A cyclosporin A/maltosyl-alpha-cyclodextrin complex for inhalation therapy of asthma.

The inhalation of cyclosporin (Cs)A to the lung is limited by its hydrophobic properties. In order to improve the poor solubility of CsA, cyclodextrin (CD) was evaluated for its suitability for dry powder inhaler formation, and the benefit of an inhaled CsA/CD complex in vivo was demonstrated. The solubilising effect of CDs on CsA was measured by high-performance liquid chromatography. Ciliostatic activity and haemolysis were determined to assess some safety profiles of CDs. The efficacy of an inhaled CsA/CD complex was evaluated by eosinophil infiltration into the bronchoalveolar lavage fluid in actively sensitised mice. CDs markedly improved the poor solubility of CsA. The ciliostatic and haemolytic activities of maltosyl-alpha-CD were the weakest of all the tested CDs. CsA inhaled alone showed inhibitory effects on allergen-induced eosinophilia. Inhalation of the complex of CsA with maltosyl-alpha-CD, where the dose of CsA was approximately nine-times less than that of CsA inhaled alone, also inhibited eosinophil accumulation significantly, with a longer duration of action in comparison with the response to CsA alone. Thus the effective dose of cyclosporin A could be reduced by formation of a complex with maltosyl-alpha-cyclodextrin, and a wider therapeutic safety margin by inhalation of cyclosporin A as a complex with maltosyl-alpha-cyclodextrin could be expected.

Effect of a novel bifunctional endothelin receptor antagonist, IRL 3630A, on guinea pig respiratory mechanics.

This study characterized the in vitro pharmacological properties of a newly developed endothelin receptor antagonist, N-butanesulfonyl-[N-(3, 5-dimethylbenzoyl)-N-methyl-3-[4-(5-isoxazolyl)-phenyl]-(D)- alanyl]-( L)-valineamide sodium salt (IRL 3630A), and its in vivo effects on respiratory mechanics were determined. IRL 3630A showed highly balanced affinities to human endothelin ET(A) and ET(B) receptors, giving apparent K(i) values of 1.5 and 1.2 nM, respectively. This compound also potently antagonized the endothelin-1-induced intracellular Ca(2+) increases in both embryonic bovine tracheal (EBTr) cells expressing endothelin ET(A) receptors and human Girardi heart (hGH) cells expressing endothelin ET(B) receptors. In guinea pig isolated tracheas having both endothelin ET(A) and ET(B) receptors, IRL 3630A greatly inhibited endothelin-1-induced contraction (pA(2)=7.1), which was partially or scarcely suppressed by the endothelin ET(A) receptor antagonist cyclo[-(D)-Trp-(D)-Asp-(L)-Pro-(D)-Val-(L)-Leu-] (BQ-123) or the endothelin ET(B) receptor antagonist N-(3, 5-dimethylbenzoyl)-N-methyl-3-(4-phenyl)-(D)-phenylalanyl-(L)-t ryptop han (IRL 2500), respectively. Bolus i.v. injections of IRL 3630A administered into anaesthetized guinea pigs at 10 and 30 microg/kg inhibited endothelin-1 (1.3 microg/kg)-induced changes in respiratory resistance and compliance in a dose dependent manner, whereas both sodium 2-benzo[1, 3]dioxol-5-yl-4-(4-methoxy-phenyl)-4-oxo-3-(3,4, 5-trimethoxy-benzyl)-but-2-enoate (an endothelin ET(A) receptor antagonist: PD 156707) and IRL 2500 at doses of up to 30 microg/kg did not affect endothelin-1-induced changes in respiratory mechanics, reflecting the in vitro results. IRL 3630A is thus an effective bifunctional endothelin receptor antagonist, and will be useful in clarifying the role of endothelin in pulmonary diseases such as bronchial asthma.

Right-sided ulcerative colitis.

This report describes a case of right-sided ulcerative colitis in which multiple shallow ulcers and erosion with symmetric luminal stenosis were distributed segmentally from the ascending colon to the cecum, with a skip lesion composed of superficial erosions in the right half of the transverse colon. Both the rectum and the left colon were spared at the time of onset. Biopsies taken from the lesions showed non-specific inflammation, while those from the rectum and sigmoid colon showed no abnormal findings. A 5-year follow-up study was made based on radiography and endoscopy. Other inflammatory bowel diseases, such as Crohn's disease, tuberculosis, Yersinosis, Behçet's disease, and ischemic colitis were all ruled out, based on the macroscopic and microscopic findings as well as the clinical course. To our knowledge, this is the first report of right-sided ulcerative colitis that has been followed for a long period.

Augmentation of apoptosis in bronchial exuded rat eosinophils by cyclosporin A.

The effect of cyclosporin A on apoptosis in eosinophils was examined to clarify the inhibitory mechanisms of cyclosporin A on allergen-induced eosinophilia in the airway. Eosinophils in bronchoalveolar lavage fluid from sensitized rats after inhaling an allergen were used. More than 50% of the eosinophils that died spontaneously by apoptosis within 24 hour incubation in RPMI 1640 medium contained 10% fetal calf serum. The addition of cyclosporin A or dexamethasone significantly enhanced the eosinophils apoptosis at concentrations of more than 0.1 microM. Apoptosis in eosinophils was considerably suppressed in the presence of culture supernatant of activated splenocytes as a source of various cytokines. Even in the presence of culture supernatant of activated splenocytes, cyclosporin A or dexamethasone facilitated apoptosis in eosinophils. These results suggest that apoptotic death of activated eosinophils is augmented with cyclosporin A and that accelerated apoptosis in eosinophils of the airway may account for the inhibitory effect of cyclosporin A on eosinophilia.

Effect of the eradication of Helicobacter pylori on duodenal ulcer healing and ulcer relapse: randomized controlled study in Japan.

To investigate the effect of the eradication of Helicobacter pylori on the healing and relapse of duodenal ulcers. 50 patients with active duodenal ulcer and H. pylori infection were randomly allocated to two treatment groups. One group (cimetidine group) received cimetidine 400 mg twice daily for 6 weeks and the other group (double-therapy group) received 300 mg amoxicillin granules and 250 mg metronidazole thrice daily for 2 weeks, in addition to the same regimen of cimetidine as the cimetidine group. Forty-two patients completed the study. After confirmation of ulcer scar, all patients were followed up for 6 months while receiving treatment with teprenone, an agent that does not affect acid secretion or the eradication of H. pylori. The healing rates at 6 weeks were 90% in the cimetidine group and 95.5% in the double-therapy group. H. pylori eradication occurred in 0% of the cimetidine group and in 73.7% of the double-therapy group (P = 0.004). The cumulative relapse rates in the two groups at 6 months were 64.3% and 11.1%, respectively (P = 0.0007). In the double-therapy group, the cumulative relapse rate at 6 months in the patients in whom H. pylori persisted was 50% (2/4); the rate was 0% (0/14) in the patients in whom H. pylori had been eradicated (P = 0.005). Histological gastritis significantly improved compared with the baseline in the double-therapy group, but no such improvement was seen in the cimetidine group. White scarring was found in 7.1% of the cimetidine group and in 83.3% of the double-therapy group after 6 months (P < 0.0001). The eradication of H. pylori markedly decreased the relapse rate in duodenal ulcer patients, and it significantly improved both the grade of gastritis and the quality of the ulcer scar.

Preoperative diagnosis of a small, depressed rectal cancer complicating ulcerative colitis. Report of a case.

PURPOSE: This study was designed to clarify the importance of detecting small, depressed colorectal cancer complicating ulcerative colitis. METHODS: A 39-year-old Japanese male, who had an 18-year history of left-sided ulcerative colitis, was admitted to Fukuoka University Hospital for further evaluation of his colitis. Colonoscopy with a dye spraying method clearly revealed a small, depressed lesion and flat plaque-like lesions in the rectum. Biopsies taken from a depressed lesion and plaque-like lesions revealed a signet-ring cell carcinoma and dysplasia, respectively. Total colectomy was performed. RESULTS: Sections from the depressed rectal lesion, measuring 7 x 8 mm in size, revealed a signet-ring cell carcinoma that diffusely invaded the muscularis propria. Lymph node metastasis was evident. Flat plaques and mucosa around the depressed lesion were positive for dysplasia. CONCLUSION: This is the first report of a case of small, depressed rectal cancer complicating ulcerative colitis diagnosed by preoperative colonoscopy. To improve outcome of colonoscopic surveillance in ulcerative colitis, detecting such small, depressed lesions are important, and colonoscopy with a dye spraying method would be useful in detecting them.

[A new index of disease activity in Crohn's disease].

A comprehensive assessment including subjective and objective parameters seems to be essential for evaluation of disease activity in Crohn's disease. The aim of the present study was to develop an activity index (AI) of Crohn's disease, composed of both subjective and objective variables. Data composed of a total 132 clinical examinations in 100 patients with Crohn's disease were used to determine the AI. Three physicians independently evaluated disease severity in each patient. Stepwise multiple regression analysis was carried out with the overall evaluation as a dependent variable, and with 18 parameters as independent variables. Analysis showed that the following seven variables had a significant correlation with physician's overall evaluation: abdominal pain, abdominal tenderness, complications, C-reactive protein (CRP), body temperature, diarrhea, and body mass index (BMI). AI was expressed as follows: AI = 3.5 x abdominal pain + 3 x abdominal tenderness + 3 x complications + CRP + 3 x body temperature + 4 x diarrhea - 0.4 x BMI. AI values below 15 corresponded to inactive disease, values between 15 and 25 to mild disease, values between 25 and 35 to moderate disease, and values above 35 to severe disease. This study suggests that the activity index is useful for evaluation of the effect of medical treatment in Crohn's disease.

Allergic bronchospasm and airway hyperreactivity in the guinea pig.

In passively sensitized guinea pigs, show infusion of an amount of ovalbumin insufficient to evoke airway obstruction induces hyperreactivity of the airways. A wide range of changed responsivity was observed for different test spasmogens, with leukotriene C4 > histamine > prostaglandin F2 alpha > bradykinin > leukotriene E4 > serotonin > acetylcholine. Injection of ovalbumin as a bolus produced pronounced airway obstruction without hyperreactivity. Airway obstruction due to vascular engorgement (dextran infusion) or edema (histamine infusion) did not result in hyperreactivity. Infusion of PAF induced pronounced airway obstruction together with hyperreactivity, but with a rank order of histamine > leukotriene C4 > serotonin > bradykinin > leukotriene E4 > acetylcholine. It can be concluded that allergic airway hyperreactivity in the guinea pig is spasmogen-selective and largely independent of airway obstruction. These observations question the presumption of non-selective hyperreactivity in allergic asthma and cast doubt upon the proposal that airway hyperreactivity is secondary to airway obstruction.

Exacerbation of airway hyperreactivity by (+/-)salbutamol in sensitized guinea pig.

In guinea pigs passively sensitized to ovalbumin, sustained (6 days) subcutaneous infusion of (+/-)salbutamol (1 mg/kg/day) induced significant airway obstruction and heightened responsivity to airway spasmogens. Of these animals, a substantial proportion (78/235) were too responsive to injected spasmogens to permit infusion of ovalbumin or died following infusion of ovalbumin; yet there were few deaths (2/166) amongst the sensitized animals not exposed to (+/-)salbutamol. In comparison to the animals not exposed to (+/-)salbutamol, infusion of ovalbumin led to exaggerated responsivity of the airways to leukotriene C4, leukotriene E4, histamine, serotonin and acetylcholine, but not to prostaglandin F2 alpha or bradykinin. The capacity of sustained exposure to high doses of (+/-)salbutamol to induce airway hyperreactivity to allergic mediators may account for an association between asthma death and regular, excessive use of sympathomimetics.

Secretion and purification of hepatitis C virus NS1 glycoprotein produced by recombinant baculovirus-infected insect cells.

Recombinant baculoviruses that produce a putative non-structural protein 1 (NS1) of hepatitis C virus (HCV), predicted to be the second envelope glycoprotein, were constructed. The recombinant NS1 protein (re-NS1) produced in infected insect cells was localized on the cell surface and was apparently glycosylated, because it was susceptible to treatment with both tunicamycin and N-glycanase. Furthermore, re-NS1 was effectively secreted into the culture supernatant when the putative NS1 signal peptide (SP) was replaced by the SP of rabies virus G protein, and the C-terminal hydrophobic region was eliminated. The secreted re-NS1 was tagged with six His residues at the C terminus and purified simply by native Ni(2+)-nitrilotriacetic acid (Ni(2+)-NTA) affinity column chromatography. An enzyme-linked immunosorbent assay (ELISA) was developed for the serological diagnosis of HC using purified re-NS1. Anti-NS1 antibody (Ab) was detected in 55 of 60 patients (92%) with chronic HC liver diseases. Thus, this ELISA for Ab directed against HCV re-NS1 produced in insect cells is useful for the detection of chronic HC patients.

Ketotifen inhibits exacerbation of allergic airway hyperreactivity by racemic salbutamol in the guinea pig.

In passively sensitized anesthetized guinea pigs, intravenous infusion of low doses of antigen ovalbumin induced a marked increased responsivity of the airways to intravenous injection of leukotriene C4. Sustained infusion of racemic salbutamol intensified responses to leukotriene C4 both before and after infusion of ovalbumin. Hyperreactivity as a result of infusion of ovalbumin was inhibited by intraduodenal injection of either hydrocortisone or ketotifen at doses that did not diminish responses to leukotriene C4 in animals not exposed to antigen. Ketotifen, but not hydrocortisone, inhibited the enhanced hyperreactivity associated with infusion of racemic salbutamol.

A survey of lead poisoning in wild waterfowl in Japan.

Ingested shotgun pellets were found in 13 of 56 hunter-killed wild waterfowl between October and December 1991 from two hunting grounds in Japan. Four of 33 other waterfowl found dead in lightly hunted areas between January 1991 and March 1992 were diagnosed as having lead poisoning. We propose that lead poisoning maybe a threat to waterfowl in Japan.

A comparative study of once-a-day morning and once-a-day bedtime administration of 40 mg famotidine in treating duodenal ulcers.

A randomized controlled study comparing once-a-day morning and once-a-day bedtime administration of 40 mg famotidine in treating duodenal ulcers was carried out in 99 Japanese patients. Endoscopic examinations were performed at the baseline and repeated at 3-week intervals until healing was confirmed. Eighty-two patients fulfilled the evaluation criteria (38 in the morning group and 44 in the bedtime group). In 13 of these patients the antisecretory effects of these regimens were also assessed by 24 h intragastric pH monitoring. The healing rates were 66% after 3 weeks and 95% after 6 weeks in the morning group, and 57% after 3 weeks and 80% after 6 weeks in the bedtime group. The differences were insignificant between the two groups, but there was a higher healing rate tendency after 6 weeks in the morning group (0.05 less than P less than 0.10). Regarding pain subsidence, there were no significant differences between the two groups. Both treatments were significantly superior to the control group in increasing 24 h intragastric pH. The morning regimen was significantly superior to the bedtime regimen in suppression of daytime acidity. On the contrary, the bedtime regimen was significantly superior to the morning regimen in suppression of nocturnal acidity. These findings suggest that suppression of nocturnal acidity is important but not essential to promote duodenal ulcer healing and suppression of daytime acidity is equally important. Thus, once-a-day morning administration of 40 mg famotidine seems to be at least as effective as once-a-day bedtime administration of 40 mg famotidine in treating duodenal ulcers.

Histamine(H1) antagonists and airway hyperreactivity in the guinea-pig.

Ketotifen inhibits development of airway hyperreactivity in guinea-pigs exposed to PAF, (+/-)isoprenaline, immune complexes or endotoxin. Ketotifen is not a competitive histamine(H1) antagonist, so that it cannot be concluded that there is mandatory involvement of histamine in the development or expression of these forms of airway hyperreactivity. This conclusion has been reinforced by determining the efficacy of other histamine(H1) antagonists as inhibitors of PAF-induced airway hyperreactivity. When compounds were administered intravenously, at a dosage (1 mg/kg) which fully abolished responses to intravenous histamine, the observed rank order for inhibition of PAF-induced hyperreactivity was: ketotifen greater than cetirizine greater than acrivastine greater than KB-2413 greater than oxatomide greater than azelastine greater than terfenadine = astemizole = clemastine = mepyramine = loratadine = saline. Terfenadine may lack inhibitory activity because of a capacity to induce airway hyperreactivity in the guinea-pig. It can be concluded that inhibition of the development of airway hyperreactivity is not a characteristic of histamine(H1) antagonists.

Purification and some properties of thiosulphate-cleaving enzyme from Thiobacillus novellus.

A thiosulphate-cleaving enzyme was purified from Thiobacillus novellus and some of its properties studied. The enzyme showed an absorption peak at 279 nm and no peaks between 300 and 650 nm. Its Mr was 38,000. Although the crude enzyme cleaved thiosulphate to form sulphite without addition of cyanide, the purified enzyme required cyanide to cleave thiosulphate. The Km values for thiosulphate and cyanide of the purified enzyme were 1.0 mM and 0.3 mM, respectively. One mol of the enzyme formed 10 mol of thiocyanate per s from thiosulphate and cyanide. The thiosulphate-cleaving activity of the enzyme was strongly inhibited by cysteine, while beta-mercaptoethanol was less inhibitory. The factor which accepted sulphur from thiosulphate in the crude preparation of thiosulphate-cleaving enzyme seemed to be a relatively labile compound with an Mr of 10,000 x 20,000.