46 research outputs found
The importance of cross-reactivity in grass pollen allergy
According to the data obtained from in vivo and in vitro testing in Serbia, a significant number of patients have allergic symptoms caused by grass pollen. We examined the protein composition of grass pollens (Dactylis glomerata, Lolium perenne and Phleum pratense) and cross-reactivity in patients allergic to grass pollen from our region. The grass pollen allergen extract was characterized by SDS-PAGE, while cross-reactivity of single grass pollens was revealed by immunoblot analysis. A high degree of cross-reactivity was demonstrated for all three single pollens in the sera of allergic patients compared to the grass pollen extract mixture. Confirmation of the existence of cross-reactivity between different antigenic sources facilitates the use of monovalent vaccines, which are easier to standardize and at the same time prevent further sensitization of patients and reduces adverse reactions
Igg vezivanje alergena i alergoida polena pelina prethodno izloženih simuliranim uslovima gastrointestinalnog trakta mereno domaÄim elisa testom
This study considers the influence of exposure to simulated gastrointestinal conditions (saliva, gut, intestine and acidic conditions of the gut) on IgG binding of unmodified allergens and three types of LMW allergoids of Artemisia vulgaris pollen extract obtained by means of potassium cyanate succinic and maleic anhydride. It also concerns the optimization of a self-developed ELISA assay for comparison of the specific IgG binding of mugwort pollen extract and modified mugwort pollen derivatives. The ELISA was conducted with a mugwort pollen extract coupled to the plate, using the sera from 12 mugwort- pollen allergic patients. The exposure to saliva fluid for 2 min did not influence the IgG binding properties of allergens and allergoids. Exposure of mugwort pollen allergens and LMW allergoids to the acidic conditions of the gut did not dramatically change their IgG binding properties. By exposing mugwort pollen extract and LMW derivatives to the SGF conditions for 1 h, the percent of IgG binding epitopes was reduced to a half of its starting value in the extract and to about 30%in all the allergoid samples. After prolonged exposure only the carbamyl derivative showed reduced IgG binding. Changes of the IgG binding potential of all four samples after exposure in SIF followed a similar pattern.Predmet ovog rada je ispitivanje promena IgG vezivanja nemodifikovanih alergena polena A. vulgaris i tri tipa alergoida malih molekulskih masa dobijenih tretmanom sa kalijum-cijanatom, i anhidridima Äilibarne i meleinske posle izlaganja uslovima gastrointestinalnog trakta (saliva, želudac, tanko crevo i kisela sredina želudaÄnog soka) i optimizacija domaÄeg ELISA testa za odreÄivanje IgG vezivanja alergena i alergoida polena pelina.UELISA testu je za ploÄicu kuplovan ekstrakt polena pelina i koriÅ”Äeni su serumi 12 pacijenata alergiÄnih na pelin. Izlaganje salivi u trajanju od 2 minuta ne utiÄe na IgG vezujuÄe osobine alergena i alergoida. Izlaganje kiseloj sredini želudaÄnog soka znaÄajno ne utiÄe na IgG vezujuÄe osobine alergena i alergoida polena pelina. Posle izlaganja simuliranom želudaÄnom soku alergena i derivata u trajanju od 1 sata, procenat IgG vezujuÄih epitopa u nemodifikovanom uzorku se smanjuje na polovinu poÄetne vrednosti i na oko 30 % kod sva tri derivata. Jedino se kod karbamil-derivata % IgG vezivanja dodatno smanjuje sa produžavanjem izlaganja SGF-u.Promene u IgG vezujuÄem potencijalu sva 4 uzorka posle izlaganja simuliranim uslovima tankog creva prate sliÄan obrazac
Allergenic potency of kiwi fruit during fruit development
Food allergies, including kiwi fruit allergy, have been the subject of extensive research in the last few years. The aim of this study was to examine a possible relationship between the developmental stage of kiwi fruit and its allergenic potency. The protein and allergen patterns of kiwi fruit extracts in September, October, November and December fruit in the period from 2000-2002 were analysed. One of the factors that may contribute to the difficulties in proposing well-defined and standardized fruit extracts should also be the time of fruit harvesting. In this particular case, when the kiwi fruit was edible throughout November and December, we showed discrepancies in allergen content and potencies both in qualitative and quantitative terms. Two major allergens of kiwi fruit, Act c 1 and Act c 2, mainly accounted for the highest allergenic potential of November kiwi extract in vivo and in vitro. Not only the content of major allergens, but also the ratio of different proteins and even isoforms of the same allergen (Act c 2) change with fruit ripening. These findings should be taken into account during preparation of extracts for allergy diagnosis
Izolovanje i karakterizacija 68 kD alergena iz ekstrakta kuÄnih grinja
House dust mites (HDM) represent a major source of allergens, contributing to the increasing incidence of type I hypersensitivity disease worldwide. Over 30 different IgE-binding proteins from the HDM extract were detected. Although group 1 and 2 have been identified as major allergens, due to the safety and efficacy of allergy diagnosis and immunotherapy, there is a need to carefully evaluate the clinical relevance of other allergens present in the HDM extract. In regard to this, a high molecular mass allergen of about 68 kD was purified from the HDM extract using a combination of gel permeation chromatography and reversed-phase chromatography. The IgG and IgE reactivity of the purified protein were preserved during the purification process, as confirmed by Western blot analysis with polyclonal rabbit antibodies and dot blot analysis with a pool of sera from subjects with house dust mite allergy, respectively. In addition, the IgE reactivity was confirmed using ELISA testing with nine patient sera. The biological potency of the 68 kD allergen was confirmed by skin prick testing in five allergic subjects, suggesting that the high molecular mass allergen is a good candidate for component-resolved diagnosis of house dust mite allergy and eventual therapeutic treatment.Grinje iz kuÄne praÅ”ine predstavljaju jedan od glavnih izvora alergena koji su u znaÄajnoj meri doprineli porastu prvog tipa preosetljivosti. Preko 30 IgE-vezujuÄih proteina iz kuÄne praÅ”ine je detektovano do danas. Alergeni grupe 1 i 2 oznaÄeni su kao glavni alergeni kuÄne praÅ”ine. MeÄutim, da bi se poboljÅ”ala sigurnost i efikasnost dijagnoze i terapije alergijskih oboljenja izazvanih grinjama iz kuÄne praÅ”ine, neophodno je odrediti kliniÄki znaÄaj svih alergena iz ovog alergenskog izvora. U ovom radu izolovan je alergen visoke molekulske mase od 68 kD iz ekstrakta kuÄne praÅ”ine kombinovanjem gel-permeacione hromatografije i reversno-fazne hromatografije. IgG i IgE reaktivnost preÄiÅ”Äenog proteina je proverena u 'Western blot'-u i 'dot blot'-u sa poliklonskim zeÄijim antitelima na ekstrakt kuÄne praÅ”ine i 'pool'-om seruma osoba alergiÄnih na kuÄnu praÅ”inu, redom. 64 % pacijenata je pokazalo IgE reaktivnost na preÄiÅ”Äeni protein u ELISA testu. BioloÅ”ka reaktivnost preÄiÅ”Äenog alergena je potvrÄena u kožnim probama na pet pacijenata, ukazujuÄi da je preÄiÅ”Äen alergen dobar kandidat za dijagnozu alergije na kuÄnu praÅ”inu pojedinaÄnim komponentama i eventualni terapeutski tretman
Digestibilnost alergoida polena pelina u simuliranim uslovima gastrointestinalnog trakta
Chemically modified allergens (allergoids) have found use in both traditional and novel forms of immunotherapy of allergic disorders. Novel forms of immunotherapy include local allergen delivery, via the gastrointestinal tract. This study conveys the gastrointestinal stability of three types of mugwort pollen allergoids under simulated conditions of the gut. Allergoids of the pollen extract of Artemisia vulgaris were obtained by means of potassium cyanate, succinic and maleic anhydride. Gastrointestinal tract conditions (saliva, and gastric fluid) were simulated in accordance with the EU Pharmacopoeia. The biochemical and immunochemical properties of the derivatives following exposure to different conditions were monitored by determining the number of residual amino groups with 2,4,6-trinitrobenzenesulfonic acid, SDS PAGE, immunoblotting and inhibition of mugwort-specific IgE. Exposure to saliva fluid for 2 min did not influence the biochemical and immunochemical properties of the derivatives. In the very acidic conditions of the simulated gastric fluid, the degree of demaleylation and desuccinylation, even after 4 h exposure, was low, ranging from 10 to 30 %. The digestion patterns with pepsin proceeded rapidly in both the unmodified and modified samples. In all four cases, a highly resistant IgE-binding protein the Mwof which was about 28-35 kD, was present. Within the physiological conditions, no new IgE binding epitopes were revealed, as demonstrated by immunoblot and CAP inhibition of the mugwort specific IgE binding. An important conclusion of this study is the stability of the modified derivatives in the gastrointestinal tract of patients, within physiological conditions. The means that they are suitable for use in much higher concentrations in local forms of immunotherapy than unmodified ones.U ovom radu su prikazani rezultati ispitivanja stabilnosti tri tipa alergoida polena pelina u simuliranom želudaÄnom soku. KoristeÄi kalijum-cijanat anhidrid Äilibarne i anhidrid maleinske kiseline, napravljeni su alergoidi polena pelina (Artemisia vulgaris). Saliva i želudaÄni sok su simulirani na osnovu evropske farmakopeje. Biohemijske i imunohemijske osobine derivata posle izlaganja razliÄitim uslovima, praÄene su: odreÄivanjem broja slobodnih amino grupa u reakciji sa TNBS, SDS PAG elektroforezom, imunoblotom i odreÄivanjem pelin-specifiÄnog imunoglobulina E (IgE). Izlaganje salivi u trajanju od 2 minuta ne utiÄe na biohemijske i imunohemijske osobine derivata. U kiseloj sredini želudaÄnog soka ne dolazi do znaÄajnog demaleilovawa i desukcinilovanja. Äak i posle ÄetvoroÄasovnog izlaganja, taj procenat je u opsegu 10-30 %. Alergoidi pelina se trenutno digestuju pepsinom, sa izuzetkom visoko rezistentne proteinske trake molekulske mase 28-35 kD, koja odgovara važnom IgE-vezujuÄem proteinu polena pelina. Imunoblotom i CAP-inhibicijom je pokazano da, u okviru fizioloÅ”kih uslova, ne dolazi do stvaranja novih IgE-vezujuÄih epitopa. Hemijska stabilnost modifikovanih derivata u simuliranim uslovima želudaÄnog soka omoguÄuje da se tokom imunoterapije mogu primenjivati veÄe doze alergoida nego nemodifikovanog ekstrakta polena pelina
Plasma fucosylated glycans and C-reactive protein as biomarkers of HNF1A-MODY in young adultāonset nonautoimmune diabetes
OBJECTIVE Maturity-onset diabetes of the young (MODY) due to variants in HNF1A is the commonest type of monogenic diabetes. Frequent misdiagnosis results in missed opportunity to use sulfonylureas as first-line treatment. A nongenetic biomarker could improve selection of subjects for genetic testing and increase diagnosis rates. We previously reported that plasma levels of antennary fucosylated N-glycans and hs-CRP are reduced in individuals with HNF1A-MODY. In this study, we examined the potential use of N-glycans and hs-CRP in discriminating individuals with damaging HNF1A alleles from those without HNF1A variants in an unselected population of young adults with nonautoimmune diabetes. RESEARCH DESIGN AND METHODS We analyzed the plasma N-glycan profile, measured hs-CRP, and sequenced HNF1A in 989 individuals with diabetes diagnosed when younger than age 45, persistent endogenous insulin production, and absence of pancreatic autoimmunity. Systematic assessment of rare HNF1A variants was performed. RESULTS We identified 29 individuals harboring 25 rare HNF1A alleles, of which 3 were novel, and 12 (in 16 probands) were considered pathogenic. Antennary fucosylated N-glycans and hs-CRP were able to differentiate subjects with damaging HNF1A alleles from those without rare HNF1A alleles. Glycan GP30 had a receiver operating characteristic curve area under the curve (AUC) of 0.90 (88% sensitivity, 80% specificity, cutoff 0.70%), whereas hs-CRP had an AUC of 0.83 (88% sensitivity, 69% specificity, cutoff 0.81 mg/L). CONCLUSIONS Half of rare HNF1A sequence variants do not cause MODY. N-glycan profile and hs-CRP could both be used as tools, alone or as adjuncts to existing pathways, for identifying individuals at high risk of carrying a damaging HNF1A allele
Inflammatory bowel disease associates with proinflammatory potential of the immunoglobulin g glycome
BACKGROUND: Glycobiology is an underexplored research area in inflammatory bowel disease (IBD), and glycans are relevant to many etiological mechanisms described in IBD. Alterations in N-glycans attached to the immunoglobulin G (IgG) Fc fragment can affect molecular structure and immunological function. Recent genome-wide association studies reveal pleiotropy between IBD and IgG glycosylation. This study aims to explore IgG glycan changes in ulcerative colitis (UC) and Crohn's disease (CD). METHODS: IgG glycome composition in patients with UC (n = 507), CD (n = 287), and controls (n = 320) was analyzed by ultra performance liquid chromatography. RESULTS: Statistically significant differences in IgG glycome composition between patients with UC or CD, compared with controls, were observed. Both UC and CD were associated with significantly decreased IgG galactosylation (digalactosylation, UC: odds ratio [OR] = 0.71; 95% confidence interval [CI], 0.5ā0.9; P = 0.01; CD: OR = 0.41; CI, 0.3ā0.6; P = 1.4 Ć 10(ā9)) and significant decrease in the proportion of sialylated structures in CD (OR = 0.46, CI, 0.3ā0.6, P = 8.4 Ć 10(ā8)). Logistic regression models incorporating measured IgG glycan traits were able to distinguish UC and CD from controls (UC: P = 2.13 Ć 10(ā6) and CD: P = 2.20 Ć 10(ā16)), with receiverāoperator characteristic curves demonstrating better performance of the CD model (area under curve [AUC] = 0.77) over the UC model (AUC = 0.72) (P = 0.026). The ratio of the presence to absence of bisecting GlcNAc in monogalactosylated structures was increased in patients with UC undergoing colectomy compared with no colectomy (FDR-adjusted, P = 0.05). CONCLUSIONS: The observed differences indicate significantly increased inflammatory potential of IgG in IBD. Changes in IgG glycosylation may contribute to IBD pathogenesis and could alter monoclonal antibody therapeutic efficacy. IgG glycan profiles have translational potential as IBD biomarkers