RESUMEN
BACKGROUND: Surprisingly, IgE cross-reactivity between the major peanut allergens Ara h 1, 2, and 3 has been reported despite very low sequence identities. OBJECTIVE: We investigated the unexpected cross-reactivity between peanut major allergens. METHODS: Cross-contamination of purified natural Ara h 1, 2, 3, and 6 was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blot test, liquid chromatography-tandem mass spectrometry (LC-MS/MS), and sandwich enzyme-linked immunosorbent assay (ELISA). IgE cross-reactivity was studied with sera of peanut-allergic patients (n = 43) by ELISA and ImmunoCAP inhibition using both intact natural and recombinant allergens and synthetic peptides representing postulated Ara h 1 and Ara h 2 cross-reactive epitopes. RESULTS: Both purified nAra h 1 and nAra h 3 were demonstrated to contain small but significant amounts of Ara h 2 and Ara h 6 (<1%) by sandwich ELISA, SDS-PAGE/Western blot analysis, and LC-MS/MS. IgE cross-inhibition between both 2S albumins and Ara h 1 and Ara h 3 was only observed when using natural purified allergens, not recombinant allergens or synthetic peptides. Apparent cross-reactivity was lost when purified nAra h 1 was pretreated under reducing conditions, suggesting that Ara h 2 and Ara h 6 contaminations may be covalently bound to Ara h 1 via disulfide interactions. CONCLUSION: True cross-reactivity of both peanut 2S albumins with Ara h 1 and Ara h 3 could not be demonstrated. Instead, cross-contamination with small quantities was shown to be sufficient to cause significant cross-inhibition that can be misinterpreted as molecular cross-reactivity. Diagnostic tests using purified nAra h 1 and nAra h 3 can overestimate their importance as major allergens as a result of the presence of contaminating 2S albumins, making recombinant Ara h 1 and Ara h 3 a preferred alternative.
Asunto(s)
Alérgenos , Hipersensibilidad al Cacahuete , Humanos , Alérgenos/química , Proteínas de Plantas/química , Arachis , Antígenos de Plantas/metabolismo , Cromatografía Liquida , Inmunoglobulina E , Espectrometría de Masas en Tándem , Albuminas 2S de Plantas , Péptidos/metabolismo , Albúminas/metabolismo , Hipersensibilidad al Cacahuete/diagnósticoRESUMEN
BACKGROUND AND AIMS: With the intention to gain support for the hypothesis that incident ischemic complications of atherosclerotic disease involve a stochastic aspect, we performed a histological, qualitative evaluation of the epidemiology of coronary atherosclerotic disease in a cohort of aortic valve donors. PATIENTS AND METHODS: Donors (n = 695, median age 54, range 11-65 years) were dichotomized into a non-cardiovascular (non-CVD) and a cardiovascular disease death (CVD) group. Consecutive 5 mm proximal left coronary artery segments were Movat stained, and the atherosclerotic burden for each segment was graded (revised AHA-classification). RESULTS: Non-CVD and CVD groups showed steep increase of atherosclerosis severity beyond the age of 40, resulting in an endemic presence of advanced atherosclerosis in men over 40 and women over 50 years. In fact, only 19% of the non-CVD and 6% of the CVD donors over 40 years were classified with a normal LCA or a so called non-progressive lesion type. Fibrous calcified plaques (FCP), the consolidated remnants of earlier ruptured lesions, dominated in both non-CVD and CVD donors. Estimates of the atherosclerosis burden (i.e. average lesion grade, proportion of FCPs, and average number of FCPs per cross-section) were all higher in the CVD group (p<1.10-16, p<0.0001, and p<0.05, respectively). CONCLUSIONS: Dominance of consolidated FCP lesions in males over 40 and females over 50 years, show that plaque ruptures in the left coronary artery are common. However, the majority of these ruptures remain asymptomatic. This implies that the atherosclerotic process is repetitive. A relative difference in disease burden between CVD and non-CVD donors supports the concept that complications of atherosclerotic disease involve a stochastic element.