Accumulation of plasma cells expressing CXCR3 in the synovial sublining regions of early rheumatoid arthritis in association with production of Mig/CXCL9 by synovial fibroblasts.

Accumulation of plasma cells in the synovium is one of the diagnostic hallmarks in the histopathological manifestations of rheumatoid arthritis (RA). This seems to be prominent even prior to significant B cell infiltration and/or formation of lymphoid follicles in the synovium. To clarify the mechanism of early plasma cell accumulation, we examined in situ expression of chemokines and their receptors using synovial targeting biopsy specimens, which were obtained under arthroscopy from early RA patients. By immunohistochemical staining, plasma cells were found to express a chemokine receptor CXCR3, while synovial fibroblasts in the synovial sublining regions expressed its ligand, Mig/CXCL9. By reverse transcription-polymerase chain reaction (RT-PCR), using targeted lesions of synovial tissues obtained by laser capture microdissection, expression levels of Mig/CXCL9 in the synovial sublining regions were remarkably high and were likely to be associated with interferon (IFN)-gamma expression. Furthermore, cultured synovial fibroblasts were confirmed to produce Mig/CXCL9 upon stimulation with IFN-gamma. Our results indicate that in the early stage of RA, plasma cells expressing CXCR3 may be recruited directly from the circulation into the synovial sublining regions by its ligand, Mig/CXCL9, produced by synovial fibroblasts.

Immunohistochemical localization of senescence marker protein-30 (SMP30) in the submandibular gland and ultrastructural changes of the granular duct cells in SMP30 knockout mice.

Senescence Marker Protein-30 (SMP30) is a calcium-regulating protein that decreases in an androgen-independent manner as aging occurs. An enzyme-labeled antibody technique has demonstrated that SMP30 localized to the ducts (granular, intercalated, and striated ducts) of mouse submandibular glands. Immunoelectronmicroscopy demonstrated that the granular duct cells were strongly positive for SMP30, but that pillar cells in the granular duct were negative for the protein. In SMP30-knockout (KO) mice, the granular ducts were smaller in diameter. Swelling of mitochondria in the granular duct cells was observed; however, this phenomenon was not observed in the pillar cells. After administration of alpha-isoproterenol, a beta-adrenergic stimulant, a large numbers of small secretory granules were present in the granular duct cells and an expansion of the rough endoplasmic reticulum in SMP30-wild type (WT) mice; in contrast, little change was observed in SMP30-KO mice. These results suggest that SMP30 may be closely related to a signal transduction pathway in the granular duct cells of submandibular glands.

Analysis of sputum taken from wheezy and asthmatic infants and children, with special reference to respiratory infections.

Children who are destined to develop asthma are considered to be susceptible to a variety of respiratory pathogens. To elucidate respiratory inflammation among these children, we measured the levels of eosinophil cationic protein (ECP) and tryptase in sputum taken from three different groups of wheezy infants and young children: those with a first wheeze (n = 15); those with recurrent wheeze (n = 27); and those with recurrent wheeze with respiratory distress, namely asthma (n = 56). The numbers of eosinophils or metachromatic cells determined by microscopic analysis of sputum samples were also evaluated in combination with the ECP and tryptase levels. Although neither sputum ECP nor tryptase was a clear discriminative marker that differentiated the three different types of wheezy disease, ECP levels in sputum from the asthma group were significantly higher (2,269.2 +/- 6,216.8 ng/g) than those in the recurrent wheezy group (440.3 +/- 1,199.8 ng/g) or in the first-wheeze group (209.0 +/- 172.9 ng/g). A similar trend was observed with tryptase levels in sputum, but there were no significant differences among the three groups. Sputum taken from asthmatic children showed a marked accumulation of eosinophils. However, an accumulation of eosinophils in sputum (even in the presence of an elevated level of sputum ECP) was not identified in the asthmatic infants < 1 year of age. An accumlation of eosinophils in sputum was not evident until children became > 1 year old and thereafter the eosinophils rapidly increased in number until the children reached 5 years of age. It was noteworthy that sputa positive for pathogenic bacteria, taken from the 1- and 2-year-old asthmatic infants, had a tendency to show high levels of ECP but a reduced number of eosinophils. Along with the wheezy episodes induced by viral infection, primarily and occasionally in combination with secondary bacterial infection, eosinophil activation and infiltration may develop. These predestined immune reactions to various pathogens might be associated with triggering the onset of asthma.

[Effects of institutional therapy and factors which influence the prognosis].

We examined the effect of institutional therapy on asthmatic children and evaluating the factors influencing the prognosis. We analyzed the questionnaires and laboratory findings obtained form the 565 children who had been treated with the therapy in National Higashi-Saitama Hospital from April 1978 to March 1997. The institutional therapy seems to be effective in the following 3 aspects. 1) Pulmonary function test such as FEV1.0, and bronchial hypersensitivity test such as acetylcholine inhalation test were improved. 2) Exercise abilities such as free running were improved. 3) More than 40% of the patients had no asthma attack or only a few mild asthma attacks requiring no medication. When compared the group whose symptoms were not improved with the remission group, who had no asthma attack, significant differences were seen in the following points, 1) The dose and the number of the medicines at the end of the hospitalization, 2) the bronchial hypersensitivity test, 3) the age of onset, 4) the age of the symptoms become perennial and 5) the term from the symptoms become perennial to the hospitalization were significantly different. The therapy principles including the recipes for exercise training were not different between the two groups. It is concluded that we should consider institutional therapy or, a combination of physical training and environmental control as one of the most important therepeutic methods when asthmatic symptoms become perennial in spite of the conventional medication.

An improved synthesis of taurine- and glycine-conjugated bile acids.

A simple and efficient method for the synthesis of taurine- and glycine-conjugated bile acids is described. The condensation reaction was achieved by the simple mixing of unconjugated bile acid (1.0 eq.), taurine (2.0 eq.) (or glycinate ester), diethyl phosphorocyanidate (1.2 eq.) in the presence of triethylamine at room temperature for 30-60 min. Sample clean-up was effected by the use of a prepacked Sep-Pak C18 cartridge for reversed-phase solid extraction or by direct recrystallization, yielding the desired taurine and glycine conjugates in 89-93 and 92-96% isolated yields, respectively.

[Measurement of portal and splenic venous flow volume (PV and SV), congestion index (CI) and SV/PV% in various liver diseases using by Doppler echo-sonography].

We measured the portal and splenic venous flow volume (PV and SV), congestion index (CI) and SV/PV% in various stages of the liver diseases chronic inactive hepatitis (CIH), chronic active hepatitis (CAH), liver cirrhosis without esophageal varices (LCvarix(-)), and liver cirrhosis with esophageal varices (LCvarix(+))¿, and normal volunteers (NC). The results were as follows: PV was 869.4 +/- 184.0 ml/min in NC, 920.4 +/- 242.5ml/min in CIH, 900.0 +/- 216.9ml/min in CAH, 841.8 +/- 253.0 ml/min in LCvarix(-) and 909.7 +/- 430.7ml/min in LCvarix(+). SV was 241.0 +/- 80.0 ml/min in NC, 289.4 +/- 131.6 ml/min CIH, 286.4 +/- 108.8 ml/min CAH, 272.7 +/- 135.7 ml/min in LCvarix(-), 398.0 +/- 280.5 ml/min in LCvarix(+). SV/PV% was 28.1 +/- 10.1 in NC, 31.4 +/- 14.0 CIH, 32.1 +/- 9.6 in CAH, 32.4 +/- 16.0 in LCvarix(-), 43.1 +/- 23.7 in LCvarix(+). CI was 0.06 +/- 0.019 in NC, 0.07 +/- 0.028 in CIH, 0.09 +/- 0.05 in CAH, 0.11 +/- 0.03 in LCvarix(-), 0.145 +/- 0.047 in LCvarix(+). These results suggested that: (1) From the measurement of CI, portal venous pressure is begun to increase at the stage of chronic active hepatitis. (2) Increasing of splenic venous flow volume is begun at the stage of chronic hepatitis and it effects to the portal hypertension of liver cirrhosis. (3) The change of component of intrahepatic portal venous blood flow and decreasing of liver function tests was affected by increasing of splenic venous flow volume. (4) SV/PV% may be useful parameter to evaluate the decreasing of liver function tests and to estimate the complication of esophageal varices at the liver cirrhosis.

Study of liver function in infants with atopic dermatitis using the 13C-methacetin breath test.

Serum glutamic-oxaloacetic transaminase (GOT) levels were determined in 214 infants (133 males and 81 females) with atopic dermatitis during their first visit to the Department of Allergy, National Children's Hospital, Tokyo, Japan. Compared with the normal hospital range, their levels were found to be significantly higher, a tendency which was more conspicuous in lower age groups. We carried out a 13C-methacetin breath test (MBT), administering the stable-isotope-labeled compound to 11 children with higher serum GOT values and 5 within the normal range to investigate hepatic metabolism of methacetin in infants with atopic dermatitis. 13C-methacetin was given orally, and the 13CO2 level in the breath was determined immediately before and after administration, by mass spectrometry. Compared to the normal controls, the atopic infants demonstrated significantly lower 13CO2 peak excretion and delayed peak time. The clearance rate of 13CO2 was also decreased. These results suggest some relationship between atopic dermatitis and liver function in infants.

Pharmacokinetics and pharmacodynamics of the tulobuterol patch, HN-078, in childhood asthma.

BACKGROUND: Recently, a novel percutaneous adhesive tulobuterol preparation, HN-078, has been developed and tests using healthy adult subjects have indicated it to be effective for controlling exacerbations early in the morning if applied at bedtime. In children, percutaneous application is very important to eliminate side effects, including abdominal pain and appetite loss. OBJECTIVE: We report the pharmacokinetics and pharmacodynamics of tulobuterol patch, HN-078, in the treatment of childhood asthma. METHODS: Single applications of HN-078 were applied transdermally in six children with asthma who had been admitted to a hospital. Subjects weighing less than 30 kg received 1 mg of tulobuterol while subjects weighing 30 kg or above received 2 mg on the chest for 24 hours. Serum tulobuterol levels and peak expiratory flow rate were determined before and after each application. RESULTS: Cmax of tulobuterol was determined to be 1.33 +/- 0.21 ng/mL, Tmax was 14.0 +/- 2.0 hours, and AUCO-t was 27.1 +/- 4.2 ng.hr/mL. These pharmacokinetic parameters per body surface area of children were nearly equivalent to those of adults obtained in other studies. Peak expiratory flow rate values obtained after application of HN-078 significantly increased in comparison to those obtained before application. No significant changes were observed in pulse rate or blood pressure, and no side effects were found with regard to the subjective symptoms and skin conditions. CONCLUSIONS: These results suggest that the patch formulation of tulobuterol, HN-078, will be very useful for the treatment of pediatric asthma. It is especially significant that no side effects were observed.

[Investigation into the therapeutic efficacy of continuous isoproterenol inhalation in relation to respiratory infections in asthmatic children].

In our Department, continuous isoproterenol inhalation therapy has been conducted on status asthmatics over the last 10 years. In the present study, we investigated whether there are any differences in the therapeutic efficacy of continuous inhalation of isoproterenol in the presence or absence of respiratory infections which may induce or aggravate asthma attacks. The treatment period was significantly longer in patients with pneumonia. Further prolongation of the treatment period was noted in patients with atelectasis. However, there were no apparent differences in therapeutic efficacy according to age or the severity of attacks. These findings suggest that continuous inhalation of isoproterenol is very effective for status asthmatics if respiratory infections are thoroughly controlled.

[Adrenocortical function in children with near fatal asthma].

The adrenal function of children with near fatal asthma was evaluated by a modified rapid ACTH test. The rapid ACTH tests, which were performed within six months of each life-threatening asthmatic episode, showed extremely low responsiveness. The response in the subjects was significantly lower than that of patients who had received high dose of corticosteroid therapy. It was suggested that the adrenocortical function of children with near fatal asthma had been already suppressed and that adrenal suppression could easily occur in such patients. We advocate the following measures: (1) environmental control, education of patients and their families, physical training accurate medication should be supplied to reduce the use of corticosteroids, (2) sufficient doses of corticosteroids should, however be given to patients with acute exacerbation, (3) not only lung function tests or allergic examinations, but also adrenocortical function tests should be performed on severe asthmatic patients.

[A comparative evaluation of three commercially available peak flowmeters before and after being used 1000 times].

The differences in values, accuracy, reproducibility and interdevice variability of three models of portable peak flowmeters (standard and low range of Assess, mini-Wright, Pulmo-Graph) before use and after they had been used 1000 times. A computerized standard flow generator was used to evaluate. Standard range; The three models showed differences in values before and after 100 uses of less than 3 percent at flow rates below 480 L/min. However, the used flowmeters, especially the Assess, showed higher values compared with the new ones at higher flow ranges. There was no significant difference in accuracy between the new and used mini-Wright and Pulmo-Graph models. The used Assess, howevers, showed higher values at higher ranges, thus demonstrating improved accuracy over the new ones. The reproducibilities of the three models deteriorated at high flow rates. The interdevice variability of the used Pulmo-Graph flowmeter was better than that the new one. No significant changes were observed in the mini-Wright, while the Assess showed deteriorated interdevice variability. Low range; The three models showed no significant differences in values, accuracy, reproducibility or interdevice variability before and after use.

Creatinine-clearance estimates for predicting gentamicin pharmacokinetic values in obese patients.

The Cockcroft-Gault and Salazar-Corcoran equations were compared with respect to prediction of gentamicin pharmacokinetic values in obese and nonobese patients, and the results were used to formulate guidelines for calculating initial gentamicin dosages in obese patients. Creatinine clearance (CLcr) was estimated by applying the Cockcroft-Gault equation using total body weight (TBW), ideal body weight (IBW), and dosage weight (DW) and with Salazar-Corcoran equations using fat-free body mass (FBM) in 100 obese and 100 nonobese patients. Gentamicin pharmacokinetic values (k, CL, and t1/2) were estimated by using CLcr estimated by each method and standardized to a body surface area of 1.73 sq m. Actual pharmacokinetic values were determined by using steady-state gentamicin concentrations and a modified Sawchuk-Zaske equation; these values were compared with the predicted values. In the obese patients, pharmacokinetic values predicted from standardized CLcr by the Cockcroft-Gault equation using estimated DW were not significantly biased, compared with actual values; most predictions produced by the other methods were significantly biased. Predictions produced by the DW method were generally more precise than those resulting from the other methods. In nonobese patients, k values estimated by the Cockcroft-Gault equation using IBW were not significantly biased, while values obtained with all other methods were biased. All methods were biased when predicting CL and t1/2 in nonobese patients. Significant correlations existed between standardized estimates of CLcr (by all methods) and pharmacokinetic values in both groups. Predictions of gentamicin k, CL, and t1/2 were best overall when CLcr was estimated by the Cockcroft-Gault equation using DW, rather than by other methods.(ABSTRACT TRUNCATED AT 250 WORDS)

[Changes in histamine levels in skin chambers. Application for clinical evaluation of atopic dermatitis--report 2].

We measured changes in histamine levels in fluids from children with moderate or severe atopic dermatitis by the skin chamber method and evaluated the correlation with clinical symptoms. Skin chambers were applied to the forearm skin which had been scratched with a needle, and extract of Dermatophagoides farinae (mite antigen) at 50 microns filters as a challenge. We measured the concentrations of histamine 2, 6, 12 and 24 hr after challenge. The skin chamber test was performed before and after beach camp therapy or hospitalization therapy. We treated 26 children with beach camp therapy, and compared the effects with hospitalization in Tokyo (20 children). All the children in both groups showed results of mediator releasability in skin chamber was different in the two groups. The histamine levels were dramatically increased 24 hr after challenge with mite antigen. The levels of histamine in the chambers 24 hr after challenge decreased with treatment along with improvements in skin condition. A marked increase in histamine levels in the skin chamber fluids was observed after 24 hours, whereas no increase was observed when they were challenged with vehicle saline. After 7 days of beach camp therapy, challenge with mite antigen produced only a very small increase in histamine levels even after 24 hours, which was significantly lower than the levels before therapy. In the cases involving hospitalization in Tokyo, histamine was observed to be increased at 24 hr (24.9 +/- 8.0 ng/ml SEM), similarly to the beach campgroup (19.0 +/- 7.3 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluating new and traditional methods for aminoglycoside dosing in patients with various degrees of renal function.

Aminoglycosides are widely used, and clinicians continue to seek newer and better methods for initial dosing of these agents. Recently, three new methods were introduced: Thomson, Reesor Nimmo, and dosing in renopathy by easy-to-use multipliers (DREM). In comparing them with older, traditional dosing methods in patients with various degrees of renal function, the pharmacokinetic variables of gentamicin were determined from steady-state peak (Cmax) and trough (Cmin) serum concentrations using individualized regimens in 88 patients. Dosages were determined in each patient using the method of Hull-Sarubbi, rule of eights, and the three new methods, and the resultant Cmax and Cmin values were calculated from dosages generated by each method. The daily doses and Cmax values derived with the Hull-Sarubbi, Thomson, and Reesor Nimmo methods were not significantly different (p > 0.05). The Hull-Sarubbi was the most precise (root mean squared prediction error 1.3) and least biased (mean prediction error -0.05) of the five methods in predicting target gentamicin serum peak concentrations (Cmax 6.5 mg/L). The Hull-Sarubbi (69%), Thomson (86%), and Reesor Nimmo (70%) methods yielded therapeutic Cmax (5-8 mg/L) in a significantly higher percentage of patients than did the rule of eights (32%) and DREM (35%), (p < 0.05). Therefore, if gentamicin serum concentrations are not available, the first three appear to be reasonable methods for initiating gentamicin dosage regimens, but the last two may not be desirable to use in a clinical setting. These conclusions are based on the assumption that patients are adults with stable renal function and relatively stable clinical conditions.

[An evaluation of three commercially available peak flowmeters].

The accuracy, reproducibility and interdevice variability of three models of portable peak flowmeters (standard and low range) were evaluated by using a computerized standard flow generator. Standard range; The mini-Wright peak flow meter generally overestimated peak flow from 120 L/min to 540 L/min and the Assess peak flowmeter underestimated peak flow at full range. The Pulmo-Graph peak flowmeter showed a similar pattern to the mini-Wright but with more accuracy. The Assess showed the best linearity. The reproducibilities of the three models deteriorated over 20 L/min at high flow rates. The interdevice variability of the mini-Wright was better than those of the Assess or the Pulmo-Graph. Low range: The accuracy and interdevice variability of the Pulmo-Graph were better than those of the Assess or the mini-Wright. The reproducibility of Assess seemed to be inferior to the other's. The measured flow from the low ranges was not the same as that of the standard ranges. The three peak flowmeter models have different spring mechanisms for their readings, and the measured flows by one model should not be compared with other models.

[Does clinical stage have any influence on the results of provocation tests in allergic children? I. The best clinical stage for food provocation tests].

To investigate the best clinical stage for food provocation tests, twenty allergic patients who had early allergic reactions induced by any kind of food were tested. The patients were classified into two groups. Group I; open-food challenge tests were performed on eleven patients within 48 hours of admission. Group II; the tests were performed on nine patients seven or more days after admission. In these food provocation tests, there were nine positive results in Group II (p < 0.05). The percentage of eosinophils in the blood was higher in cases of positive provocation than in cases of negative provocation (p < 0.05). Moreover the percentage of eosinophils decreased after admission day by day in the provocation-negative patients of Group II, but there was no change in the provocation-positive patients. These results suggest that individual sensitivity to food decreased after admission. Therefore, in order to assess the actual sensitivity of allergic children it is necessary that food provocation tests be performed within two days of admission or before admission.

Growth in methylcellulose of human mast cells in hematopoietic colonies stimulated by steel factor, a c-kit ligand.

We examined the effect of Steel factor (SLF) on the development of human mast cells in hematopoietic colonies from cord blood mononuclear cells in methylcellulose culture. When cord blood cells were cultured for 3 weeks, SLF increased the cellular tryptase levels detected in total cultured cells. It also stimulated the formation of small-cell colonies consisting mainly of polymorphonuclear granulocytes and immature blastoid cells in a concentration-dependent manner but not the formation of colonies consisting of large macrophagic cells. A low percentage of tryptase-positive mast-cell-like cells was found in 39 out of 100 granulocyte/blastoid cell colonies. Four of the 100 colonies contained 10-20% tryptase-positive cells, but we failed to observe colonies consisting of > 20% of tryptase-positive cells. These results suggest that the effect of SLF on mast cell growth is brought on by stimulating the growth of primitive hematopoietic progenitors.

[Dermal reactions in children with atopic dermatitis--changes in histamine/tryptase levels in skin chambers. Report 1].

In the present study, we used the skin chamber method to determine histamine and tryptase released by continuous loading of a single antigen on the skin in order to evaluate dermal reactions produced and thereby obtain greater insight into the role of type I allergic reactions in children with atopic dermatitis. The subjects were 46 children, 23 males and 23 females aged 10.6 +/- 4.3 years on the average, who were being treated at the National Children's Hospital for moderate or severer atopic dermatitis. Mite antigen stimulation was carried out by the skin chamber method upon admission and histamine and tryptase levels in collected antigen solution were determined. Histamine levels in the chamber were increased significantly at 6, 12 and 24 hrs after stimulation compared to the control levels (p < 0.01). Tryptase levels were increased 2 hrs after stimulation but decreased with time thereafter. Histamine and tryptase levels were significantly correlated 2 hrs after stimulation, with a correlation coefficient of 0.954 (p < 0.01). No significant correlation was observed 24 hrs after stimulation. These findings indicate that children with severe atopic dermatitis have great skin reactivity and very sensitive to stimulation by antigens. Mast cells and basophils are thought to be involved in immediate and delayed type reactions, respectively.