Ikuo Akutsu

Clinical significance of measuring levels of sputum and serum ECP and serum IL-5 in bronchial asthma.

Although the association of asthma and eosinophils had been well-known for many years (1, 10,40,46), the function of the eosinophils in asthma was not clear until the middle of the 1980s. Several in vitro experimental results suggest the unfavorable role of eosinophils for asthma (23), i.e., the induction of damage to the respiratory epithelium with eosinophil granule basic proteins such as major basic protein (MBP), eosinophil peroxidase (EPO), and eosinophi1 cationic protein (ECP) (1 1 , 27) resulting in development of airway hyperresponsiveness (8,9), the mast cell degranulation with MBP, ECP, and EPO (18, 33, 54), the release of leukotriene C4 (45, 52) and platelet activating factor (2 1) with bronchoconstrictive properties, and the induction of airway hyperresponsiveness with direct instillation of MBP to the rat trachea (5,43). Recently much attention has been focused on the interaction between eosinophils and T lymphocytes. For example, lymphocytes migrate into the bronchial mucosa of asthma patients (20), and the number correlates with that of eosinophils (35). Among the lymphocytes migrating into the bronchial mucosa in asthma, CD4 + lymphocytes predominate with a marker of activation (13,30), and correlate with the number ofeosinophils in the mucosa (2, 16). In the study utilizing bronchoalveolar lavage fluids evels IL-5 of in

Inhibition of Antigen-Induced Late Asthmatic Response and Bronchial Hyperresponsiveness by Cyclosporin and FK 506

Using a guinea pig model of asthma, we have shown that the administration of cyclosporin, a T-lymphocyte-selective immunosuppressive agent, from the beginning of the immunization period inhibits the development of the late asthmatic response and bronchial hyperresponsiveness after antigen challenge. Similar results were obtained with FK 506, a new potent immunosuppressive agent. Since these compounds have been shown to suppress the activation of guinea pig T lymphocytes, the present data suggest that T lymphocytes may be important for the elicitation of the late asthmatic response and bronchial hyperresponsiveness.

Detection of interleukin-5 messenger RNA and interleukin-5 protein in bronchial biopsies from asthma by nonradioactive in situ hybridization and immunohistochemistry☆☆☆★★★

Recently direct evidence for the role of interleukin-5 (IL-5) in eosinophilic inflammation in the airways of persons with asthma has been provided by an in situ hybridization study that used radioisotope-labeled IL-5 complementary RNA probes. Radioisotope-labeled probes, although sensitive, require autoradiographic detection, which is time-consuming. In the most recent study we attempted to detect IL-5 messenger RNA in the bronchial biopsy specimens from patients with asthma using nonradioactive in situ hybridization, which gives rapid results. Bronchial biopsy specimens were obtained from eight patients with asthma and seven diseased control subjects. IL-5 complementary DNA probes were labeled with digoxigenin-deoxyuridine triphosphate and hybridized to permeabilized sections. Hybridization signals were visualized by an immunohistochemistry technique. Positive hybridization signals were observed in six of the eight biopsy specimens from patients with asthma. Pretreatment with ribonuclease or hybridization with an unrelated probe produced negative results. Immunohistochemical staining of serial sections with a monoclonal antibody to IL-5 revealed that a few cells within the mucosa positively stained, suggesting active synthesis of IL-5. Biopsy results from the seven diseased control subjects did not show any hybridization signal. These results confirm and extend previous observations of IL-5 messenger RNA expression in the airways of patients with asthma, and suggest that digoxigenin-labeled IL-5 complementary DNA probes would be a powerful research tool.

151 In vitro model of eosinophil migration into the site of acute allergic reactions

PEROXIDE GENERATION BY EOSINOPHIS IN ALLERGIC RHINITIS. H.Oqasawara,M.D.,S.Yoshimura, M.D. and T.Kumoi,M.D.,Nishinomiya,Japan We previously showed that H202 which could be produced by eosinophils or neutrophils from peripheral blood itself induced histamine release ,and that low concentration of H202 augmented anti-IgE or antigen mediated histamine release, while high concentration of H202 which were not cytotoxic inhibited anti-IgE or antigen mediated histamine release. The function of eosinophils in nasal secretions or peripheral blood in house dust nasal allergic patients was studied. Nasal secretion were collected, soon the patient were challenged with house dust, and 5,15 and 30 minutes later respectively,additional nasal secretion were collected. Leukocytes were isolated using five different concentrations(50,56,64,68 and 72%)of Percoll solution from heparinized venous blood. After nasal challenge,the population of eosinophils was increased with various times elapsed to 47,64,65 and 73%,the concentration of H202 in nasal secretion was B&739,728 and 697 pmol/g,the activity of peroxidase was 0.90,0.65,1.67 and 0.91 microgram/g respectively, and the activity of arylsulfatase was not change during this time. Eosinophi chemotactic activity was seen in 5 minute nasal secretion. The generation of H202 increased with time elapsed;stimulation with zymosan enhanced the H202 generation. The rate of phagocytosis for zymosan in eosinophils was not changed. In the high IgE group, oxidative metabolic function in eosinophils from peripheral blood was higher than in the low IgE group. Eosinophil chemotactic activity seemed to stimulate oxidative metabolic function. H202 plays a significant role in vivo was suggested.

Wasp venom allergy: effect of anti-IgE antibody on wasp venom anaphylaxis in a mouse model.

BACKGROUND Although anti-IgE antibody (Ab) therapy was recently shown to be effective in patients with bronchial asthma, no study has reported the effect of IgE therapy in the prevention of wasp venom anaphylaxis. In this study, we used a mouse model of wasp venom allergy to investigate the effect of anti-IgE Ab on wasp venom anaphylaxis. METHODS We developed a mouse model of wasp venom allergy by intraperitoneally (i.p.) injecting wasp venom into BALB/c mice twice on experimental day (day) 0 and 7. On day 20, a group of mice received an i.p. injection of mouse anti-IgE Ab as a pretreatment, and another group received rat anti-IgG1 Ab. On day 21, the animals were challenged by i.p. injection of wasp venom, and 30 min later, body temperature was measured and serum levels of leukotriene (LT) B4 and LTC4 were determined using enzyme immunoassay. RESULTS The body temperature of mice treated with anti-IgE Ab and controls before and after wasp venom challenge was 37.8±0.2 vs 37.7± 0.3°C before challenge and 37.8±0.2 vs 37.1± 0.3°C after challenge, respectively, showing that anti-IgE Ab treatment significantly prevented body temperature from falling (p <0.05). Furthermore, anti-IgE Ab treatment reduced total serum IgE levels in the treated mice (42.2±15.9 pg/ml), compared with controls (105.9±23.1 pg/ml, p <0.05), and inhibited the secretion of LTC4 in the treated mice (32.0±18.8 pg/ml), but not in the controls (162.4±12.4 pg/ml, p <0.05), following challenge with wasp venom. CONCLUSION The results of the present study indicate that anti-IgE Ab treatment is an effective preventive measure against wasp venom-induced anaphylaxis.

841 Eosinophil chemotactic activity in the supernatant of mononuclear cell culture stimulated with specific antigen

Peripheral blood mononuclear cells (PBMC) separated from patients with asthma who were sensitive to Dermatophagoides farinae (Df) were cultured in alpha-medium for 5 days at 37 degrees C in 5% CO2, in the presence or absence of 10 ng/ml of Df antigen. Eosinophils were purified from the peripheral blood of patients with eosinophilia who were not sensitive to Df. Eosinophil chemotactic activity (ECA) was tested using a modified Boyden chamber method. ECA in the supernatant of PBMC stimulated with Df antigen was detectable after 24 hrs, peaked at 72 hrs and continued throughout the experiment. ECA was not observed in the supernatant of PBMC culture from subjects who were not sensitive to Df, and negligible activity was also observed when PBMC were stimulated with an unrelated antigen. The activity was unchanged by heating at 56 degrees C for 30 min, but was inactivated at 100 degrees C for 10 min. CV-6209, a specific PAF antagonist, failed to inhibit this chemotactic activity. The molecular weight of this eosinophil chemotactic factor (ECF) was greater than 30,000 daltons as determined by an ultrafiltration study. In conclusion, these data suggest that in asthmatic patients sensitive to Dermatophagoides farinae mononuclear cells stimulated with a related antigen produce one of cytokine(s) which possess(es) ECA, and may play an important role in the recruitment of eosinophils in chronic asthma.