<p>Contact allergy is one of the most common skin diseases with great social and economical impact. The origin and nature of contact allergens (haptens) capable of inducing T-cell mediated allergic reactions are diverse, ranging from organic molecules to metal ions. Most of the current knowledge on T-cell responses to haptens in humans with contact allergy have been established by studies on the metal ion nickel (Ni), the most common cause of contact allergy, whereas reactivity to the large group of organic haptens has been less studied.</p><p>Haptens are not immunogenic by themselves but must bind carrier molecules prior to their presentation on MHC class I or II molecules and subsequent recognition by T cells. Due to differences in their chemical nature, haptens interact with host molecules by different mechanisms and differences in their solubility can influence their access to different antigen-presenting pathways.</p><p>The aim of the present study was to define immune responses elicited by haptens of different chemical nature including Ni (hydrophilic metal ion), methylisothiazolinones (hydrophilic organic molecule) and parthenolide (lipophilic organic molecule). The immune response displayed by subjects with allergy to these substances, and non-allergic control subjects, was assessed by measuring hapten-induced cytokine production in peripheral blood mononuclear cells (PBMC) with a focus on ELISpot analysis of T-cell type 1 (e.g. IFN-g and IL-2) and type 2 (e.g. IL-4, IL-5 and IL-13) cytokines. For Ni and parthenolide, the phenotype of the hapten-reactive T cells was determined. The allergic status of subjects was defined by clinical history and patch testing. The latter is the established diagnostic method for contact allergy, based on applying various haptens to the subjects’ back and grading the skin reaction after 2-3 days.</p><p>All three haptens elicited a concomitant T-cell type 1 and 2 response in subjects with contact allergy to the corresponding hapten, suggesting the induction of a functionally related cytokine profile, irrespective of the chemical character of the hapten. The cytokine response was related to the degree of the subjects’ patch test reactivity; PBMC from a vast majority of subjects with strong and moderate patch test reactivity displayed detectable cytokine responses to the corresponding haptens, whereas subjects with weak or no (controls) patch test reactivity did not. Despite the similar cytokine profile induced, the phenotype of the reactive T cells was found to differ between haptens with Ni eliciting CD4+ T cells and parthenolide eliciting CD8+ T cells. This difference may be explained by a better ability of a lipophilic hapten to gain access to the MHC class I-restricted antigen-presentation pathway. Moreover, the data suggest that analysis of cytokine responses to haptens may facilitate future development of <i>in vitro</i>-based diagnostics assay for contact allergy.</p><p>Finally, the relationship between the variation over time in patch test reactivity and systemic reactivity to Ni, in terms of cytokine responses to Ni <i>in vitro</i>, was investigated. The degree of patch test reactivity is known to vary over time, in particular in subjects with weak reactivity. Ni-allergic subjects were patch tested three times with three month intervals and PBMC obtained at the same time points were assessed for<i> in vitro</i> reactivity to Ni. The overall reactivity in the patch test and the <i>in vitro</i> test was well correlated confirming that both methods provide a good and comparable estimate of the systemic reactivity to Ni. However, fluctuations in the patch test reactivity over time were not well correlated with variations in the cytokine response elicited <i>in vitro</i> suggesting that other parameters besides changes in the systemic reactivity could significantly contribute to the variation in patch test reaction over time.</p>

Contact allergy is one of the most common skin diseases with great social and economical impact. The origin and nature of contact allergens (haptens) capable of inducing T-cell mediated allergic reactions are diverse, ranging from organic molecules to metal ions. Most of the current knowledge on T-cell responses to haptens in humans with contact allergy have been established by studies on the metal ion nickel (Ni), the most common cause of contact allergy, whereas reactivity to the large group of organic haptens has been less studied. Haptens are not immunogenic by themselves but must bind carrier molecules prior to their presentation on MHC class I or II molecules and subsequent recognition by T cells. Due to differences in their chemical nature, haptens interact with host molecules by different mechanisms and differences in their solubility can influence their access to different antigen-presenting pathways. The aim of the present study was to define immune responses elicited by haptens of different chemical nature including Ni (hydrophilic metal ion), methylisothiazolinones (hydrophilic organic molecule) and parthenolide (lipophilic organic molecule). The immune response displayed by subjects with allergy to these substances, and non-allergic control subjects, was assessed by measuring hapten-induced cytokine production in peripheral blood mononuclear cells (PBMC) with a focus on ELISpot analysis of T-cell type 1 (e.g. IFN-g and IL-2) and type 2 (e.g. IL-4, IL-5 and IL-13) cytokines. For Ni and parthenolide, the phenotype of the hapten-reactive T cells was determined. The allergic status of subjects was defined by clinical history and patch testing. The latter is the established diagnostic method for contact allergy, based on applying various haptens to the subjects’ back and grading the skin reaction after 2-3 days. All three haptens elicited a concomitant T-cell type 1 and 2 response in subjects with contact allergy to the corresponding hapten, suggesting the induction of a functionally related cytokine profile, irrespective of the chemical character of the hapten. The cytokine response was related to the degree of the subjects’ patch test reactivity; PBMC from a vast majority of subjects with strong and moderate patch test reactivity displayed detectable cytokine responses to the corresponding haptens, whereas subjects with weak or no (controls) patch test reactivity did not. Despite the similar cytokine profile induced, the phenotype of the reactive T cells was found to differ between haptens with Ni eliciting CD4+ T cells and parthenolide eliciting CD8+ T cells. This difference may be explained by a better ability of a lipophilic hapten to gain access to the MHC class I-restricted antigen-presentation pathway. Moreover, the data suggest that analysis of cytokine responses to haptens may facilitate future development of in vitro-based diagnostics assay for contact allergy. Finally, the relationship between the variation over time in patch test reactivity and systemic reactivity to Ni, in terms of cytokine responses to Ni in vitro, was investigated. The degree of patch test reactivity is known to vary over time, in particular in subjects with weak reactivity. Ni-allergic subjects were patch tested three times with three month intervals and PBMC obtained at the same time points were assessed for in vitro reactivity to Ni. The overall reactivity in the patch test and the in vitro test was well correlated confirming that both methods provide a good and comparable estimate of the systemic reactivity to Ni. However, fluctuations in the patch test reactivity over time were not well correlated with variations in the cytokine response elicited in vitro suggesting that other parameters besides changes in the systemic reactivity could significantly contribute to the variation in patch test reaction over time.

En allergisk person påverkas på fysisk, psykisk och social nivå. Unga vuxna allergiska personers erfarenheter och upplevelser av allergi är relativt outforskat. Syftet med studien är att förstå hur unga vuxna personer med allergi upplever att allergin påverkar deras vardag och hur de förhåller sig till ett liv anpassat efter allergin. En kvalitativ intervjustudie genomfördes med 10 deltagare i åldrarna 18-22 år som hade olika former av allergier. Resultatet visade att allergi har en betydande roll för allergiska personers identitetsskapande och många beskriver en strävan efter att passa in socialt trots sina besvär. I denna strävan görs en avvägning mellan de risker som det innebär och värdet av den aktuella situationen. Intervjupersonerna upplevde att förståelsen av allergiers konsekvenser är bristfällig vilket kan försvåra för en allergisk person i dess vardag. Resultatet visade att deltagarnas livskvalitet påverkades negativt på grund av allergin, samtidigt beskrevs livskvaliteten på ett motsägelsefullt sätt av deltagarna.

Proteins are important building blocks of all living organisms. They are composed of a defined sequence of different amino acids, and fold into a specific three-dimensional, ordered structure. The three-dimensional structure largely determines the function of the protein, but protein function always requires motion. Small movements within the protein structure govern the functional properties, and this thesis aims to better understand these discrete protein movements. The motions within the protein structure are governed by thermodynamics, which therefore is useful to predict protein interactions. Nuclear magnetic resonance (NMR) is a powerful tool to study proteins at atomic resolution. Therefore, NMR is the primary method used within this thesis, along with other biophysical techniques such as Fluorescence spectroscopy, Circular Dichroism spectroscopy and in silico modeling. In paper I, NMR in combination with molecular engineering is used to show that the folding of the catalytical subdomains of the enzyme Adenylate kinase does not affect the core of the protein, and thus takes a first step to linking folding, thermodynamic stability and catalysis. In paper II, the structure of the primary allergen from Brazil nut, Ber e 1, is presented along with biophysical measurements that help explain the allergenic potential of the protein. Paper III describes the need for a specific Brazil nut lipid fraction needed to induce an allergenic response. NMR and fluorescence spectroscopy is used to show that there is a direct interaction between Ber e 1 and one or several components in the lipid fraction.

Benzylpenicilloyl-poly-L-cysteine is prepared by reacting benzylpenicillenic acid with poly-L-cysteine at 370° in water at pH 8.3 or buffer at pH 7.98. The preparations of penicilloyl-cysteine, S-acetamidomethyl-polycysteine, and S-acetamidomethyl-penicilloyl-polycysteine are also described. C14-labeling and penamaldate assays are used to determine penicilloyl content. The iodoacetic acid method is used to ascertain thiol content.The ultraviolet spectra for penicilloyl-polycysteine, pencilloyl-cysteine, poly-L-cysteine, poly-S-carbobenzoxy-L-cysteine, S-acetamidomethyl-polycysteine and S-acetamidomethyl-penicilloyl-polycysteine are reported. The infrared spectra of penicilloyl-polycysteine, penicilloyl-cysteine, poly-S-carbobenzoxy-L-cysteine, S-acetamidomethyl-polycysteine, and S-acetamidomethyl-penicilloyl-polycysteine are reported. The nmr spectra of poly-L-cysteine, penicilloyl-cysteine, S-acetamidomethyl-polycysteine, and S-acetamidomethyl-penicilloyl-polycysteine are discussed.The kinetic rates of reaction of benzylpenicillenic acid at 37.5 ± 0.50 in buffer, cysteine, N, S-di-CBZ L-cysteine, poly-L-cysteine, poly-S-CBZ L-cysteine andβ-mercaptoethylamine are compared.Ball State UniversityMuncie, IN 47306

Benzylpenicillin, a-aminobenzylpenicillin, phenoxymethylpenicillin, and 2,6-dimethoxy-phenylpenicillin were incubated with whole human blood. Migration patterns into human erythrocytes were compared spectrophotometrically.A possible inhibition effect by chloroquine on benzyl-[14C]-penicillin in red blood cells was studied via a Beckman LS-100C Liquid Scintillation Counter.An intraerythrocytic protein, carbonic anhydrase, was investigated as being responsible for the hydrolysis of benzylpenicillin. Three carbonic anhydrases (a, b, and c) were quantitatively measured via a Beckman IR 4250 Spectrophotometer for their effect upon the a-lactam ring of benzylpenicillin.Ball State UniversityMuncie, IN 47306

Polymorphonuclear neutrophils (PMNs) are important effector cells in host defense and the inflammatory response to antigen. The involvement of PMNs in inflammation is mainly mediated by the Fc receptor family, including IgE receptors. Recently, we have shown that human PMNs from allergic asthmatic subjects express the high affinity receptor, FceRI. In this study, we have examined the regulation of FceRI by human PMNs in vitro and in vivo during the allergic pollen season. First we studied the pattern of expression of FceRI in PMNs during the pollen allergic and outside the pollen season. Peripheral blood neutrophils were isolated from adult atopic asthmatics (AA) (n=17), allergic non asthmatics (ANA) (n=15) and healthy donors (n=16) by dextran, ficoll gradient centrifugation and magnetic cell sorting (MACS). Surface, total protein and mRNA expression of FceRI were investigated in the three groups by FACS, immunocytochemistry (ICC) and fluorescent in situ hybridization (FISH) respectively. Secondly, we investigated the effect of Th-2 cytokines which are known to regulate IgE receptor expression. PMNs from atopic asthmatic subjects were stimulated in vitro with Th-2 cytokines (IL-4, IL-9, GM-CSF) and Th-1 cytokine IFN-gamma. Finally we determined whether the expression of FceRIbeta chain correlated with the surface expression of FceRIalpha chain in PMNs. Irrespective of the season, PMNs from atopic asthmatic subjects showed increased expression of FceRIalpha chain in surface, total protein and mRNA compared to atopic non asthmatics and healthy donors (n=20). Interestingly, FceRIalpha chain surface and mRNA expression increased significantly during pollen season compared to non pollen season (P=0.001) in PMNs isolated from AA (n=9) in contrast to healthy donors and ANA (n=8). Furthermore similar pattern of FceRI expression were observed in vitro when PMNs were stimulated with Th2 cytokines. IL-4, IL-9 and GM-CSF showed increased protein and mRNA expression of FceRIalpha chain at 6 and 18hrs (n=6) whereas IFN-gamma down regulated the mRNA expression of FceRIalpha chain at 6hrs. Also, irrespective of season AA (n=11) subjects showed increased expression of FceRI beta chain when compared to ANA (n=10) and healthy donors (n=9). Western blot analysis showed increased FceRI beta protein in atopic asthmatic subjects (n=4). Interestingly irrespective of the groups, there was a positive correlation r = 0.8054 between total protein expression of beta chain with surface expression of alpha chain of FceRI in neutrophils. Our data suggest that the expression of FceRI in neutrophils of atopic asthmatic patients is highly regulated. Our in vitro studies provide evidence that Th-2 cytokines such as IL-9, IL-4 and GM-CSF up-regulate the expression of FceRI. Furthermore we show evidence of increased expression of FceRIbeta chain in neutrophils of atopic asthmatic subjects. Collectively these results suggest that FceRI mediated neutrophil dependent activation may play a key role in allergic diseases. / May 2005

Syftet var att beskriva omvårdnaden kring nyfödda barns tillmatning på svenska BB-avdelningar, om föreskrifter följdes och om hänsyn togs till eventuell allergihereditet. Metod: Deskriptiv tvärsnittsstudie med kvantitativ ansats Resultat: Av alla nyfödda barn vid 26 barnkliniker i Sverige vecka 26 år 2009 (n=849) tillmatades 18 % . Moderns bröstmjölk gavs till 16% medan 78 % tillmatades med komjölksbaserad modersmjölksersättning. Den vanligaste orsaken till tillmatning av ersättning var att mammans egen mjölkproduktion inte hade kommit igång (24 %). Andra vanligt förekommande anledningar till tillmatning var lågt blodsocker (18 %) samt sugproblem hos barnen (16 %). Sexton barn tillmatades trots ett blodsockervärde motsvarande &gt;2.2mmol/l. Vad gäller allergiförekomst tillfrågades endast 37 % av barnens föräldrar före tillmatning. Dokumentation om tillmatning fanns noterat i 80 % av barnens journaler. Motsvarande siffra för mammornas journaler var 34 %. Konklusion: Bristfälliga omvårdnadsåtgärder vidtas i samband med tillmatning av nyfödda spädbarn på BB-avdelningar, där allergifrågor ställs i för låg utsträckning. BB-kliniker har varierande referensvärde för vad som räknas som lågt blodsocker. Omvårdnadsbehov för nyfödda barn och deras föräldrar, där förmedlad kunskap om amningens betydelse, tas inte på allvar och oklara riktlinjer skapar förvirring. Föreskrifter och riktlinjer bör följas i större utsträckning an vad denna studie visar.

<p>The objective with this Master’s thesis was to develop, implement and evaluate an iterative procedure for hierarchical clustering with good overall performance which also merges features of certain already described algorithms into a single integrated package. An accordingly built tool was then applied to an allergen IgE-reactivity data set. The finally implemented algorithm uses a hierarchical approach which illustrates the emergence of patterns in the data. At each level of the hierarchical tree a partitional clustering method is used to divide data into k groups, where the number k is decided through application of cluster validation techniques. The cross-reactivity analysis, by means of the new algorithm, largely arrives at anticipated cluster formations in the allergen data, which strengthen results obtained through previous studies on the subject. Notably, though, certain unexpected findings presented in the former analysis where aggregated differently, and more in line with phylogenetic and protein family relationships, by the novel clustering package.</p>

published_or_final_version / Community Medicine / Master / Master of Public Health