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63 巻, 6 号
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是木 一也, 藤平 隆司, 白川 文彦, 渡辺 堅一, 江藤 澄哉1987 年 63 巻 6 号 p. 719-726
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーHLA-DR antigens are not expressed on normal circulating T lymphocytes, but recently it has become apparent that HLA-DR antigens are expressed on immunologically activated T lymphocytes, as well as monocytes, macrophages, and B lymphocytes. Namely, the HLA-DR antigens are considered to be one of the activated T cell antigens.
It is apparent from the previous studies that increased numbers of HLA-DR positive T cells frequently appear in the circulation in systemic autoimmune diseases, such as RA and SLE. Furthermore, in such diseases, it is reported that the variations in circulating HLA-DR positive T cells are related to the activity of the diseases.
In this communication, we examined the variations in HLA-DR positive T cells in the peripheral blood of the patients with autoimmune thyroid diseases. HLA-DR positive T cells were detected by cytotoxicity test using anti HLA-DR mouse monoclonal antibody (Leu-HLA-DR antibody) and rabbit complement.
The results indicate that; 1) The percentage of HLA-DR positive T cells were increased in the patients with autoimmune thyroid diseases. 2) The changes of HLA-DR positive T cells accompanied with the stimulation by non-specific mitogens in vitro in autoimmune thyroid diseases did not differ from those in the normal controls. 3) The percentage of HLA-DR positive T cells increased by the stimulation of TSH-receptor and thyroid microsome in Graves' disease, on the other hand, it occurred by the stimulation of thyroglobulin and thyroid microsome in Hashimoto's thyroiditis. 4) The percentage of HLA-DR positive T cells were correlated with TRAb (TSH receptor Ab assayed by Smith's kit) in Graves' disease.
These results suggested that T cells are already activated by the thyroid antigens in vivo in autoimmune thyroid diseases, and that the thyroid-specific antigens which stimulate T cells are different between Graves' disease and Hashimoto's thyroiditis.抄録全体を表示PDF形式でダウンロード (678K) -
I. 副腎髄質オピオイド受容体とオピオイドの結合村瀬 寛, 上久保 啓太, 村山 正憲, 安田 圭吾, 鶴見 介登, 三浦 清1987 年 63 巻 6 号 p. 727-740
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーWe studied the binding of [3H] D-Ala2-D-Leu5-enkephalin ([3H] DADLE) and [3H] diprenorphine to crude plasma membrane fraction obtained from the bovine adrenal medulla (bovine adrenal medullary membranes) in order to characterize adrenal medullary opioid receptors. The [3 H] diprenorphine binding was the highest in crude plasma membrane-mitochondrial fraction among all subcellular fractions studied. The amount of [3 H] diprenorphine bound to bovine adrenal medullary membranes was proportional to the protein concentration. Association kinetics of the [3H] diprenorphine binding to bovine adrenal medullary membranes showed that the maximal binding was achieved following 8 min in-cubation and that the binding conformed the second-order kinetics. [3 H] DADLE and [3H] diprenorphine bound to bovine adrenal medullary membranes with high affinities. The Kd and Bmax for the [3H] DADLE binding were found to be 2.9 nM and 57.5 fmole/ mg protein, respectively, while those for the [3H] diprenorphine binding were 0.31 nM and 250 fmole/mg protein, respectively. Displacement studies showed that the [3H] diprenorphine binding was inhibited dose-dependently by levorphanol, dynorphin (1-13), β-endorphin and DADLE. Levorphanol was at least 1000-fold more potent to inhibit the [3H] diprenorphine binding than dextrorphan, indicating stereospecificity of the [3H] diprenorphine binding.
Na+, Li+ and K+ (100 mM) diminished the [3H] DADLE binding and enhanced [3H] diprenorphine binding. Na++ (100mM) increased the Kd value for the [3H] DADLE binding from 2.9 nM to 14.1 nM. Mn++, Ca++ and Mg++ diminished the [3H] diprenorphine binding. Mn++ (1 mM) increased the Bmax value for the [3H] DADLE binding from 95 fmole/mg protein to 450 fmole/mg protein. These effects of Na+ and Mn++ on the [3 H] diprenorphine binding were found to be dose-dependent. [3 H] Diprenorphine binding to the digitonin-solubilized opioid receptor was also inhibited dose-dependently by Mn++.
These results suggest that bovine adrenal medullary membranes contain high affinity and stereospecific opioid receptors and that the binding of opioids to the bovine adrenal medullary opioid receptors is influenced by cations.
Binding study also revealed the presence of opioid receptors in human malignant pheochromocytoma. The Kd and Bmax of the [3H] diprenorphine binding to crude mem-brane fraction obtained from malignant pheochromocytoma were found to be 0.14 nM and 10.4 fmole/mg protein, respectively.抄録全体を表示PDF形式でダウンロード (1168K) -
II.副腎髄質オピオイド受容体とGTP結合蛋白質の連関村瀬 寛, 上久保 啓太, 村山 正憲, 安田 圭吾, 鶴見 介登, 三浦 清1987 年 63 巻 6 号 p. 741-751
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーWe studied possible coupling of opioid receptors to GTP-binding proteins to clarify the mechanism (s) of opioid action in bovine adrenal medullary membranes. Guanylyl imidodiphosphate (Gpp (NH) p) reduced the binding of [3H] D-Ala2-D-Leu5-enkephalin ([3 H] DADLE) to bovine adrenal medullary membranes dose-dependently, and enhanced the binding of [3H] diprenorphine to them. Gpp (NH) p (0.1 mM) enhanced the Kd value of the [3 H] DADLE binding from 2.9nM to 3.9nM, but did not change its Bmax. Pretreatment of bovine adrenal medullary membranes with pertussis toxin (PT) reduced the [3 H] DADLE binding. The Gpp (NH) p inhibition for [3 H] DADLE binding was diminished by the PT-pretreatment. On the other hand, the [3H] diprenorphine binding to PT-pretreated membranes was higher than that to control membranes. Levorphanol inhibited the adenylate cyclase activity of the rat caudate nucleus crude synaptosomal fraction, but did not change that of bovine adrenal medullary membranes.
These results suggest that opioid receptors in bovine adrenal medullary membranes are coupled to PT-sensitive GTP-binding protein which may not influence on adenylate cyclase.抄録全体を表示PDF形式でダウンロード (1063K) -
野村 隆, 桜田 俊郎, 吉田 克己, 海瀬 和郎, 海瀬 信子, 板垣 洋一, 山本 蒔子, 斉藤 慎太郎, 平山 伊久江, 水柿 道直, ...1987 年 63 巻 6 号 p. 752-772
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーWe studied the role of serum free fatty acid (FFA) in the elevation of serum dializable fraction of T4 (DFT4) and evaluated the serum free T4 (FT4) level in low T3 syndrome.
Serum DFT4 and DFT3 were measured by equilibrium dialysis method with phosphate buffer, and serum FFA concentration was obtained by gas chromatography.
In patients with nonthyroidal systemic illnesses who showed reduced serum total T3 (TT3) and normal total T4 (TT4) (loW T3 group, TT3 48± 14 ng/dl, n = 10), mean (-± SD) DFT4 value (0.032±0.05%) was significantly higher than that of the group of systemic illnesses with normal TT3 and TT4 (normal T3 group, TT3 79±7 ng/dl, n = 10). Mean TT4 value of low T3 group (6.5± 1.0μg/dl) was lower than that of normal T3 group (8.6±2.4μg/dl, p<0.02). There was no difference in mean absolute FT4 (AFT4) value between the two groups (2.07±0.46 vs 2.19±0.53 ng/dl). On the other hand, there was no significant difference in DFT3 value between the groups (0.274±0.043 vs 0.247±0.035%), and mean AFT3 of low T3 group (1.29±0.39 pg/ml) was low than that of normal T3 group (1.92±0.26 pg/ml, p<0.01). In cases of low and normal T3 groups (n = 20), serum thyroxine binding globulin (TBG) concentration had a negative correlation with DFT3 (r =-0.707, p<0.001), but not with DFT4. Although there were no significant differences in serum albumin and TBG concentrations between the two groups, the mean serum total FFA concentration and molar ratio of FFA to albumin in low T3 group (579± 249 μM and 1.31±0.61) were significantly higher than those in normal T3 group (345±170 μM and 0.75±0.32, p<0.05 and <0.025, respectively). All of FFA concentrations in low T3 group, especially oleate, were higher than those in normal T3 group. Moreover, the higher the total FFA concentration was, the greater was the percent fraction of oleate.
DFT4 values were significantly increased by the addition of 1 mM oleate to the sera of low T3 group, and this effect was more marked in the sera of the patients with lower albumin concentration. This effect of oleate on FT4 fraction was also observed by column adsorption chromatography method with Sephadex LH-20. The effect of oleate on DFT4 of subject with TBG deficiency was remarkable, and this was getting less as TBG concen-tration was increased by the addition of the serum containing high TBG to the original serum. In the study of 1-T4 load to the normal serum, this effect of oleate on DFT4 became greater as the amount of 1-T4 loaded was above the maximal binding capacities of TBG and/or thyroxine binding prealbumin (TBPA). And this effect of oleate on DFT4 was not attenuated with barbital buffer which inhibits the binding of T4 to TBPA.
We also studied the ability of oleate in displacing 125I-T4 from HSA or purified TBG (Ka for T4 : 1010M-1). One half maximal B/F were obtained at 3.58 × 10-5M oleate and 1.54 × 10-5M HSA (molar ratio : 2.32) or at 1.96 × 10-5 oleate and 7.7 × 10-9M TBG (molar ratio : 2,545). Although oleate was about 3,000 times less potent than T4 in dis-placing 125I-T4 from TBG,抄録全体を表示PDF形式でダウンロード (1877K) -
是木 一也, 藤平 隆司, 白川 文彦, 渡辺 堅一, 八幡 勝也, 江藤 澄哉, 坂本 久浩, 鈴木 秀郎1987 年 63 巻 6 号 p. 773-781
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーWe investigated the presence of circulating monocyte-specific antibodies (monocyto-toxic activities) by cytotoxicity tests and also the relationships between these monocyto-toxic activities and the clinical data in autoimmune thyroid diseases.
Subjects of this investigation include 16 patients with Graves' disease, 20 patients with Hashimoto's thyroiditis and 10 normal controls. To obtain monocyte-rich population, peripheral blood mononuclear cells of type 0 healthy donor were incubated on culture dishes for 12 hours and dish adherent cells were separated by pipetting. These monocyte-rich population were incubated with sample sera which were previously absorbed by lymphocytes of many different kinds of HLA types to eliminate anti-lymphocyte anti-bodies, and thereafter cytotoxicity tests were performed by adding rabbit complements. The monocytotoxic activities were expressed by %cytotoxicities and the value of %cyto-toxicity over 3.9% (mean + 4SD of %cytotoxicities of normal controls) was considered to be positive in monocytotoxic activity.
The results indicate that; 1) 8 patients (50%) of Graves' disease and 10 patients (50%) of Hashimoto's thyroiditis had positive values of monocytotoxic activity.
2) These positive values of monocytotoxic activity were markedly decreased after absorp-tion of sample sera by monocytes, and these patients who had positive values of monocytotoxic activities to allogenic monocytes also had positive values of monocytotoxic activities to autologous monocytes.
3) Patients who had positive monocytotoxic activities also had high levels of TSH receptor antibody (TRAb) and anti-microsomal antibody, besides, monocytotoxic activity was significantly correlated with levels of TRAb in Graves' diesease.
4) Changes of monocytotoxic activities were correlated with the changes of titers of TRAb, but not with the changes of values of thyroid hormone in the clinical course of a patient with Graves' disease.
These results suggested that monocytotoxic activities increased in some patients with autoimmune thyroid diseases and that the monocyte-specific antibody (monocytotoxic activity) was significantly related to the immunological activity of autoimmune thyroid diseases.抄録全体を表示PDF形式でダウンロード (699K) -
志伊 光瑞1987 年 63 巻 6 号 p. 782-797
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーMonoclonal antibodies to a cytosolic insulin-degrading enzyme (IDE) were produced by fusing spleen cells from mouse immunized highly purified human erythrocyte IDE with mouse myeloma cells.
Four monoclonal antibodies were identified by their ability to bind to 125 I-insulin covalently linked to a cytosolic IDE from human erythrocytes. All four antibodies were found to remove more than 90% of the insulin-degrading activity from erythrocytes extracts, demonstrating that these antibodies were directed against an enzyme which accounts for most of this activity. By immunoprecipitation from metabolically labelled cells and immunoblot procedure, the enzyme from a variety of tissue was shown to be composed of a single polypeptide chain of apparent Mr = 110 kDa.
One of these antibodies; 31H7 was coupled to Affi-Gel 10 and used for the purification of this enzyme. Immobilized antigen was eluted at more than 85% efficiency with buffers consisting of either pH2.3, 2.5M MgCl2 or with 6M urea. However, the antigen eluted under 6M urea retained the highest antigenecity (44%) and biological activity (8%) and the yield of the enzyme obtained from this procedure increased up to 17 fold as com-pared with the conventional method. NaDodSO4/polyacrylamide gel electrophoresis showed a single band of this protein with apparent Mr 110kDa.
These monoclonal antibodies and the purified enzyme will be useful tools for a better understanding of this enzyme, so may lead to the design of specific inhibitors of this enzyme that may be used to treat patients with excessive insulin degradation.抄録全体を表示PDF形式でダウンロード (2003K) -
Euglycemic Glucose Clamp法を用いて李 起東 (哲児)1987 年 63 巻 6 号 p. 798-808
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーTo clarify the mechanism of insulin resistance in pregnancy, we have used the eu-glycemic glucose clamp technique in estradiol (E) treatment (n =6), progesterone (P) treatment (n=28), and Control (n =29) female rats.
E (10μg/day) and P (10mg/day) were injected subcutaneously into female rats for 14 days, to increase E and P concentrations to pregnant levels.
Glucose production and glucose utilization were measured by using [3-3F] -glucose. The results were as follows, 1) Glucose production was almost suppressed at hyperinsu-linemia (11,000μU/ml) both Control and P treatment rats. Then at hyperinsulinemia, glucose utilization rate was almost equal to glucose infusion rate. 2) In P treatment rats glucose utilization was significantly lower (p<0.05) than in Control rats at hyperinsulinemia (11,000 μU/ml). 3) In P treatment rats glucose infusion rate was significantly lower than in Control rats at plasma insulin concentrations of 1,000μU/ml (p<0.02), and 11,000μU/ml (p<0.01), and lower than in E treatment rats at plasma insulin concentrations of 11,000μU/ml (p< 0.05). 4) In a dose-response curve for the effects of four different concentrations of insulin on glucose infusion rate, the insulin resistance induced by progesterone is characterized by a decreased responsiveness to insulin.
The results suggest that progesterone may play an important role in inducing insulin resistance in pregnancy.抄録全体を表示PDF形式でダウンロード (1045K) -
上芝 元1987 年 63 巻 6 号 p. 809-822
発行日: 1987/06/20
公開日: 2012/09/24
ジャーナル フリーTo solve the problem of cross-reaction in immunoassay and determine various steroid hormones simultaneously in a small amount of sample, a method for the quantitative analysis of steroid hormones was developed. This method is a combination of high-performance liquid chromatography (HPLC) and radioimmunoassay (RIA). The purpose of the study is the comprehensive analysis of steroid hormones profiles in normal subjects and adrenocortical diseases.
One hundred μl of plasma was extracted by ether and the ether layer was evaporated. The residue was redissolved and separated by HPLC. Then fractions of steroid hormones were taken and determined by RIA. In this study, cortisol (F), androstenedione (A), 17α-hydroxyprogesterone (17-OHP), testosterone (T), progesterone (P), estrone (E1) and estradiol (E2) were analyzed in normal adults, congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency and Cushing's syndrome. Results in normal adults were similar with those had been previously reported. In CAH, F was remarkably low and 17-OHP, A, T and P were remarkably high before treatment. During treatment some cases showed that 17-OHP, A, T and P were high, and 17-OHP and P tended to be within normal range, if F had been kept higher than about 20μg/dl. In the analysis of Cushing's syndrome before treatment, there were definite differences between adenoma and hyperplasia. A, 17-OHP, T, E1 and E2 were higher in hyperplasia than those in adenoma. It is suggested that it is possible to diagnose the type of Cushing's syndrome with a small amount of plasma using this method.
As a mass screening method of CAH at present, 17-OHP in dried blood on filter paper is determined, therefore the quantitative analysis of 17-OHP in dried blood on filter paper (9 mm disc) was attempted. The quantitative analysis proved to be possible, and it was considered to be applied as the secondary screening method of CAH by the use of dried blood on filter paper.抄録全体を表示PDF形式でダウンロード (1289K)
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