Keiko Takeoka

Increase in Serum Levels of Autoantibodies after Attack of Seasonal Allergic Rhinitis in Patients with Graves’ Disease

Background: The prevalence of allergic disease is increasing worldwide, but its influence on the clinical course of autoimmune diseases is unknown. Objective: The purpose of this study was to assess the effect of seasonal allergic rhinitis on the clinical course of Graves’ disease, which has been considered a Th2-dominant autoimmune disease. Methods: Ten patients with Graves’ disease, who were considered to be in a state of remission or near remission, were serially examined for 18 months starting from August. Five of them had seasonal allergic rhinitis due to Japanese cedar pollen, and the remaining patients had no such allergic disorders. Peripheral eosinophil counts, serum concentrations of cedar-pollen-specific IgE, anti-TSH-receptor antibody, anti-thyroid-peroxidase antibody and antithyroglobulin antibody were assessed at 2- to 4-month intervals. Serum thyroid hormones and TSH levels were also measured to evaluate disease activity. Results: All patients with pollinosis had attacks of allergic rhinitis caused by cedar pollen in early March. Subsequently, peripheral eosinophil counts, pollen-specific IgE activity and serum levels of anti-thyroid-peroxidase and antithyroglobulin autoantibodies markedly increased. Serum levels of anti-TSH-receptor antibody increased in 3 patients in association with an increase in serum thyroid hormones but were always negative in 2 patients. The control patients without pollinosis showed no consistent change of these parameters. Conclusions: Seasonal allergic rhinitis aggravated the clinical course of Graves’ disease and induced an increase in serum antithyroid autoantibody concentrations as well as an increase in pollen-specific IgE concentration. These data suggest that environmental antigens induce not only local allergic reactions, but also stimulate thyroid immune reactions toward Th2 proliferation, and finally aggravate Th2-dependent autoimmune thyroid disease.

Enzyme-linked immunosorbent assay for anti-tropomyosin antibodies and its clinical application to various heart diseases

Tropomyosin is one of the key proteins for muscle contraction. We developed an enzyme-linked immunosorbent assay for antibodies to porcine muscle tropomyosin and measured serum anti-tropomyosin antibodies in patients with heart diseases and in normal controls. The mean values of absorbance in the ELISA assay of patients with ischemic heart disease (n=36, P<0.001), dilated cardiomyopathy (n=28, P<0.005), valvular heart disease (n=27, P<0.05), and collagen disease (n=38, P<0.05) were significantly higher than those of normal controls (n=53), but the value in patients with hypertrophic cardiomyopathy (n=19) was not significantly different from that of normal controls. When the cut-off value was fixed at the mean+2 SD of absorbance in normal controls, positive reactions were found in 19.4%, 7.1%, 18.5% and 15.8% of patients with ischemic heart disease, dilated cardiomyopathy, valvular heart disease, and collagen disease, respectively. An inhibition study revealed that anti-tropomyosin antibodies were different from anti-myosin antibodies, but there was a partial cross-reactivity between the two. Thus, there was a weak correlation of the titers of the two types of antibody within the group of heart diseases. These data indicate that measurement of anti-tropomyosin antibodies by ELISA is helpful for detecting autoimmune abnormalities in various heart diseases.

The stability of immunological and biological activity of human thyrotropin in buffer: its temperature-dependent dissociation into subunits during freezing.

The stability of thyroid-stimulating hormone (TSH) in buffer under various storage conditions was studied with regard to its immunoreactivity measured by an immunoradiometric assay and its bioactivity by a sensitive FRTL-5 cell bioassay. The immunoreactivity was well retained at 4 degrees C or 24 degrees C throughout the study period of 90 days. At -20 degrees C, however, it decreased proportionately with the storage time. The mean reduction was 42.1% at 90 days compared with that when stored at -80 degrees C. The bioactivity showed a similar course of change with its reduction of 44.3% at -20 degrees C in 90 days. The loss of both activities was attributed to the dissociation of human (h) TSH molecule into its subunits. The concentration of the alpha subunit of hTSH in those samples stored at -20 degrees C gradually increased from the initial undetectable level to that almost equivalent on a molar basis to the loss of immunoreactivity. The enrichment of albumin in the buffer to a level of more than 1.0% was effective in preventing the occurrence of such a phenomenon. These data indicate that hTSH, when frozen at -20 degrees C in buffer is gradually dissociated into its subunits, despite its outstanding stability for 90 days at both 4 degrees C and 24 degrees C. However, no apparent inconsistency between immunological and biological activities, was observed at any temperature between -20 degrees C and 24 degrees C.

Influence of breast-feeding on the production of cytokines

PROBLEM: Recently, we reported increases in the production of interferon‐γ (IFN‐γ), interleukin‐2 (IL‐2), and IL‐4 during the postpartum period. The present study was undertaken to investigate whether these increases might be explained by increased prolactin while breast‐feeding.
 METHOD: Whole blood from 41 women who were breast‐feeding, 13 women not breast‐feeding, and 31 healthy non‐pregnant women was stimulated with phorbol 12‐myristate 13‐acetate and ionomycin, and the levels of cytokines in the supernatant were measured by enzyme‐linked immunosorbent assay. Their serum levels of prolactin were measured by enzyme immunoassay.
 RESULTS: Increases in IFN‐γ, IL‐2, IL‐4, and IL‐10 production were observed in women who were breast‐feeding but not in women who were not breast‐feeding. Serum levels of prolactin correlated with the levels of IFN‐γ in culture supernatant.
 CONCLUSIONS: These results suggest that breast‐feeding induces production of cytokines and that IFN‐γ production is enhanced by physiological concentrations of prolactin.

A solution for distinguishing Le(a−b−) sera in CA19-9 assays using SphereLight 180 and Architect i2000 assays

BACKGROUND Measurement of carbohydrate antigen (CA) 19-9 is not applicable in patients with Lewis (Le) blood type, Le(a-b-). It is important to distinguish cases with Le(a-b-) before CA19-9 measurement. Therefore, we prepared a cut-off solution that gives a clear index to distinguish Le(a-b-) sera. METHOD The frequencies of Le blood types and the distribution of the CA19-9 values in each Le blood type were examined in 188 healthy subjects. The CA19-9 values for all Le(a-b-) sera and for a portion of Le(a-b+) sera exist below the limit of quantitation as measured by the SphereLight 180 kit. The cut-off solution, which gives a clear cut-off index (COI), was prepared to differentiate Le(a-b-) from Le(a-b+), and was evaluated using the SphereLight 180, Architect i2000, UniCel DxI 800, Elecsys 2010, and Lumipuls ƒ kits. RESULTS The COI was calculated as the mean +3 SD of the CA19-9 values of a cut-off solution that is adjusted to the limit of detection. Both the sensitivities and specificities of the COIs were 100% using the SphereLight 180 kit and 100% and 91.7%, respectively, using the Architect i2000 kit, but these values were not satisfactory using the other CA19-9 assay kits. CONCLUSION The COIs, determined by the cut-off solution, correctly identified all Le(a-b-) sera as Le(a-b-) and differentiated Le(a-b-) sera from other types of sera in CA19-9 assays using only the SphereLight 180 and Architect i2000 kits.