bluuebunny/crossref_metadata_2025 huggingface dataset:
Biological Effects of Newly Synthesized Cholecystokinin Analogs
Authors: Verspohl Eugen J. , LaMura Marco
Published: April 2003

Cholecystokinin (CCK) is a gut hormone that regulates pancreatic endocrine functions via CCKA receptors. CCK4 (Trp-Met-Asp-Phe-NH2) has an insulinotropic effect, but is 1,000-fold less potent than CCK8 in rodents. The in vitro potencies with respect to binding, the biological effects and the selectivity of newly synthesized CCK4 analogs constructed by computer modelling experiments were investigated in vitro in rat pancreas and brain, INS-1 cells, and guinea pig ileum. Exchanging various amino acids, e.g. Met by either Pro or Nle, and modifying Phe by adding various substituents in different positions led to compounds which were more effective as insulin secretagogues than CCK4 itself and even show insulinotropic effects comparable with those of CCK8 (e.g. compounds M1 and M2 being substituted at Phe). Some compounds which possess electron withdrawing groups on the C-terminal Phe and possess a Pro instead of a Met were especially effective. The CCKA receptor antagonist L-364,718, but not by the CCKB receptor antagonist L-365,260, inhibited the insulinotropic effects. The synthetic CCK4 compounds were not selective for the endocrine pancreas: e.g. M1 and M2 had binding activity with respect to rat brain homogenates but no activity with respect to contraction of the guinea pig ileum. The data indicate that some of the newly synthesized CCK tetrapeptides exhibit a high affinity for the CCK receptor of β-cells and have an insulinotropic effect much higher than CCK4.
Antiproliferative Effect and Cell Cycle Modulation by Melatonin on GH3 Cells
Authors: Fornas Oscar , Mato M. Eugenia , Webb Susan M.
Published: April 2003

We have investigated the effect of melatonin on cell proliferation and modulation, in the GH3 experimental rat pituitary cell line; the expression of oncogenes c-myc, c-jun and the tumor suppressor gene p53 were also analyzed basally and after exposure to melatonin (10–6, 10–8 and 10–10 M). Melatonin exhibited an antiproliferative effect at all the doses tested, decreasing the proliferating index by 50%. After exposure to melatonin, a decrease in Ki67 and Proliferation cell nuclear antigen occurred acute- and transiently (at 2 h) after a single dose which recovered at 4 h, as well as chronically after repeated 12-hour doses which persisted at 48 h; a similar behavior was observed both acute- and chronically for c-myc and c-jun, while it was opposite for p53, rising acute- and transiently as well as after repeated exposure. These results demonstrate that melatonin modulates the proliferation mechanisms of the GH3 cells.
Theophylline Suppresses the Release of Interleukin-4 by Peripheral Blood Mononuclear Cells
Authors: Tohda Yuji , Nakahara Hiroshi , Kubo Hirokazu , Muraki Masato , Fukuoka Masahiro , Nakajima Shigenori
Published: April 2003

To elucidate the mechanism of the anti-inflammatory effect of theophylline, peripheral blood mononuclear cells (PBMCs) collected from patients with bronchial asthma who had a RAST score for Dermatophagoides farinae (Df) > = 3 were stimulated with Df antigen, and the interleukin-4 (IL-4) production by the stimulated PBMCs was determined in the presence and absence of theophylline. The stimulated IL-4 production in the PBMCs was significantly suppressed by simultaneous treatment with theophylline (8 μg/ml). Furthermore, the intracellular 3′,5′-adenosine monophosphate (cAMP) level following stimulation with Df was significantly higher in PBMCs simultaneously treated with theophylline (8 μg/ml) than in nontreated controls. These results suggest that theophylline suppresses the IL-4 production in inflammatory cells by increasing the intracellular cAMP level; this effect may thus provide an additional basis for the benefit of theophylline in antiasthmatic therapy.
IgE /Anti-IgE Immune Complexes in Sera from Patients with Crohn’s Disease Do Not Contain Food-Specific IgE
Authors: Huber Andrea , Genser Dieter , Spitzauer Suzanne , Scheiner Otto , Jensen-Jarolim Erika
Published: April 2003

Background: An association of Crohn’s disease (CD) with food allergy has been discussed, but the role of food-specific IgE has not been clarified yet. Since CD is combined with immune complex formation, we examined in the present study whether anti-IgE autoantibodies in such complexes might hinder the determination of specific IgE. Methods: In order to elucidate the role of food-specific IgE in CD, we tested sera from CD patients (n = 107), healthy controls (n = 65) and allergics subjects (n = 7) for their IgE binding to food antigens (yeast, corn, celeriac, wheat) by an immunodot assay. After determining levels of IgE/IgG anti-IgE immune complexes, we purified them from serum pools of patients with CD, allergic subjects and healthy controls by affinity absorption using a monoclonal anti-IgE antibody. These purified immune complexes were treated by low pH (pH = 4) in order to dissolve them and to increase the detectability of food-specific IgE by RAST and CAP assay. Results: In CD sera no food-specific IgE could be detected, but levels of immune complexes of IgE and IgG anti-IgE autoantibodies were statistically significantly increased compared to healthy controls. pH treatment of purified IgE/IgG anti-IgE immune complexes resulted in a significant increase in specific IgE to yeast, corn, wheat and celeriac detected by RAST, however, only in the serum sample purified from allergic subjects. After pH treatment of CD immune complexes, specific IgE levels remained still very low. Conclusion: Thus, even if IgE seems to represent an autoantigen in CD, it is unlike to specifically participate in the pathophysiology of the putative food adverse reactions.
Interleukin-12 Inhibits Production of Interleukin-5 but Not of Granulocyte/Macrophage Colony-Stimulating Factor by Antigen-Stimulated Blood Mononuclear Cells in Allergic Bronchial Asthmatics
Authors: Okano Yoshio , Nakamura Yoichi , Sano Takahiro , Azuma Masahiko , Sone Saburo
Published: April 2003

Bronchial asthma, characterized by eosinophilic inflammation in airways, may involve Th2-type cytokines such as interleukin-5 (IL-5). IL-12, a newly established cytokine, induces IFN-γ production, which may have a regulatory effect on the production of Th2-type cytokines. We examined the effects of IL-12 on the productions of IL-5 and granulocyte/macrophage colony-stimulating factor (GM-CSF) by antigen (Dermatophagoides farinae, Df)-stimulated mononuclear cells (MNCs) from asthmatic patients in vitro. IL-12 enhanced IFN-γ production and inhibited the production of IL-5 but not of GM-CSF by Df-stimulated MNCs from asthmatic patients. Exogenous IFN-γ directly inhibited IL-5 production by Df-stimulated MNCs and the inhibition of IL-5 production by IL-12 was partially blocked by anti-IFN-γ antibody in culture, indicating that inhibitory effect of IL-12 on IL-5 production by antigen-stimulated MNCs is partially dependent on IFN-γ production. IL-12 also inhibited the release of eosinophil survival-stimulating factor from MNCs. These results indicate that IL-12 may be therapeutically beneficial in correcting Th1/Th2 imbalance in bronchial asthma.
Allergen-Specific Nasal Challenge: Response Kinetics of Clinical and Inflammatory Events to Rechallenge
Authors: Ciprandi Giorgio , Ricca Vittorio , Landi Massimo , Passalacqua Giovanni , Bagnasco Marcello , Canonica G. Walter
Published: April 2003

Allergen-specific nasal challenge is a valid and reliable tool for studying the pathophysiological mechanisms involved in allergic inflammation. Nasal challenge induces an immediate clinical response in allergic subjects and a concomitant appearance of an inflammatory infiltrate. The mucosal inflammation may persist up to 48–72 h after allergen exposure. If the subjects are rechallenged within this period the response is more pronounced: the so-defined priming effect. The aim of the study was to evaluate the effects of the nasal rechallenge, performed at different time intervals: 3 days, 1, 2 and 4 weeks after the first challenge. Forty allergic subjects underwent two nasal challenges: at baseline and after the periods mentioned above (10/group). Symptoms and inflammatory cells (eosinophils and neutrophils recovered by nasal brushing) were assessed. The 3-day-interval group showed a hyperreactive response (priming effect), the 1- and 4-week-interval groups showed a response similar to baseline, and the 2-week-interval group showed a hyporeactive response (‘tolerogenic effect’). The last phenomenon may be due to a possible immunologic response similar to that achievable during local specific immunotherapy. The present results further elucidate the kinetics of allergen-driven inflammatory events and highlight the importance of the time period chosen for rechallenge. The latter fact may be of primary importance in clinical trials involving specific challenge.
A Role for Histamine and Substance P in Immediate Allergic Responses in Guinea Pig Airways: Characterization of MDL 108,207DA, a Dual H1 / NK-1 Receptor Antagonist
Authors: Kudlacz Elizabeth , Shatzer Scott , Logan Deborah , Olsen Karl , Knippenberg Robert , Hsieh Lily , Esteve Hector , Maynard George
Published: April 2003

Background: Histamine is a critical mediator of immediate hypersensitivity reactions. Sensory neuropeptides, such as substance P (SP), may also contribute to acute inflammatory responses. A compound which antagonizes both H1 and NK-1 receptors, such as MDL 108,207DA, may present a significant therapeutic advantage over pure antihistamines. Methods: The binding affinity of MDL 108,207DA for H1 and NK-1 receptors was evaluated and its potency of antagonism evaluated in vitro. The in vivo antagonism of SP- or histamine-induced microvascular leakage in guinea pig airways was examined. A role for these mediators in antigen-induced microvascular leakage in ovalbumin-sensitized guinea pig airways was examined using MDL 108,207DA as well as the NK-1-selective antagonist FK888 and the H1-selective antagonist pyrilamine alone or in combination. Results: The affinity of MDL 108,207DA for H1 and NK-1 receptors is similar to that of receptor-selective antagonists. The compound inhibits both receptors in vitro and in vivo with comparable potencies for each. The efficacy of FK888 in combination with pyrilamine and MDL 108,207DA on antigen-induced microvascular leakage in sensitized guinea pig airways supports a role for both SP and histamine in early allergic responses. Conclusion: The contribution of both SP and histamine to immediate hypersensitivity reactions supports the utility of NK-1 and H1 receptor antagonist therapy. MDL 108,207DA incorporates both activities into the same compound and, as a result, may be useful in the treatment of allergic diseases.
Differential Endothelial Adhesion Molecule Expression in Early and Late Whealing Reactions
Authors: Haas Norbert , Schadendorf Dirk , Henz Beate M.
Published: April 2003

In order to clarify the pathomechanisms of fleeting versus more persistent wheals, expression of endothelial adhesion molecules was studied in biopsies of lesional and uninvolved skin of 15 patients with different types of whealing reactions, using immunohistochemistry. In wheals of ≤ 30 min duration, no increase of ELAM-1 and ICAM-1 was noted. GMP-140 expression was absent in prick tests, but could be demonstrated in lesions of cholinergic and cold urticaria, with a gradual increase of the latter with time. In wheals of ≥ 6 h duration, GMP-140 was only weakly expressed whereas ELAM-1 and ICAM-1 were markedly up-regulated in lesional and less so in nonlesional skin of acute, chronic recurrent and delayed pressure urticaria. This differential expression of endothelial adhesion molecules may reflect the activity of mast cell-derived and other mediators during the elicitation phase and explains the persistence of wheals in different types of urticaria.
Molecular and Immunological Characteristics of Latex Allergens
Authors: Breiteneder Heimo , Scheiner Otto
Published: April 2003

Natural rubber latex proteins are a well-recognized cause of type-I allergic reactions that increasingly afflict health-care workers, housekeeping personnel, and other persons using latex gloves or latex products. More than a dozen individual latex allergens have been identified of which eight have received an international nomenclature designation. To study the biochemical and immunological properties in detail, it is desirable to clone and express each of these allergens. Proteins that are involved in rubber synthesis are most likely confined to latex whereas other enzymes, such as chitinases or glucanases, are also present in fruits and may account for the symptoms observed in the latex-fruit syndrome. The available data on the molecular characterization of latex allergens presented here illustrate the importance of this research in the production of better diagnostic tests and, perhaps, tools for immunotherapy.
Contractile Responses to Allergen and Cholinergic Stimulation in Bronchus and Trachea of Cockroach Allergen-Sensitized Guinea Pigs
Authors: Chen Gang , Zhou Daohong , Kim Jin  -Tai , Wu Zhongxin , Lee Lu  -Yuan , Kang Bann C.
Published: April 2003

Airway hyperreactivity is a hallmark of asthma. Yet, the role of allergic sensitization in the hyperreactivity remains controversial. This study examined the effects of airborne cockroach allergen (CRa) sensitization on the contractility of bronchial (BSM) and tracheal (TSM) smooth muscle rings to cholinergic stimulations, and to specific antigen, in vitro. Guinea pigs were actively immunized with CRa, 5 mg (high dose, HD), 0.5 mg (low dose, LD), or saline aerosols (2 ×/day, 5 days/week, for 4 weeks) and challenged by CRa inhalation 5 days after last exposure. Twenty-four hours postchallenge, the contractions of the TSM and BSM to electrical field stimulation (EFS), acetylcholine (ACh), and to CRa were measured in vitro. In addition, the contraction to these stimuli were also examined in the passively sensitized TSM with the sera of CRa-immunized guinea pigs. The contractile responses of actively immunized and passively sensitized tissues to EFS and ACh were very similar to those of control tissues. CRa induced dose-dependent contractions in both actively immunized and passively sensitized, but not in control, tissues. The CRa-induced contraction was stronger in the HD group than in the LD group (p < 0.0001). Leukotriene C4/D4 receptor antagonist LY-171883 inhibited the CRa-induced contraction by 86% in actively immunized TSM, and by 9% in the passively sensitized TSM. Pyrilamine inhibited the contraction by 57% in actively immunized TSM and 70% in passively sensitized TSM. The results indicate that CRa sensitization does not cause increased airway smooth muscle contractility to cholinergic stimulations, but induces antigen-specific contractions in vitro. Leukotrienes appear to play a significant role in the CRa-specific contractions, more in the actively immunized than those in the passively sensitized tissues, while histamine exerts a moderate effect on the CRa-induced contractions.
DNA Immunotherapeutics: New Potential Treatment Modalities for Allergic Disease
Authors: Goodman Justin S. , van Uden John H. , Kobayashi Hiroko , Broide D. , Raz Eyal
Published: April 2003

Genetic immunization is a relatively new approach to vaccination, one that has generated considerable interest for its potential to prevent or treat a number of types of disease processes. Although initial work focused on potential applications in infectious disease, it was soon recognized that the antigen–specific Th1 response typically generated by genetic immunization protocols could also be useful for the treatment of allergic disease. In this review we present a summary of genetic vaccination and the related topic of immunostimulatory DNA sequences – which we collectively designate DNA immunotherapeutics – and discuss their potential for the prevention and therapy of allergic disease.
Bronchoprotective Effect of an Intrabronchial Administration of Cilostazol Powder and a Nebulized PDE1 and PDE4 Inhibitor KF19514 in Guinea Pigs
Authors: Fujimura Masaki , Tachibana Hideki , Myou Shigeharu , Kita Toshiyuki , Matsuda Tamotsu
Published: April 2003

Background: Both oral and intravenous inhibitors of cyclic nucleotide phosphodiesterase (PDE) 3 and PDE4 have been shown to have bronchodilator and/or pulmonary antiallergic effects but their adverse effects limit the clinical use. The purpose of the present study was to assess the bronchoprotective effect of intrabronchial administration of a selective PDE3 inhibitor, 6–[4–(1–cyclohexyl–1H–tetrazol–5–yl)butoxy]–3,4–dihydro–2(1H)–quinolinone (cilostazol), in dry power form which is proposed to be desirable rather than propellant–driven metered–dose inhalers. Methods: Effect of an intrabronchial administration of cilostazol powder on aerosolized histamine– and antigen–induced leukotriene–mediated bronchoconstriction were examined in anesthetized and artificially ventilated guinea pigs, in comparison with the effects of ultrasonically nebulized 5–phenyl–3–(3–pyridyl)methyl–3H–imidazo[4,5–c] [1,8] naphthyridin–4(5H)–one (KF19514), a selective PDE1/4 inhibitor. Results: Cilostazol powder and KF19514 solution inhibited histamine– and antigen–induced bronchoconstriction in a dose–dependent manner. When assessing the resulting ED50 values, the activity of cilostazol powder against antigen–induced broncho–constriction was weaker than its antagonism of histamine–induced bronchoconstriction (60.4 vs. 7.0 μg), while those of KF19514 were the opposite (0.40 vs. 2.81 μg/ml). Conclusion: These results suggest that intrabronchially administered cilostazol powder has a bronchodilator effect, and ensure that PDE4 inhibitors have an antiallergic activity in addition to their bronchodilator effect.
Molecular Cloning of IgE–Binding Fragments of Alternaria alternata Allergens
Authors: De Vouge Michael W. , Thaker Ajay J. , Zhang Lei , Muradia Gauri , Rode Harold , Vijay Hari M.
Published: April 2003

Exposure to the hyphomycete Alternaria alternata is recognized as an important risk factor for asthma. IgE immunoblotting has been used to catalogue the number and Mr of allergens in A. alternata extracts, with estimates ranging from 10 to 30, although few are present in nearly all extracts studied. Several A. alternata allergens have been cloned, including a subunit of the major allergen Alt a 1, ribosomal P2 phosphoprotein, aldehyde dehydrogenase and a yeast YCP4 homolog. We have cloned two sequences encoding IgE–binding fragments of allergens from an A. alternata λgt11 cDNA library using pooled atopic sera from A. alternata–sensitive individuals. One is homologous to a region near the C–terminus of hsp70 from Cladosporium herbarum; a near–complete isoallergen variant of A. alternata ribosomal P2 protein was also cloned. Their lacZ fusion proteins had reactivities of 5 and 14%, respectively, with individual atopic sera, indicating that the corresponding allergens are both minor. This study describes one new A. alternata allergen candidate and implicates ribosomal P2 protein as an allergen thtat is stable between independently isolated clones.
Inhibition of Guinea Pig Skin Allergic Reactions by Nonpeptide Bradykinin B2 Receptor Antagonist FR173657
Authors: Mori Takeshi , Imamura Takahisa
Published: April 2003

Background: Kinins, which cause vascular permeability enhancement (VPE) through bradykinin (BK) B2 receptors, are released at allergic reaction sites; however, the role kinins play in the lesions is still unknown. To determine whether kinins contribute to allergic reactions, the effect of a potent, nonpeptide BK B2–receptor–specific antagonist, FR173657, on VPE induced in the reaction sites was investigated. Methods: Type I allergic reaction was induced by intradermal injections of dinitrophenol (DNP)–bovine serum albumin (BSA) into guinea pigs having received an intravenous injection of anti–DNP antiserum a week before. Type III allergic reaction was induced by intradermal injections of BSA into animals sensitized with subcutaneous injections of complete Freund's adjuvant–emulsified BSA 2 weeks before. VPE and swelling were measured by the leakage of intravenously injected dye and double thickness, respectively, at allergic reaction sites. FR173657 (4 mg/kg body weight) was administered orally 30 min before antigen injection. Results: In the type I allergy model, FR173657 inhibited 34% of the early–phase VPE, which occurs a few minutes after antigen injection, and 50% of the subsequent swelling at the allergic reaction sites. In the type III allergy model, FR173657 also inhibited 30% of the early VPE but neither the late VPE, which occurs from 1 h to at least 24 h after the antigen injection, nor the following swelling at the allergic reaction sites. FR173657 inhibited BK–induced VPE almost completely without affecting histamine–induced VPE. Conclusions: Kinins contribute significantly to the VPE induced in the animal allergy models. FR173657 may be useful for the therapy of human allergic diseases.
Local and Systemic Tolerance to Orally Administered Dinitrochlorobenzene Is Not Broken by Cholera Toxin
Authors: Oliver Alfred R. , Silbart Lawrence K.
Published: April 2003

Background: It is widely acknowledged that oral administration of many antigens induces antigen–specific systemic tolerance. In this study we tested the hypothesis that oral administration of a low dose of dinitrochlorobenzene (DNCB) could induce local tolerance in the absence of systemic tolerance. We also hypothesized that the mucosal adjuvant cholera toxin (CT), which prevents the induction of local and systemic tolerance to coadministered antigens, would be unable to break an established tolerance to an orally administered antigen. Methods: Tolerance was induced in BALB/c mice by oral administration of either a high (5.0 mg) or a low (0.05 mg) oral dose of the contact–sensitizing agent DNCB. This pretreatment was followed by parenteral or oral administration of dinitrophenyl (DNP)–carrier protein conjugates. Results: As anticipated, the high DNCB dose induced systemic tolerance, as evidenced by depressed delayed type hypersensitivity responses to DNCB and reduced serum IgG anti–DNP responses. Oral pretreatment with the high dose of DNCB also induced local tolerance, as indicated by reduced fecal IgA and IgG anti–DNP responses. Conversely, oral pretreatment with the low dose of DNCB induced only local, not systemic tolerance. In addition, CT was incapable of breaking the preexisting tolerance induced by oral DNCB pretreatment. Conclusion: This study and others support the notion that the mucosal arm of the immune system is more sensitive to the induction of tolerance than the systemic arm, and that CT may not be an effective adjuvant for antigens to which the mucosal immune system has previously been exposed.
Analysis of the Cross–Reactivity between BtM and Der p 5, Two Group 5 Recombinant Allergens from Blomia tropicalis and Dermatophagoides pteronyssinus
Authors: Caraballo Luis , Mercado Dilia , Jiménez Silvia , Moreno Liliana , Puerta Leonardo , Chua Kaw Yan
Published: April 2003

Background: In tropical climates, sensitization to Bloma tropicalis and Dermatophagoides pteronyssinus is high and mainly directed to species–specific allergens. There is some cross–reactivity between extracts of these mites, probably due to the group 5 allergens that have high sequence homology. Objective and Methods: We used the radioallergosorbent test (RAST), RAST inhibition and immunoblotting inhibition experiments to investigate the cross–reactivity between the recombinant allergens BtM and Der p 5, expressed as glutathione S–transferase fusion proteins, to detect the epitopes involved and to analyze the importance of this cross–reactivity. Results: Seventy–nine percent of 48 patients sera were RAST positive to both recombinants, with a strong correlation (r = 0.8, p<0.0001). BtM inhibited 25 and 21.1% of IgE–binding to B. tropicalis and D. pteronyssinus extracts respectively and Der p 5 inhibited 22 and 24% of IgE–binding to D. pteronyssinus and B. tropicalis extracts. Furthermore, BtM inhibited 74.5% of IgE binding to Der p 5 and Der p 5 inhibited 72.4% of IgE–binding to BtM. RAST inhibition with BtM–derived synthetic peptides showed that peptide 4 (residues 35–50) and peptide 5 (residues 46–61) inhibited 37 and 16% of IgE–binding to BtM while peptides 5 and 2 (residues 14–30) were able to inhibit the IgE binding (32 and 28%, respectively) to Der p 5. Conclusion: There is cross–reactivity between BtM and Der p 5, which explains almost all the cross–reactivity between the two mite extracts. This cross–reactivity seems to be related to epitope(s) at the C–terminal segment of these allergens.
Expression of Adhesion Molecules in Cultured Human Nasal Mucosal Microvascular Endothelial Cells Activated by Interleukin–1â or Tumor Necrosis Factor–·: Effects of Dexamethasone
Authors: Yamamoto Yuji , Ikeda Katsuhisa , Watanabe Masako , Shimomura Akira , Suzuki Hideaki , Oshima Takeshi , Imamura Yoko , Ohuchi Kazuo , Takasaka Tomonori
Published: April 2003

Adhesion molecules of microvascular endothelial cells play a key role in the inflammatory processes involved in nonallergic sinusitis. We investigated the cytokine–regulated expression of intercellular adhesion molecule–1 (ICAM–1), E–selectin and vascular cell adhesion molecule–1 (VCAM–1), and the effect of dexamethasone on these expressions in cultured human nasal microvascular endothelial cells (HNMEC). ICAM–1 was enhanced, and E–selectin and VCAM–1 were induced in a dose–dependent fashion following stimulation with IL–1β or TNF–α. HNMEC differed from human umbilical vein endothelila cells in that (1) maximal upregulation of ICAM–1 expression induced by IL–1β or TNF–α required more time (2) TNF–α was more potent than IL–1β in VCAM–1 expression, and (3) dexamethasone inhibited the upregulation of E–selectin expression alone. These findings contribute to a better understanding of the characteristic features of leukocyte infiltration into inflamed tissue and the effect of glucocorticoid in nonallergic chronic sinusitis.
Identification of the Schistosoma japonicum 22.6–kDa Antigen as a Major Target of the Human IgE Response: Similarity of IgE–Binding Epitopes to Allergen Peptides
Authors: Santiago Mario Luis , Hafalla Julius Clemence R. , Kurtis Jonathan D. , Aligui Gemiliano L. , Wiest Peter M. , Olveda Remigio M. , Olds G. Richard , Dunne David W. , Ramirez Bernadette L.
Published: April 2003

Human resistance to reinfection with Schistosoma mansoni and Schistosoma haematobium correlates with elevated IgE titers against worm antigens (soluble worm antigen preparation, SWAP). In S. mansoni infection, low levels of reinfection following chemotherapy are associated with the recognition of a cloned tegumental protein Sm22.6. Because of potential species–specific differences in resistance to schistosomes, we attempted to identify Schistosoma japonicum antigens recognized by human IgE. Following a survey of 176 infected individuals in Leyte, Philippines, we show that IgE antibodies from the majority of older, high–IgE/SWAP responders recognize antigens in the 22 (Sj22)–, 45–, 78– and 97–kDa range in SWAP. Limited IgE cross–reactivity between Sj22 and Sm22 was observed following a comparison of Filipino IgE responses to these antigens. The antigen was cloned from an adult S. japonicum λ–ZAP cDNA library (Mindoro strain) by immunoscreening with pooled high–titer IgE antisera and a rabbit anti–Sj22 polyclonal antibody. The deduced amino acid sequence of the identified cDNA clone, MJ–1, showed significant homology to Sm22.6 (74%) and Sj22.6 (99%). Although the molecular sequence of Sj22.6 has already been reported, this is the first demonstration of its recognition by human IgE, thereby strengthening its potential as a vaccine candidate. Using an overlapping peptide approach, four IgE–binding epitopes were identified in Sj22.6, two of which exhibited similarities to known IgE–binding epitopes from codfish (Gad c 1) and β–lactoglobulin–related allergens. These findings suggest that allergy and protective immunity to helminth infection may be linked by the structural similarities of epitopes recognized by human IgE.
Bronchial Hyperresponsiveness, Hypersensitivity to Analgesics and Urinary Leukotriene E4 Excretion in Patients with Aspirin–Intolerant Asthma
Authors: Yoshida S. , Nakagawa H. , Yamawaki Y. , Sakamoto H. , Akahori K. , Nakabayashi M. , Sakamoto M. , Hasegawa H. , Shoji T. , Tajimab T. , Amayasu H.
Published: April 2003

This study was designed to investigate the protective effect of cromolyn sodium on airway sensitivity to sulpyrine, and bronchial responsiveness to methacholine, and to investigate whether this protective activity is associated with reduction in aspirin–induced excretion of urinary leukotriene E4 (u–LTE4), a marker of the cysteinyl LT overproduction that participates in the pathogenesis of aspirin–induced asthma. We assessed the effects of pretreatment with cromolyn sodium on bronchoconstriction precipitated by inhalation of methacholine and sulpyrine in 16 adult patients with mild or moderate aspirin–intolerant asthma; those who were in stable clinical condition and were hypersensitive to a sulpyrine provocation test were included in this study. A double–blind, randomized, crossover design was used. u–LTE4 was measured using combined reverse–phase high–performance liquid chromatography enzyme immunoassay. Cromolyn sodium protected against analgesic–induced bronchoconstriction through mechanisms that are not related to its bronchodilator property, but to the improvement of both bronchial hyperresponsiveness and hypersensitivity to analgesics (p<0.01 and p<0.001). Although excretion of u–LTE4 did not increase after the methacholine provocation test, it significantly increased after sulpyrine provocation (p<0.01). Furthermore, after pretreatment with cromolyn sodium, the maximum level of u–LTE4 after the sulpyrine provocation test was significantly lower than in controls (p<0.01). These results support the hypothesis that cysteinyl LT is one of the most important components in the pathogenesis of aspirin–intolerant asthma. Cromolyn sodium improves both hypersensitivity to analgesics, and bronchial hyperresponsiveness in aspirin–intolerant asthma.
Hemopoietic Mechanisms in Allergic Airway Inflammation
Authors: Denburg Judah A. , Inman Mark D. , Sehmi Roma , Uno Masashi , O'Byrne Paul M.
Published: April 2003

The bone marrow actively participates in the production of IgER–positive inflammatory cells (eosinophils, basophils and mast cells), which are typically recruited to tissues in atopic individuals. Understanding the signalling between the tissue and the bone marrow at the molecular level may well open up new avenues of therapy for allergic inflammation.
Characterization of Cogon Grass (Imperata cylindrica) Pollen Extract and Preliminary Analysis of Grass Group 1, 4 and 5 Homologues Using Monoclonal Antibodies to Phleum pratense
Authors: Kumar Lalit , Sridhara Susheela , Singh B.P. , Gangal S.V.
Published: April 2003

Background: Previous studies have established the role of Imperata cylindrica (Ic) pollen in type I allergic disorders. However, no systematic information is available on the allergen composition of Ic pollen extract. Objectives: To characterize the IgE–binding proteins of Ic pollen extract and to detect the presence of grass group 1, 4 and 5 allergen homologues, if any. Methods: Pollen extract of Ic was analyzed by in vivo and in vitro procedures such as intradermal tests (ID), enzyme–linked immunosorbent assay (ELISA), ELISA–inhibition, thin–layer isoelectric focusing (TLIEF), sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS–PAGE) and immunoblotting. Dot blot assay was carried out to check the presence of well–known group 1, 4, and 5 allergen homologues in Ic pollen extract. Results: Out of 303 respiratory allergies patients skin–tested, 27 showed sensitivity to Ic pollen extract. Specific IgE levels were elevated in all 15 serum samples tested. The extract prepared for this study was found to be highly potent since it required only 400 ng of homologous proteins for 50% inhibition of binding in ELISA inhibition assays. TLIEF of Ic pollen extract showed 44 silver–stained bands (pI 3.5–7.0) while SDS–PAGE resolved it into 24 Coomassie–Brilliant–Blue–stained bands (MW 100–10 kD). Immunoblotting with individual patient sera recognized 7 major IgE–binding bands (MW 85, 62, 57, 43, 40, 28 and 16 kD) in Ic pollen extract. A panel of monoclonal antibodies, specific to group 1, 4 and 5 allergens from Phleum pratense pollen extract identified group 5 and group 4 homologues in Ic pollen extract. Conclusion: Ic pollen extract was characterized for the protein profile by TLIEF and SDS–PAGE. IgE reactivity was determined by ELISA and immunoblot. Monoclonal antibodies to group 5 and group 4 allergens reacted weakly showing that this pollen contains group 5 and group 4 homologous allergens.
CD8+ T Cells Are the Effectors of the Contact Dermatitis Induced by Urushiol in Mice and Are Regulated by CD4+ T Cells
Authors: López Carolina B. , Kalergis Alexis M. , Becker María Inés , Garbarino Juan A. , De Ioannes Alfredo E.
Published: April 2003

Background: The exposure of human skin to leaves and branches of litre (Lithraea caustica), a Chilean endemic tree, induces a severe contact dermatitis characterized by swelling and pruritus in susceptible individuals. The allergenic priniciple of litre is 3–pentadecyl (10–enyl) catechol (litreol), which is structurally similar to the allergens isolated from poison oak and poison ivy. All of them belong to a family of compounds named urushiols. As a proelectrophilic allergen, litreol must be intracellularly activated before modifying proteins of individuals exposed to it. As a result, self–peptides derived from litreol–modified intracellular proteins would be presented in the context of class I MHC molecules. We hypothesized that CD8+ T lymphocytes would play a major role during the effector phase of the immune response induced by those modified peptides. In order to test this hypothesis, we investigated the cellular immune response to litreol in Balb/cJ mice. The role of the different lymphocyte subpopulations in this response was assessed by immunodepleting mice of CD4+ or CD8+ T lymphocytes using specific monoclonal antibodies (mAbs). We report the observation that the contact dermatitis induced by litreol has two components: a primary response which does not require TCRαβ+ T cells, and a secondary response mediated mainly by CD8+ T cells and regulated by CD4+ T cells. Our results show that CD8+ lymphocytes play a central role as effectors of the secondary response to litreol. Furthermore, our data suggest that two functionally different CD4+ T subpopulations serve as regulators of the CD8+ T cell function: a CD4+ T helper population sensitive to a low dose of the depleting mAb, and CD4+ T suppressor population which is eliminated only with a high dose of depleting mAb.
Endothelial Cell Apoptosis in Lipopolysaccharide–Induced Lung Injury in Mice
Authors: Fujita Masaki , Kuwano Kazuyoshi , Kunitake Ritsuko , Hagimoto Naoki , Miyazaki Hiroyuki , Kaneko Yumi , Kawasaki Masayuki , Maeyama Takashige , Hara Nobuyuki
Published: April 2003

Background: Studies have shown the importance of apoptosis in vascular injury in vitro. We postulated that apoptosis of the endothelium contributes to vascular injury in vivo and may be involved in acute lung injury. Methods: To test this hypothesis, we investigated the incidence of endothelial cell apoptosis in acute lung injury induced in mice by the administration of lipopolysaccharide (LPS). Male ICR mice were administered LPS (20 mg/kg body weight) intravenously and sacrificed at specified times thereafter. Results: Histologic findings were consistent with acute lung injury which increased with time from 3 to 48 h after injection. Electrophoretic analysis of DNA that was extracted from lung tissue and 3'–end–labeled with digoxigenin demonstrated a fragmentation of DNA starting at 6 h. In situ terminal deoxynucleotidyl transferase–mediated dUTP biotin nick end–labeling (TUNEL) demonstrated DNA strand breaks in the endothelial cells. TUNEL also revealed DNA strand breaks in bronchial and alveolar epithelial cells as well as inflammatory cells in the interstitium. These TUNEL–positive cells appeared 6 h after injection. Electron–microscopic examination of the endothelium strongly suggested the morphological characteristics of apoptosis. Conclusion: Apoptosis was induced by LPS administration in endothelial cells in vivo. A role for such apoptosis is suggested in acute lung injury.
A Species–Specific Monoclonal Antibody to Cynodon dactylon
Authors: Lovborg U. , Baker P.J. , Tovey E.R.
Published: April 2003

Background: It is usually difficult to differentiate between the pollens of different grass species on the basis of their appearance under a microscope, as they often appear similar. Such distinctions are important when interpreting the clinical relevance of pollens in air samples as individuals can differ in their allergic responses to different grass species. As this allergenic distinction occurs at the level of presence and differences of epitopes on the allergens associated with different species, it could be anticipated that species–specific monoclonal antibodies could provide such distinctions between pollens. Method: Monoclonal antibodies raised against Cynodon dactylon were screened and characterised in ELISA assays and blotting, using a range of grass pollen extracts, to identify clones which were species specific. Results: The most specific monoclonal raised to C. dactylon did not react at a level of greater than 1.2% to extracts of 10 other grass pollens in a direct ELISA assay and showed no detectable cross–reactivity in a particle blotting assay. Conclusion: It has been possible to produce a monoclonal antibody that is functionally species specific to C. dactylon.
Antigen Interaction and Heat Inactivation Expose New Epitopes on Human IgE
Authors: Pachlopnik Jana M. , Stämpfli Martin R. , Rudolf Michael P. , Aebischer Iwan , Kricek Franz , Miescher Sylvia , Stadler Beda M.
Published: April 2003

It is well established that heat–denatured IgE is no longer capable of binding to FcεRI. We have found an antibody that interacts with heat–denatured IgE. Interestingly, this antibody can also be used to detect some serum IgE, but not IgE synthesized de novo in vitro. However, native IgE can be transformed into an IgE that is recognized by this antibody, if antigen is added. Our data indicate that physiological mechanisms exist that biologically inactivate IgE which might still be mistaken for 'functional' IgE by assays based on polyclonal antibodies.
Granulocyte/Macrophage–Colony Stimulating Factor in Allergen–Induced Rhinitis: Cellular Localization, Relation to Tissue Eosinophilia and Influence of Topical Corticosteroid
Authors: Nouri-Aria K.T. , Masuyama K. , Jacobson M.R. , Rak S. , Lowhagen O. , Schotman E. , Hamid Q. , Durham S.R.
Published: April 2003

Background: Allergen–induced late nasal responses are associated with recruitment of T lymphocytes and eosinophils, and preferential messenger RNA (mRNA) expression of 'TH2–type' cytokines. We previously showed that topical steroid inhibited the late response and associated tissue eosinophilia. In this study we tested the hypothesis that granulocyte/macrophage–colony stimulating factor (GM–CSF) may contribute to late–responses and tissue eosinophilia and is inhibitable by topical corticosteroid. Methods: Nasal biopsies were taken before and 24 h after nasal allergen provocation following 6 weeks of treatment with either a nasal corticosteroid spray (fluticasone propionate) or a matched placebo nasal spray twice daily. Cryostat sections were processed by immunohistochemistry and in situ hybridization to assess cytokine mRNA expression for GM–CSF. Results: Increases in T lymphocytes and eosinophils were seen in the nasal mucosa after allergen challenge (p = 0.01) which were accompanied by a 5–fold increase in cells expressing mRNA for GM–CSF (p = 0.01). Double immunohistochemistry/in situ hybridization demonstrated that the majority of GM–CSF mRNA+ cells were co–localized to CD68+ (40%), or T cells (40%) with a lesser contribution from eosinophils (<20%). Topical steroid treatment was accompanied by a decrease in both the CD3+ and major basic protein (MBP+) cells expressing GM–CSF mRNA (p = 0.01) with a corresponding proportionate increase in the % of macrophages expressing GM–CSF. Conclusions: The results indicate that after allergen provocation, eosinophils are recruited to the nasal mucosa and that, at least in part, this may be due to GM–CSF. Topical nasal corticosteroid inhibits late responses and the associated eosinophilia, possibly indirectly by decreasing GM–CSF from T lymphocytes or reducing autocrine production of GM–CSF from eosinophils.
Effect of IL–4, IFN–Á and IL–12 on Cytokine Production from Human CD45RA and CD45RO CD4 T Cell Precursors
Authors: Sasama J. , Vyas B. , Vukmanovic-Stejic M. , Kemeny D.M.
Published: April 2003

The aim of this study was to compare the effects of IL–4, IL–12 and IFN–γ on the production of T helper–1 (Th1) and T helper–2 (Th2)–type cytokines from human peripheral blood 'naive' CD45RA and 'memory' CD45RO CD4 T cells. CD45RA or CD45RO CD4 T cells were cultured for 4 days with phorbol myristate acetate (PMA) and ionomycin and either IL–4, IFN–γ or IL–12 and their ability to proliferate and secrete IFN–γ and IL–4 determined. Purified CD45RO CD4 T cells stimulated with PMA and ionomycin secreted higher levels of IL–4 and IFN–γ, as measured by ELISA, than CD45RA CD4 T cells which secreted little IL–4 or IFN–γ. However, CD45RA and CD45RO CD4 T cells proliferated to the same extent and IL–4, IFN–γ and IL–12 had no effect on this. IL–12 and IFN–γ had no effect on the amount of IL–4 secreted by PMA and ionomycin–stimulated CD45RO CD4 T cells, but culture with IL–4 enhanced IL–4 production 7–fold. IL–12 increased the amount of IFN–γ produced by CD45RO CD4 T cells 2– to 3–fold. Small amounts of IFN–γ production were induced in CD4 CD45RA T cells by IL–12 and IFN–γ. These results indicate: (1) that CD45RA cells cannot make significant amounts of IL–4 under the conditions used, (2) that CD45RO cells can produce both Th1 and Th2 cytokines immediately upon restimulation, (3) that IL–12 favours Th1 cytokine production in both CD45RA and CD45RO CD4 T cells, and (4) that IFN–γ favours IFN–γ production in CD45RA but not CD45RO cells.
In situ Localization of Pollen Allergens by Immunogold Electron Microscopy: Allergens at Unexpected Sites
Authors: Grote Monika
Published: April 2003

The windborne pollen grains of many trees and grasses contain a number of highly water–soluble (glyco)proteins which upon moistening, e.g. on the human respiratory mucosa, rapidly diffuse out of the pollen grain. In susceptible individuals, they may cause allergic reactions. Because of their rapid release from the pollen, pollen allergens were expected to be located in the outermost layers of the pollen grain, i.e. on the surface and in the wall (exine, intine). First attempts to localize allergens by microscopic methods in the pollen grain supported this view. However, since conventional preparation methods were used, artificial mobilization of allergens could not be excluded. Based on new, completely anhydrous preparation protocols and on the immunogold–labeling technique, pollen allergens were electron–microscopically shown to be located in the interior of the pollen grain, i.e. in the cytoplasm, often within ribosome–rich areas. This unexpected result was obtained in all strictly anhydrously prepared pollen species. The biological function of these allergens is largely unknown, however, clues as to possible cellular functions have been obtained for the birch pollen major allergen Bet v 1.
Molecular Analysis of the Group 1 and 2 Allergens from the House Dust Mite, Euroglyphus maynei
Authors: Smith W. , Mills K.L. , Hazell L.A. , Hart B.J. , Thomas W.R.
Published: April 2003

Background: There is increasing evidence that the house dust mite Euroglyphus maynei may be a significant source of allergic sensitization. The structural information for the E. maynei allergens is largely restricted to a single partial genomic sequence of Eur m 1. Methods: A cDNA library was constructed from a culture of E. maynei. Clones encoding the major group 1 and 2 allergens were isolated by DNA hybridization and sequenced. Results: The sequence of several full length clones of Eur m 1 and Eur m 2 were obtained. The full pre–proenzyme sequence of the cysteine protease Eur m 1 was determined. The translated amino acid sequence of Eur m 1 and Eur m 2 had 84–86% sequence identity with the corresponding allergens from Dermatophagoides pteronyssinus and Dermatophagoides farinae mites. This is the same as the degree of sequence identity found between D. pteronyssinus and D. farinae despite Euroglyphus being a member of the Pyroglyphinae subfamily rather than the Dermatophagoidinae subfamily. Conclusion: The sequences of the major Eur m 1 and Eur m 2 allergens are described. Their degree of divergence from the Dermatophagoides spp. is similar to that observed between D. pteronyssinus and D. farinae group 1 and group 2 allergens.
Protein Tyrosine Phosphorylation: A Possible Common Signaling Pathway in Human Th1 and Th2 Cell Clones
Authors: Minoguchi Kenji , Yamashita Naomi , Oda Naruhito , Takeno Mitsuhiro , Kaneoka Hidetoshi , Sakane Tsuyoshi
Published: April 2003

Background: It has been reported that protein tyrosine phosphorylation is essential for cytokine production in mouse Th1 cells but not in Th2 cells. However, little is known about the difference of signal transduction between human antigen–specific Th1 and Th2 cell clones. Methods: Purified protein derivative–specific Th1 and Dermatophagoides farinae–specific Th2 cell clones were established. T cell clones were stimulated with anti–CD3 Abs and further treated with goat antimouse immunoglobulin Abs for cross–linking of TCR/CD3 complex. Results: In contrast to murine Th2 clones, tyrosine phosphorylation including both ZAP–70 and phospholipase–C–Ál was necessary for cell activation in both types of human T cell clones. This was further supported by the observation that genistein (protein tyrosine kinase inhibitor) inhibits IFN–Á production for Th1 and IL–4 for Th2 in a dose–dependent manner. Conclusion: Protein tyrosine phosphorylation is thus an essential component in transducing the activation signal from TCR–CD3 complex both in human Th1 and Th2 cells.