for sex, age, and BMI. AECA levels were determined in each subject using an enzyme-linked immunosorbent assay (ELISA). AECA levels were expressed as mean values. Results. Twenty-five subjects from group A (28%) showed elevated IgG antiendothelial cell antibodies levels vs. three from group B (3.75%, p < 0.001). IgM AECA levels were elevated in 18 subjects from group A (20%) vs. two from group B (1.5%, p < 0.001). Conclusions. The present findings suggest that healthy normotensives with HNBP have significantly higher AECA levels of both classes (IgG, IgM) compared to healthy normotensives with NBP. This may have prognostic significance for the future development of essential hypertension in this group of healthy subjects.

DETECTION OF ANTI-ENDOTHELIAL CELL ANTIBODIES IN PATIENTS AFTER HEART TRANSPLANTATION AND THEIR RELATION TO ENDOTHELIAL CELL ACTIVATION.

A284 Aims: Transplant-associated coronary artery disease is the most serious long-term complication after cardiac transplantation (TxCAD). Although often associated with chronic production of anti-endothelial cell antibodies (AECA), the precise role of antibodies in chronic rejection remains uncertain. In the present study we have screened serum of patient after heart transplantation for the presence of AECA and discussed their activation potential on endothelial cells. Methods: Up to 6 years after transplantation, 175 sequential sera were retrospectively collected from 37 patients. Serum antibody activity was measured using a cellular enzyme-linked immunosorbent assay (ELISA) in which human endothelial cells (EaHy926) were used as target cells. The effect of antibodies on human saphenous vein endothelial cells (HSVEC) and adhesion molecule expression was measured by cellular ELISA. Results Long-term transplantation time resulted in a significant increase in the antibody activity (1-2 years: 33 % vs 60 % for IgA; 3-6 years: 13 vs 40 % for IgM). There was no correlation between AECA and the number of acute rejection episodes. Only two patients showed increased levels of AECA (IgG-type) without any signs of TxCAD. In the early postoperative phase (up to 3 months) serum concentrations of interleukin-6 and tumor necrosis factor-soluble receptor I were elevated and correlated with the amount of IgG. Even 3 years after transplantation the level of IgG-AECA remained elevated in one patient (factor 3 level above probands). Incubation of HSVEC with 30 % of the serum for 48 h resulted in an increase of the proliferation activity (20-40 %). The basal expression of adhesion molecules was not influenced, whereas the tumor necrosis factor alpha induced expression of E-selectin increased by a factor of 1.8. To exclude the influence of soluble factors in serum (medication) it will be necessary to purify the antibodies. Conclusions: The presence of AECA in the serum of patients after transplantation may be a prognostic marker for endothelial injury. It remains to be seen whether antibodies of IgA and IgM-type cause cell activation.

Anti-Endothelial Cell Antibodies (AECA) in Systemic Sclerosis - Increased Sensitivity using Different Endothelial Cell Substrates and Association with Other Autoantibodies

Objective: One of the main features of systemic sclerosis (SSc) is vascular damage, the mechanism of which is not understood. In the present study we examined whether screening of SSc patients for different anti-endothelial cells antibodies (AECA) of various origins increase the sensitivity of AECA detection in SSc patients. Secondary aim was an attempt to correlate AECA with other common autoantibodies. Materials & methods: 478 SSc patients were studied for the presence AECA. anti-cardiolipin (aCL), anti-dsDNA, anti-heparin (AHA), anti-pyruvate dehydrogenase (PDH) and anti-PDC-E2 autoantibodies. AECA levels were determined using human umbilical vein EC (HUVEC), bone marrow EC (BMEC), EC hybridoma (EA.hy 926) and Kaposi sarcoma EC (KS). Results: Positive AECA were found in 49.5% of SSc patients (27.1% HUVEC: 34.3% BMEC; 26.3% EaHy 926 and 22.7% KS). The highest percent reactivity of AECA was obtained using microvascular BMEC. When combining BMEC and either other cell lines the reactivity ranged from 41.4% to 46%. A significant association between AECA on the one hand and AHA (p<0.001) and anti-PDH (p<0.05) on the other was seen. Cross-reactivity with anti-PDC-E2 was excluded by inhibition tests, but AHA and anti-PDH may be part of the spectrum of AECA. Conclusions: Since false-negative AECA may result from lack of expression of various antigens on a specific EC, analysis of AECA in SSc patients requires using several EC types, including microvascular EC.

Identification of candidate endothelial cell autoantigens in systemic lupus erythematosus using a molecular cloning strategy: a role for ribosomal P protein P0 as an endothelial cell autoantigen.

To attempt to characterize the diversity and nature of antigens recognized by anti-endothelial cell antibodies (AECA) in patients with systemic lupus erythematosus (SLE) using a molecular cloning strategy. AECA in sera of 15 SLE patients were measured by ELISA and Western blot analysis was used to examine the diversity of autoantigen targets in two clinically active patients. A human umbilical vein endothelial cell cDNA expression library was immunoscreened with sera from these two patients to identify their autoantigen targets. An anti-ribosomal P peptide antibody ELISA was used to assess the clinical significance of anti-ribosomal P protein antibodies in the sera of one patient. Significantly higher AECA levels were found in five patients with active disease and nephritis than in five patients with clinically inactive disease. Sera from two clinically active patients were found to recognize distinct spectra of autoantigens. The candidate autoantigens that were identified included (1) endothelial cell-specific plasminogen activator inhibitor; (2) the classical lupus antigen, i.e. ribosomal P protein P0; and (3) proteins never before described as putative autoantigens in SLE, including ribosomal protein L6, elongation factor 1alpha, adenyl cyclase-associated protein, DNA replication licensing factor, profilin II and the novel proteins HEAPLA 1 and HEAPLA 2 (human endothelial associated putative lupus autoantigens 1 and 2). In one patient, antibodies against ribosomal P protein P0 were predominant and levels of these antibodies correlated with total AECA levels, anti-DNA antibody titres, overall clinical score and renal disease in a longitudinal study. A panel of candidate endothelial autoantigens in SLE, which includes previously described autoantigens and novel targets, has been identified by a molecular cloning strategy. This novel molecular approach could also be applied to the identification of autoantigens in other autoimmune vascular diseases.

Anti-endothelial cell antibodies in patients with coronary atherosclerosis

Anti-endothelial cell antibodies (AECA) have been shown to possess endothelial cell activation properties and to harbor pathogenic potential in experimental animal models of autoimmune systemic disorders. Atherosclerosis is a form of an inflammatory condition in which the immune system has been shown to be involved. The aim of the present study was to assess the presence of AECA in patients with coronary atherosclerosis. A total of 134 patients admitted for chest pain of suspected anginal origin were evaluated for coronary artery atherosclerosis by angiography. Sera were drawn prior to the procedure for the determination of AECA employing cyto-ELISA. AECA positive sera were further evaluated for its ability to promote in vitro E-selectin expression by HUVEC using a cell-based ELISA. Patients with no coronary artery involvement had levels of AECA that did not differ from those obtained for patients with confirmed coronary atherosclerosis (one, two or three vessel disease). Furthermore, AECA positive sera from patients, with or without coronary atherosclerosis displayed similar capacity of inducing E-selectin expression by endothelial cells. AECA may not stand as an optimal mean of discriminating atherosclerotic from non-atherosclerotic patients. The ability of AECA to activate endothelial cells is also not unique to patients with atherosclerosis and is evident also in age-matched control subjects.