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It has been reported that the level of anticardiolipin antibodies (aCL) is probably genetically determined, as HLA-DR7 (or HLA-DR4) has been shown to be associated with persistently high levels of aCL.1 No reported data are available about genetic factors implicated in ischaemic or thrombotic events in patients with systemic lupus erythematosus (SLE) without antiphospholipid antibodies (aPL).
In this study we examined the frequency of the platelet GPIIb/IIIa-P1A2 polymorphism in a series of patients with SLE. We assessed the relationship between the A2 allele prevalence2 and the clinical and immunological manifestations, as possible predisposing biological factors to major ischaemic manifestations such as central nervous system (CNS) lupus or Raynaud’s phenomenon as defined according to the American College of Rheumatology (ACR) Committee.3
We studied a cohort of 109 patients with SLE attending the rheumatology referral centres of the universities of Udine, Pisa, and Milan, classified according to the ACR criteria. For comparison, 161 patients with rheumatoid arthritis (systemic inflammatory disease), 54 with systemic sclerosis (chronic ischaemic-thrombotic vasculopathy), and 128 healthy blood donors (HBD) recruited from the blood transfusion service were studied. The clinical and laboratory manifestations were carefully examined to detect a possible association between genotypes and symptoms, signs, and immunological characteristics. In particular, we focused on nephritis, vasculitis, Raynaud’s phenomenon, and CNS major neurological events, thought to be related to possible ischaemic-thrombotic mechanism; these were defined by the presence of one of the following: (a) acute cerebrovascular disease (stroke); (b) seizures or chorea; (c) lupus headache; (d) organic brain syndrome. Anticardiolipin antibodies (enzyme linked immunosorbent assay (ELISA) assay: normal values <15 GPLU; β2-glycoprotein I dependent) and or lupus anticoagulant (LAC) positivities were defined according to the most recent international consensus statement.4 In the study aPL were considered positive when aCL or LAC were positive on two consecutive occasions over a six week period. Antibodies to DNA were tested either on Crithidia or in an ELISA assay (normal values <30 IU/ml). When one or the other was positive antibodies to DNA were considered positive in this study. The analysis of the P1A2 gene polymorphism was performed after DNA extraction following the restriction fragment length polymorphism method as reported by Weiss et al.2
Statistical analysis was performed using the Prism software (Graph-Pad, San Diego, CA 92121, USA), which automatically calculated the odds ratios (OR) and confidence intervals (CI) with the statistical significance.
Patients with SLE had a mean (SD) age of 41.6 (14.6), mean (SD) disease duration 8.9 (7.0) years; 34 (31%) had nephritis, 21 (19%) major CNS events, 20 (18%) arthritis, 41 (38%) Raynaud’s phenomenon (Ray), 105 (96%) antinuclear antibodies, 64 (59%) antibodies to dsDNA, and 56 (51%) aPL (aCL or LAC, or both, positive). Table 1 shows that the frequency of the A2 allele tended to be slightly increased in patients with SLE compared with HBD (OR=1.30, 95% CI 0.7 to 2.0), whereas in patients with systemic sclerosis the opposite trend was seen (OR=0.7). The same trend was seen for the A2 homozygosity in SLE (OR=2.4, 95% CI 0.2 to 26.6, p=NS) as well as in rheumatoid arthritis (OR=4.1, 95% CI 0.5 to 35.3, p=NS). The A2 allele was found to be more represented in the aPL positive than in the aPL negative patients (OR=1.7, 95% CI 0.7 to 3.9, p=NS) and in Ray+ compared with Ray− patients (OR=1.7, 95% CI 0.7 to 3.9, p=NS). CNS events were far more often seen in aPL positive than negative patients as expected (OR=2.9, 95% CI 1.0 to 8.1, p=0.04). In addition, subjects carrying the A2 allele in association with the aPL positivity have an increased risk of developing Raynaud’s phenomenon (OR=2.8, 95% CI 1.0 to 7.5, p=0.04), whereas no association was found with CNS events, nephritis, cutaneous vasculitis (table 2). Antiphospholipid antibody positivity did not correlate with Raynaud’s phenomenon.
In SLE some patients in whom CNS ischaemic events occur, have no increases of aPL, and other factors should be considered as pathogenic. To clarify this point we considered the issue of possible genetic risks and we focused on the genetic features of the P1A2 allele polymorphism, which has been suggested as a predisposing factor either for myocardial or for CNS events.5 In this study we examined the prevalence of the A2 allele and its relationship with clinical manifestations. The P1A2 genetic setting (C to T replacement at nucleotide 1565) in platelet glycoprotein (GP)IIb/IIIa (integrin αIIbβ3) has been commonly reported as a possible risk factor for CNS and coronary ischaemic events, especially in younger patients.
We observed a trend towards an increased frequency of the A1/A2-A2/A2 genotypes in patients with SLE with Raynaud’s phenomenon and mostly in those with aPL. Activated GPIIb/IIIa receptor mediates platelet aggregation and stable adhesion through the interaction with von Willebrand factor and fibrinogen. When codified by the P1A2 allele, this integrin has been shown to bind more tightly to immobilised fibrinogen and to enhance platelet reactivity.6 Therefore the A2 allele might predispose to vascular damage and in association with aPL to a vasospastic phenomenon. Our data appear complementary to a previous report from the Baltimore group,7 in which the frequency of the P1A1 and P1A2 alleles was compared in aPL positive patients with and without thrombosis. In the subgroup with arterial thrombotic events 33% were homozygous or heterozygous for the P1A2 allele, whereas only 19% of the patients without thrombosis possessed the allele. The major conclusion was that although platelet GPIIIa polymorphism was not a major risk factor for all thrombosis in patients with aPL, a possible association with arterial thrombosis could be considered.
Further studies dealing with factors released from platelets of carriers of the A2 allele should be performed to define fully the functional role of the polymorphism in diseases like SLE. Our data suggest that the P1A2 allele defines a subset of patients with SLE with the following characteristics: higher risk of aPL positivity and presence of Raynaud’s phenomenon.