Variants of the FADS1 FADS2 Gene Cluster, Blood Levels of Polyunsaturated Fatty Acids and Eczema in Children within the First 2 Years of Life

Association of genetic-variants in the FADS1-FADS2-gene-cluster with fatty-acid-composition in blood of adult-populations is well established. We analyze this genetic-association in two children-cohort-studies. In addition, the association between variants in the FADS-gene-cluster and blood-fatty-acid-composition with eczema was studied. Data of two population-based-birth-cohorts in The Netherlands and Germany (KOALA, LISA) were pooled (n = 879) and analyzed by (logistic) regression regarding the mutual influence of single-nucleotide-polymorphisms (SNPs) in the FADS-gene-cluster (rs174545, rs174546, rs174556, rs174561, rs3834458), on polyunsaturated fatty acids (PUFA) in blood and parent-reported eczema until the age of 2 years. All SNPs were highly significantly associated with all PUFAs except for alpha-linolenic-acid and eicosapentaenoic-acid, also after correction for multiple-testing. All tested SNPs showed associations with eczema in the LISA-study, but not in the KOALA-study. None of the PUFAs was significantly associated with eczema neither in the pooled nor in the analyses stratified by study-cohort. PUFA-composition in young children's blood is under strong control of the FADS-gene-cluster. Inconsistent results were found for a link between these genetic-variants with eczema. PUFA in blood was not associated with eczema. Thus the hypothesis of an inflammatory-link between PUFA and eczema by the metabolic-pathway of LC-PUFAs as precursors for inflammatory prostaglandins and leukotrienes could not be confirmed by these data

Development of a hypoallergenic recombinant parvalbumin for first-in-man subcutaneous immunotherapy of fish allergy.

To access publisher's full text version of this article, please click on the hyperlink in Additional Links field or click on the hyperlink at the top of the page marked Files. This article is open access.The FAST (food allergy-specific immunotherapy) project aims at developing safe and effective subcutaneous immunotherapy for fish allergy, using recombinant hypoallergenic carp parvalbumin, Cyp c 1.Preclinical characterization and good manufacturing practice (GMP) production of mutant Cyp (mCyp) c 1.Escherichia coli-produced mCyp c 1 was purified using standard chromatographic techniques. Physicochemical properties were investigated by gel electrophoresis, size exclusion chromatography, circular dichroism spectroscopy, reverse-phase high-performance liquid chromatography and mass spectrometry. Allergenicity was assessed by ImmunoCAP inhibition and basophil histamine release assay, immunogenicity by immunization of laboratory animals and stimulation of patients' peripheral blood mononuclear cells (PBMCs). Reference molecules were purified wild-type Cyp c 1 (natural and/or recombinant). GMP-compliant alum-adsorbed mCyp c 1 was tested for acute toxicity in mice and rabbits and for repeated-dose toxicity in mice. Accelerated and real-time protocols were used to evaluate stability of mCyp c 1 as drug substance and drug product.Purified mCyp c 1 behaves as a folded and stable molecule. Using sera of 26 double-blind placebo-controlled food-challenge-proven fish-allergic patients, reduction in allergenic activity ranged from 10- to 5,000-fold (1,000-fold on average), but with retained immunogenicity (immunization in mice/rabbits) and potency to stimulate human PBMCs. Toxicity studies revealed no toxic effects and real-time stability studies on the Al(OH)3-adsorbed drug product demonstrated at least 20 months of stability.The GMP drug product developed for treatment of fish allergy has the characteristics targeted for in FAST: i.e. hypoallergenicity with retained immunogenicity. These results have warranted first-in-man immunotherapy studies to evaluate the safety of this innovative vaccine.info:eu-repo/grantAgreement/EC/FP7/20187

The lancet weight determines wheal diameter in response to skin prick testing with histamine

BACKGROUND:Skin prick test (SPT) is a common test for diagnosing immunoglobulin E-mediated allergies. In clinical routine, technicalities, human errors or patient-related biases, occasionally results in suboptimal diagnosis of sensitization. OBJECTIVE:Although not previously assessed qualitatively, lancet weight is hypothesized to be important when performing SPT to minimize the frequency of false positives, false negatives, and unwanted discomfort. METHODS:Accurate weight-controlled SPT was performed on the volar forearms and backs of 20 healthy subjects. Four predetermined lancet weights were applied (25 g, 85 g, 135 g and 265 g) using two positive control histamine solutions (1 mg/mL and 10 mg/mL) and one negative control (saline). A total of 400 SPTs were conducted. The outcome parameters were: wheal size, neurogenic inflammation (measured by superficial blood perfusion), frequency of bleeding, and the lancet provoked pain response. RESULTS:The mean wheal diameter increased significantly as higher weights were applied to the SPT lancet, e.g. from 3.2 ± 0.28 mm at 25 g to 5.4 ± 1.7 mm at 265 g (p<0.01). Similarly, the frequency of bleeding, the provoked pain, and the neurogenic inflammatory response increased significantly. At 265 g saline evoked two wheal responses (/160 pricks) below 3 mm. CONCLUSION AND CLINICAL RELEVANCE:The applied weight of the lancet during the SPT-procedure is an important factor. Higher lancet weights precipitate significantly larger wheal reactions with potential diagnostic implications. This warrants additional research of the optimal lancet weight in relation to SPT-guidelines to improve the specificity and sensitivity of the procedure

Cross-reactivity of IgE antibodies to allergens

The cross-reactivity of IgE antibodies is of interest for various reasons, three of which are discussed. Firstly, from the clinical view, it is important to know the patterns of cross-reactivity, because they often (but not always) reflect the pattern of clinical sensitivities. We discuss the cross-reactivities associated with sensitization to pollen and vegetable foods: PR-10 (Bet v 1-related), profilin, the cross-reactive carbohydrate determinant (CCD), the recently described isoflavone reductase, and the (still elusive) mugwort allergen that is associated with celery anaphylaxis; cross-reactivities between allergens from invertebrates, particularly tropomyosin, paramyosin, and glutathione S-transferase (GST); and latex-associated cross-reactivities. Clustering cross-reactive allergens may simplify diagnostic procedures and therapeutic regimens. Secondly, IgE cross-reactivity is of interest for its immunologic basis, particularly in relation to the regulation of allergic sensitization: are IgE antibodies to allergens more often cross-reactive than IgG antibodies to "normal" antigens? If so, why? For this discussion, it is relevant to compare not only the structural relation between the two allergens in question, but also the relatedness to the human equivalent (if any) and how the latter influences the immune repertoire. Thirdly, prediction of IgE cross-reactivity is of interest in relation to allergic reactivity to novel foods. Cross-reactivity is a property defined by individual antibodies to individual allergens. Quantitative information (including relative affinity) is required on cross-reactivity in the allergic population and with specific allergens (rather than with whole extracts). Such information is still scarce, but with the increasing availability of purified (usually recombinant) allergens, such quantitative information will soon start to accumulate. It is expected that similarity in short stretches of the linear amino-acid sequence is unlikely to result in relevant cross-reactivity between two proteins unless there is similarity in the protein fol

The effect of neuromuscular blockade on canine laparoscopic ovariectomy: A double-blinded, prospective clinical trial

The Effect of Neuromuscular Blockade on Canine Laparoscopic Ovariectomy: A Double-Blinded, Prospective Clinical Trial Bart Van Goethem, Diplomate ECVS, Sebastiaan Alexander van Nimwegen, PhD, Ies Akkerdaas, DVM, Joanna Claire Murrell, BVSc., PhD, Diplomate ECVAA, and Jolle Kirpensteijn, PhD, Diplomate ACVS & ECVS Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 108, 3584CM, Utrecht, The Netherlands Corresponding Author: Bart Van Goethem, Diplomate ECVS, Department of Small Animal Medicine and Clinical Biology, Faculty of Veterinary Medicine, Gent University, Gent, Belgium E-mail: [email protected] Submitted July 2010 Accepted November 2011 DOI:10.1111/j.1532-950X.2011.00962.x Objective: To evaluate the effect of neuromuscular blockade (NMB) on surgical time and various anesthetic variables during laparoscopic ovariectomy in dogs. Study Design: Prospective, double-blinded, randomized clinical trial. Animals: Female dogs (n = 40). Methods: Laparoscopic ovariectomy by bipolar electrocoagulation was performed by 1 surgeon using a standardized protocol, where 1 ovary was removed under NMB, and the other without NMB. Surgical and anesthetic (respiratory and circulatory) variables were recorded for predetermined procedural stages and were statistically evaluated. Results: Mean total surgical time was 25.1 ± 6.3 minutes (range, 16–47 minutes). With NMB, mean duration of surgical excision of the ovary (5.7 ± 2.3 minutes) was not significantly changed compared to ovariectomy without NMB (5.9 ± 1.9 minutes). Arterial blood pressure was the only recorded anesthetic variable that significantly changed under NMB (5% decrease). Occurrence of intraoperative complications did not differ. In obese dogs, total surgical time was increased by 22%. Other variables, including occurrence of intraoperative mesovarial bleeding did not influence surgical duration. Conclusions: NMB did not significantly improve laparoscopic ovariectomy times and except for a 5% decrease in arterial blood pressure did not change any of the evaluated anesthetic and surgical variables

The effect of neuromuscular blockade on canine laparoscopic ovariectomy: A double-blinded, prospective clinical trial

The Effect of Neuromuscular Blockade on Canine Laparoscopic Ovariectomy: A Double-Blinded, Prospective Clinical Trial Bart Van Goethem, Diplomate ECVS, Sebastiaan Alexander van Nimwegen, PhD, Ies Akkerdaas, DVM, Joanna Claire Murrell, BVSc., PhD, Diplomate ECVAA, and Jolle Kirpensteijn, PhD, Diplomate ACVS & ECVS Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 108, 3584CM, Utrecht, The Netherlands Corresponding Author: Bart Van Goethem, Diplomate ECVS, Department of Small Animal Medicine and Clinical Biology, Faculty of Veterinary Medicine, Gent University, Gent, Belgium E-mail: [email protected] Submitted July 2010 Accepted November 2011 DOI:10.1111/j.1532-950X.2011.00962.x Objective: To evaluate the effect of neuromuscular blockade (NMB) on surgical time and various anesthetic variables during laparoscopic ovariectomy in dogs. Study Design: Prospective, double-blinded, randomized clinical trial. Animals: Female dogs (n = 40). Methods: Laparoscopic ovariectomy by bipolar electrocoagulation was performed by 1 surgeon using a standardized protocol, where 1 ovary was removed under NMB, and the other without NMB. Surgical and anesthetic (respiratory and circulatory) variables were recorded for predetermined procedural stages and were statistically evaluated. Results: Mean total surgical time was 25.1 ± 6.3 minutes (range, 16–47 minutes). With NMB, mean duration of surgical excision of the ovary (5.7 ± 2.3 minutes) was not significantly changed compared to ovariectomy without NMB (5.9 ± 1.9 minutes). Arterial blood pressure was the only recorded anesthetic variable that significantly changed under NMB (5% decrease). Occurrence of intraoperative complications did not differ. In obese dogs, total surgical time was increased by 22%. Other variables, including occurrence of intraoperative mesovarial bleeding did not influence surgical duration. Conclusions: NMB did not significantly improve laparoscopic ovariectomy times and except for a 5% decrease in arterial blood pressure did not change any of the evaluated anesthetic and surgical variables

Severe immediate allergic reactions to grapes: Part of a lipid transfer protein-associated clinical syndrome

Background: Grape allergy is considered rare; grape lipid transfer protein (LTP; Vit v 1), an endochitinase and a thaumatin-like protein (TLP) have been reported as grape allergens. A considerable number of patients have referred to our department for severe reactions to grapes, and several IgE binding proteins were detected. Objectives: The aim of this study was to identify and characterise the allergens involved in severe allergic reactions to grapes and describe the population in which they occur. Methods: Patients with reported severe allergic reactions to grapes (n = 37) are described. Grape allergens were purified/fractionated by a combination of chromatographic techniques, identified by proteomic analysis and biochemically characterised. Immunoreactivity was assessed by blot (inhibitions) and RAST (inhibitions), and skin prick tests were performed with the isolated allergens. Results: All subjects were polyallergic, sensitised and reactive to several additional foods and pollen. All patients were sensitised to grape LTP. A 28-kDa expansin, a 37.5-kDa polygalacturonase-inhibiting protein, a 39-kDa β-1,3-glucanase and a 60-kDa protein were identified as minor grape allergens. Endochitinase and TLP did not play a role. Inhibition experiments revealed the possible cross-reactive role of LTP for clinical sensitivities to other LTP-containing plant foods, but also the involvement of cross-reactive carbohydrate determinants of minor allergens in IgE cross-reactivity. Conclusions: LTP is the major grape allergen, while additional minor allergens may contribute to clinical reactivity. Severe grape allergy presents in atopic patients who frequently react to other LTP-containing, plant-derived foods. The &apos;LTP syndrome&apos; is the appropriate term to describe this condition. Copyright © 2007 S. Karger AG

IgE cross-inhibition between Ara h 1 and Ara h 2 is explained by complex formation of both major peanut allergens

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