Recent advance in investigation of gene polymorphisms in Japanese patients with aspirin-exacerbated respiratory disease.

Aspirin-exacerbated respiratory disease (AERD) is a complex clinical syndrome characterised by severe asthmatic attack upon treatment with aspirin and/or non-steroidal anti-inflammatory drugs (NSAIDs). Genetic predisposition has been considered as a crucial determinant and candidate genes have concentrated especially on cysteinyl leukotrienes (LTs)-related genes as the inhibitory action of aspirin and NSAIDs on cyclooxygenase activity may cause overproduction of cysteinyl LTs. However, conflicting results have been reported, in parallel with replication studies in different ethnic groups. Thus, future areas of investigations need to focus on comprehensive approaches towards the discovery of other genetic biomarkers. Unfortunately, few papers have been reported about gene polymorphisms in Japanese patients with AERD. Here, we described on our recent genetic investigations on B2ADR, IL-13, IL-17A, CYP2C19, TBXA2R, CRTH2 and HSP70. This review indicates potential genetic biomarkers contributing to the early diagnosis of AERD, which may include CYP2C19 and HSP70 gene polymorphisms, and future validation studies in independent population are required to provide reassurance about our findings.

Characterization of platelet-activating factor-induced elevation of cytosolic free calcium concentration in eosinophils.

In order to evaluate the role of calcium in the activation processes in eosinophils induced by platelet-activating factor (PAF), we investigated the changes in free cytoplasmatic Ca2+ concentration using fura-2. PAF causes a rapid and transitory rise of the intracellular free calcium ion concentration [( Ca2+]i) in purified guinea pig eosinophils of approx. 1000 nM above a basal level of 120.7 +/- 36.5 nM (n = 10). The effect was dose-related with a maximum rise at 1000 nM PAF and an EC50 of 17.4 nM and specifically inhibited by the PAF antagonist WEB 2086 with an IC50 of 95.5 nM. WEB 2086 did not affect either the leukotriene B4- or the fMet-Leu-Phe-induced elevation of [Ca2+]i. The response to PAF was dependent on external Ca2+ as it was significantly inhibited by EGTA (85.6 +/- 5.4%) and Ni2+ (95.8 +/- 2.1%) but not by the dihydropyridine antagonist nimodipine. We conclude that Ca2+ entry via receptor-operated Ca2+ channels may be involved in PAF-induced degranulation of eosinophils.

Effect of YM264 on the airway hyperresponsiveness and the late asthmatic response in a guinea pig model of asthma.

We investigated the effects of YM264, a specific platelet-activating factor (PAF) antagonist, on the airway hyperresponsiveness (AH) and the late asthmatic response (LAR) of guinea pigs that were sensitized by exposure to aerosolized ovalbumin (OA). Respiratory resistance (Rrs) was determined by the oscillation technique. Airway responsiveness was evaluated by administering a dose of histamine at which the Rrs reached 200% of the baseline value (H200). Animals were administered 1 or 3 mg/kg of YM264 orally 30 min before and again at 3 h after exposure to OA. YM264 significantly suppressed AH 24 h after and 5 days after the exposure. YM264 also suppressed the development of the LAR and accumulation of eosinophils and neutrophils in the tracheal mucosa of guinea pigs. These observations suggest that PAF is involved in the AH and the development of the LAR in asthma. PAF antagonists may play a beneficial role in the treatment of asthma.

Inhibitory action of acyclovir (ACV) and penciclovir (PCV) on plaque formation and partial cross-resistance of ACV-resistant varicella-zoster virus to PCV.

Penciclovir has potent antiviral activity against varicella-zoster virus (VZV). We have characterized the inhibitory effects of penciclovir and acyclovir on the plaque formation of cell-free VZV and cross-resistance of acyclovir-resistant VZV to penciclovir. The apparent effective concentration for 50% plaque reduction (EC50) of penciclovir determined on the third day was significantly lower than that determined on the fourth or fifth day. The size of plaques was smaller in the presence of penciclovir than in the presence of acyclovir. The effective concentrations for 50% reduction of the number of infected cells per plaque were 1.40 and 5.00 micrograms/ml for penciclovir and acyclovir, respectively. Thus penciclovir suppressed spread of infection within developing plaques more efficiently than acyclovir. Five acyclovir-resistant VZV strains with altered DNA polymerase selected by acyclovir were examined for cross-resistance to penciclovir. They were 11- to 18-fold more resistant to ACV than the parent strain, but only 4- to 5-fold more resistant to PCV. Penciclovir-triphosphate carrying the 3'-hydroxyl group of 2'-deoxyribose might have better affinity to the altered viral DNA polymerase than acyclovir-triphosphate without the 3'-hydroxyl group.

Prophylactic treatment of cytomegalovirus infection with traditional herbs.

Hot water extracts of four traditional herbs, Geum japonicum, Syzygium aromaticum, Terminalia chebula and Rhus javanica, which have been shown to have anti-herpes simplex virus (HSV) activity in vivo, were examined for anti-cytomegalovirus (CMV) activity in vitro and in vivo in this study. They inhibited replication of human CMV and murine CMV (MCMV) in vitro. These anti-CMV activities in vivo were examined in an MCMV infection model using immunosuppressed mice. Mice were subcutaneously treated with various doses of cyclosporine, and immunosuppression and MCMV infection were monitored by suppression of antibody production and virus yield in the lung, respectively. Each herbal extract was orally administered to mice treated with 50 mg/kg of cyclosporine from a day before intraperitoneal infection, and the efficacy of herbs was evaluated by the reduction in the virus yield in the lung. Among them Geum japonicum, Syzygium aromaticum, and Terminalia chebula significantly suppressed MCMV yields in lungs of treated mice compared with water treatment. Efficacy of oral treatment with 750 mg/kg per day of Geum japonicum extract was similar to that of the intraperitoneal administration of 2 mg/kg per day of ganciclovir in increasing the body weight of infected mice and reducing the virus yield in the lungs. These herbs may be beneficial for the prophylaxis of CMV diseases in immunocompromised patients.

Lipogenic and lipolytic activities in isolated adipocytes from cattle with fat necrosis.

The metabolic activity and cellularity of adipocytes isolated from the abdominal adipose tissue of normal heifers and heifers with fat necrosis were compared. The basal rate of U-14C glucose incorporation into total lipids in adipocytes from the periphery of the necrotic mass was higher than that in the colonic mesentery of both the affected and normal heifers. In the affected animals. adipocytes from the mesentery of the spiral colon and adipocytes from the periphery of the necrotic mass failed significantly to increase the incorporation of labelled acetate and glucose, respectively, in response to insulin. In the presence of adrenalin, adipocytes from the colonic mesentery and the periphery of the necrotic mass of the affected heifers released more glycerol than adipocytes from the colonic mesentery of normal animals. In addition, the mean diameters of adipocytes from the colonic mesentery and the periphery of the necrotic mass of the affected heifers were significantly greater than those from the colonic mesentery of normal animals. These results indicate that excessive fattiness in abdominal adipose tissue may predispose cattle to fat necrosis.

[Effects of cefoperazone on respiratory infections of patients of advanced age and/or with underlying respiratory diseases].

Respiratory infections of 19 subjects of advanced age and/or with underlying respiratory disease were treated with cefoperazone (CPZ) and its clinical effects were studied. Sixteen subjects suffered from respiratory tract infection and 3 subjects had pneumonia. The age of the subjects ranged from 39 to 77 years with the mean of 63.8, 7 of them being more than 70 years of age. The underlying respiratory diseases included chronic pulmonary emphysema in 6 subjects, diffuse panbronchiolitis in 3, bronchiectasis in 3, silicosis in 2 and one each of chronic bronchitis, pulmonary fibrosis, lung cancer and old pulmonary tuberculosis. One case, 75 years of age, had renal insufficiency. The daily dose of CPZ was 4 grams in 18 of the 19 subjects and the duration of administration ranged 5 to 22 days. The remaining 1 subject received 2 g of CPZ daily for 6 days. Clinical effects were judged from the changes in fever, cough, amount of sputum, dyspnea, rales, cyanosis, chest X-ray, white blood cell counts, CRP, erythrocyte sedimentation rates and results of sputum culture. Clinical effects were good in 16 subjects, fair in 1, and poor in 2. Bacteriological follow-up was carried out in 13 subjects. Infecting bacteria were eliminated from 5 subjects, reduced in 2 and, in 4 subjects, they were replaced by other bacteria. In 1 subject, P. aeruginosa was isolated from sputum even after the treatment with CPZ, and in another subject H. influenzae relapsed immediately after the cessation of the CPZ treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of inhaled cyclosporin A on the allergen-induced late asthmatic response and increased in airway hyperresponsiveness in a guinea pig model of asthma.

Oral administration of cyclosporin (CsA), a potent inhibitor of helper T cell function, prevents the allergen-induced late asthmatic response (LAR) and the increase in airway hyperresponsiveness (AH) seen in actively sensitized guinea pigs. The systemic administration of this agent in humans has been associated with serious side effect, therefore, the effects of inhaled CsA were therefore examined in guinea pigs that were actively sensitized by repeated exposure to nebulized ovalbumin. Respiratory resistance (Rrs) of the animals was measured by an oscillation method and the extent of AH was inferred from the inhaled concentration of histamine required to increase Rrs by 200%. The magnitude of ovalbumin-induced immediate bronchoconstriction after sensitization was similar in CsA-treated and nontreated control animals. However, a LAR was observed in 4/5 control animals but in 0/5 CsA-treated animals. The increase in AH observed 24 hours after antigen exposure in control animals was significantly inhibited by prior CsA inhalation. Significant CsA concentrations were detected by radioimmunoassay in the lungs of CsA-treated animals. Thus, inhaled CsA should be further investigated because it may be useful treating asthma while avoiding side effects.

[Effect of BAY u3405-thromboxane A2 receptor antagonist, on biphasic airway responses induced by platelet-activating factor in actively sensitized guinea pigs].

Our previous study in activity sensitized guinea pigs demonstrated an LAR-like increase in respiratory resistance (Rrs) at 3 to 9 hr after PAF inhalation. The result suggested possible involvement of the priming effect of active sensitization and PAF. Mean while, thromboxane A2 (TXA2) is known to be induced by PAF. The present study investigated the involvement of TXA2 in the guinea pig LAR model with a new TXA2 receptor antagonist, BAY u3405. One hr after BAY u3405 administration to guinea pigs sensitized by ovalbumin, the Rrs following inhalation of PAF was subsequently determined. Infiltration of inflammatory cells in the airway tissue 9 hr after PAF inhalation was also observed. While a re-increase in Rrs was found in all the cases in the control group, the re-increase in Rrs was inhibited significantly in the BAY u3405 administration group, 4 to 9 hr after PAF inhalation. The numbers of eosinophils and lymphocytes in the airway tissue were significantly decreased in the BAY u3405 administration group, as compared with the control group. From these results, the possibility is suggested that TXA2 and its direct effect on the airway and the migration-enhancing effect on eosinophils and T lymphocytes, as well as PFA, are involved in the development of LAR by PAF.

[Effect of capsaicin on the migration of eosinophils into the bronchi of guinea pigs].

Substance P (SP), a potent neuropeptide, which is localized in the sensory nerves and released by many physiological stimuli has been implicated in airway neurogenic inflammation. We have studied the effects of capsaicin (CAP), which releases tachykinins (TK) from the sensory nerves, on eosinophil (EOS) recruitment in the airway in guinea pigs in vivo. Male guinea pigs were used. The respiratory resistance (Rrs) of the guinea pigs were measured by an oscillation technique and histological studies of the right main bronchus were carried out. Exposure to inhaled CAP resulted in a significant increase in Rrs with PC200 CAP of 0.97 +/- 0.25 (x 10(-6) M) (n = 5). This stimulation also provoked striking eosinophilia in the right bronchus in a dose-dependent manner. A neutral endopeptidase (NEP) inhibitor, phosphoramidon, potentiated CAP-induced EOS infiltration. By contrast, pretreatment with [D-Pro2, D-Trp7,9]-SP, an analogue of SP and its receptor antagonist, diminished the response. We conclude that CAP-induced tachykinin release is capable of causing striking eosinophilia in the lung in vivo. This mechanism may contribute to airway inflammation in patients with asthma. This would provide further support for a link between tachykinin and bronchial eosinophilia in asthma.

[Involvement of platelet activating factor (PAF) in ovalbumin antigen-induced late asthmatic response and increase of airway hyperresponsiveness in a guinea pig experimental model of asthma].

Platelet activating factor, a potent chemical mediator, has been implicated in the pathogenesis of asthma in terms of inflammatory cell recruitment and activation. We have recently demonstrated that repeated antigen (ovalbumin; OA) exposure by inhalation to guinea pigs results in a development of late asthmatic response (LAR) in more than 50% of the animals and significant increase in airway hyperresponsiveness (AH). We have studied the effect of WEB 2086, a specific PAF receptor-antagonist, on this model. Respiratoly resistance (Res) of guinea pigs was measured by a oscillation technique and AH was evaluated by the provocative concentration of aerosols of histamine causing 200% increase of Rrs over the baseline Rrs (PC200 Hist). Four out of 5 actively sensitized and diphenhydramine-pretreated animals developed LAR 3 to 9 hr after allergen (20 mg/ml OA, 10 min inhalation)-induced immediate bronchoconstriction (LAR). Treatment with WEB 2086 (3 mg/kg intravenously) 30 min before and 3 hr after the exposure suppressed LAR clearly without affecting the IAR. Significant increase in AH from 2.80 +/- 0.03 to 2.51 +/- 0.01 and 2.60 +/- 0.08 (p less than 0.05, n = 8) of PC200 Hist (mg/ml, log) was observed 24 hr and 5 day after the OA exposure, respectively. The WEB 2086 treatment also prevented the increase of AH after the OA exposure (PC200 Hist; 2.82 +/- 0.09 before the challenge 2.80 +/- 0.07 and 2.75 +/- 0.09 24 hr and 5 days after, respectively. n = 8). Administration of WEB 2086 did not affect baseline Rrs and PC200 Hist in normal guinea pigs without any antigen challenge. We conclude that WEB 2086 is capable of preventing the development of LAR and increase in AH, and thus PAF may play an important causal role in LAR and increased AH observed in asthma.

[Platelet activating factor (PAF)-induced late asthmatic response in sensitized but not in non-sensitized guinea pigs].

We have recently demonstrated that pretreatment with WEB 2086, a specific PAF antagonist or cyclosporine A (CsA), a potent helper T cell suppressant, resulted in preventing the development of late asthmatic response (LAR) and increase of airway hyperreactivity (AH) in guinea pig experimental models of asthma. We have now examined whether exogenously applied PAF causes LAR in these models in vivo. The respiratory resistance (Rrs) of guinea pigs was measured by an oscillation technique and histological studies of the bronchi were also made. Guinea pigs, actively sensitized by repeated antigen (ovalbumin; OA) inhalation, showed a leftward shift of the inhaled PAF dose response curve of Rrs compared with that in control animals, indicating that the sensitized animals were more sensitive to inhaled, PAF. PC200 PAF, which indicate provocative concentrations of PAF aerosols causing a 200% increase in the baseline Rrs, were 3800 +/- 604.9 micrograms/ml and 780 +/- 94.3 micrograms/ml, in the control and sensitized animals, respectively. The same magnitude of immediate bronchoconstriction after 780 and 3800 micrograms/ml of PAF exposure was observed in the actively sensitized and non-sensitized control animals, respectively. However, LAR developed 4 out of 6 animals only in the sensitized guinea pigs. We conclude that both exogenously applied PAF by inhalation and antigen exposure are capable of inducing LAR in sensitized guinea pigs, and thus the priming effect of immunization and PAF may contribute to the development of LAR observed in asthma.

[Effect of rush immunotherapy (RIT) on Hymenoptera allergy].

In our country approximately forty people die every year from anaphylaxis caused by hymenoptera stings. Between 1988 and 1996, 48 patients, who had experienced a systemic reaction to hymenoptera sting and were proved to have specific IgE antibodies to wasp, yellow or both (RAST score > or = 2), received rush immunotherapy (RIT) using venom extracts in our hospital. Fifteen patients had re-sting after RIT. Fourteen out of the 15 patients showed only local reaction to the hymenoptera re-sting and one patient had mild generalized symptoms. Although one patient showed mild generalized uriticaria during RIT, no adverse reaction occurred during and after RIT in the other subjects. Follow-up studies on the titers of serum total IgE antibodies and hymenoptera specific IgE and IgG4 antibodies revealed that total and specific IgE antibodies transiently increased one month after RIT and returned to their baseline values by 6 months after RIT, while specific IgG4 antibodies continued to gradually increase up to al least 3 years after RIT. These results demonstrates that RIT is effective in prevention of a systemic reaction to hymenoptera re-sting and an increase in the titer of hymenoptera specific IgG4 antibodies may at least partly explain the efficacy of RIT.

[Clinical and experimental study of the host defense mechanism from the aspect of tumor immunology].

The host defense mechanism was fundamentally and clinically investigated from the aspect of tumor immunology. The behavior of macrophages in the immune systems of experimental animals demonstrated different patterns of response in the bone marrow, peritoneal cavity and spleen, respectively. It was doubtless that the spleen occasionally exerted an immunosuppressive action. Ectopic immunization of syngeneic rats with normal liver cells resulted in specific inhibition of the growth of the tumor cells (AH-I30) derived from liver. This seemed to strongly suggest a local immune response. In clinical cases, the host defense mechanism was found to collapse due to surgical intervention, malnutrition and immunosuppressive substances. The relation between complements and host defense mechanism has been controversial. Our studies indicated the serum complement level to be the most reliable parameter for understanding the host defense mechanism. Since the combination of immunostimulants and anticancer drugs is likely to induce anticancer drugs to exert an adverse effect depending on the time of such combination, timing of combined administration of drugs should be chosen with great care.

[Eosinophil-derived collagenase (metalloproteinase)].

Collagenase is a highly specific neutral protease which acts by cleaving the collagen molecule into fragments, at a site three-quarters of the distance from the amino terminus. Collagenase has been assumed to be connected with the destruction of the pathological connective tissue which accompanies inflammatory conditions such as rheumatoid arthritis and interstitial lung disease. Despite the association of eosinophils with wound-healing and fibrotic processes, their collagenolytic ability has been poorly defined and it was not until 1984 that human eosinophils were shown to contain an enzymatic activity which degrades collagen. Eosinophils contain a metalloprotein that degrades types I and III collagens and no collagenolytic activity against types IV and V collagens is detected. Some experiments suggested that the eosinophil collagenase was different from neutrophil collagenase which cleaves type I collagen preferentially. Eosinophils may play a role in the alterations in connective-tissue matrices seen in physiological and pathological states.

[Cytomegalovirus infection and its possible treatment with herbal medicines].

Medicinal herbs, Geum japonicum, Syzygium aromaticum, Terminalia chebula, and Rhus javanica, with anti-herpes simplex virus therapeutic activity, inhibited replication of human cytomegalovirus(CMV) and murine CMV(MCMV) in vitro. These anti-CMV activities were examined in an MCMV infection model using immunosuppressed mice. Geum japonicum, Syzygium aromaticum, and Terminalia chebula significantly suppressed MCMV yields in lungs of treated mice compared with water treatment. Efficacy of oral treatment with 750 mg/kg/day of Geum japonicum-extract was similar to that of the intraperitoneal administration with 2 mg/kg/day of ganciclovir in increasing the body weight of infected mice and reducing the virus yield in the lungs. These herbs may be beneficial for the prophylaxis of CMV diseases in immunocompromized patients.