A leucocyte migration inhibition test (LMIT) was carried out in 28 healthy controls and 39 diabetics with special reference to age at onset, insulin-dependency, family history and retinopathy. The method employed was the agarose plate method (Clausen). The antigens used were rat liver mitochondria prepared according to Zamecnik, bovine pancreas extract prepared according to Nerup et al., and rat islet mitochondria prepared according to Howell.
The following results were obtained :
1) There was no significant difference between controls and diabetics as a whole, when the tests were performed with rat liver mitochondria as an antigen. There was also no difference between juvenile onset diabetics and maturity onset diabetics, insulin-dependent diabetics and insulin-nondependent ones, patients with a family history and others without a family history, and the patients with retinopathy and those without retinopathy as far as this antigen was used.
2) There was a tendency toward a higher incidence of a positive LMIT in juvenile onset diabetics and patients with retinopathy as compared with maturity onset diabetics and those without retinopathy when the tests were carried out with bovine pancreas extract as an antigen. But none of them reached statistically significant levels.
3) Statistically higher (p<0.05) incidences of a positive reaction in LMIT were observed in juvenile onset diabetics and insulin-dependent patients as compared with maturity onset diabetics and insulin-nondependent patients when the patients were tested with rat islet mitochondria as an antigen.
Based on the above-mentioned results, the significance of autoimmune mechanism as a possible factor in the pathogenesis of juvenile and insulin-dependent diabetes mellitus was briefly discussed.

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To study the relationship between maternal and fetal pituitary-adrenal axis, plasma concentrations of ACTH, cortisol, progesterone, and unconjugated estradiol and estriol were determined during late pregnancy and labor in women with normal and abnormal pregnancies. Plasma ACTH, progesterone, and unconjugated estradiol and estriol were determined by a radioimmunoassay technique. Unconjugated plasma cortisol was measured by a competitive protein binding method.
Results obtained were as follows :
(1) In 7 healthy women in the 10th month of pregnancy, the concentrations of plasma ACTH just before delivery were statistically higher than those found before the onset of labor. The fetal plasma ACTH concentration was much higher than the maternal level. No ACTH was detected in the cord plasma of two anencephalic infants.
(2) After ACTH infusion into 4 healthy women during labor, the concentrations of ACTH in their plasma increased to about 2.2 to 10.4 times the control values, while the concentrations in the cord plasma of their newborn infants decreased significantly. The difference between the ACTH concentrations in infants delivered from normal women and from women infused with ACTH was significant.
(3) In 6 healthy women, the concentrations of plasma cortisol just before delivery were statistically higher than the levels found before the onset of labor, and the fetal plasma cortisol level was significantly lower than the maternal level. In two women with anencephalic fetuses, the maternal plasma concentrations of cortisol were similar to those in six women with normal fetuses, while the cortisol concentrations in the cord blood were significantly lower than those in normal newborn infants.
(4) After ACTH infusion into 4 healthy women during labor, the concentrations of cortisol in their plasma increased slightly, but not significantly.
(5) In 7 healthy women, the concentrations of plasma estradiol and estriol during labor did not change, and the fetal plasma estradiol and estriol concentrations were significantly higher than the maternal levels. In three women with anencephalic fetuses, the maternal plasma concentrations of estradiol and estriol during labor were very low. It is interesting that estradiol concentrations in the cord blood of two of these fetuses were similar to those of their mothers, while estriol concentrations were higher than those of the mothers.
(6) After ACTH infusion into 4 healthy women during labor, the concentrations of estradiol and estriol in their plasma did not differ from the control values, while the levels of estradiol and estriol in the cord plasma of the newborn infants were significantly lower than those normally found in newborn infants.
(7) There were no differences among the plasma progesterone concentrations during labor in normal pregnant women, in normal pregnant women with ACTH infusion and in pregnant women with anencephalic fetuses or between those in normal newborn infants and those in anencephalic infants.
(8) ACTH was infused for 3 hr into 10 pregnant women beyond term by ten days or more. The concentrations of plasma estradiol and estriol in all the patients did not change for 2 hr following ACTH infusion, but definitely decreased 3 hr after the cessation of ACTH infusion, following a rise of the plasma cortisol levels.
These results obtained suggest that ACTH does not cross the placental barrier, that during late pregnancy and labor the maternal adrenal cortex responds rapidly to infused ACTH by increasing corticosteroids synthesis, and the corticosteroids cross the placental barrier from the maternal to the fetal side and suppress fetal adrenal secretion, and so decrease production of estrogen precursor in the fetal adrenal, resulting in lowered estrogen levels of fetal plasma, and that as far as the progesterone is concerned, the human placenta is a unique site of synthesis and this synthesis is under self-control.

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The mechanism of the enhanced anti-inflammatory activity of synthetic glucocorticoids has not been established. The most potent and useful glucocorticoids, dexamethasone (16α-methyl-9α-fluoro-prednisolone) and betamethasone (16β-methyl-9α-fluoro-prednisolone), have the same anti-inflammatory potency in humans. But in the rat they have a different anti-inflammatory potency. That is, in the rat dexamethasone is considered to have a potency 160 to 190 times that of cortisol, whereas betamethasone is considered to have a potency 10 to 70 times that of cortisol. This difference is thought to depend upon the fact that in the dexamethasone molecule, the 16-methyl radicle is in the ot-position, whereas in the betamethasone molecule it is in the β-position.
This paper describes the plasma concentration, the tissue distribution and the excretion of dexamethasone and betamethasone following intramuscular administration in Sprague-Daulay male rats. In addition, the urinary metabolites of these glucocorticoids in the free fraction are analysed. These experimental data demonstrate the relationship between the structure and the metabolic fate of these two compounds and clarify the mechanism of enhancement in the anti-inflammatory activity of glucocorticoids by structural changes. The radioisotope-labelled glucocorticoids used were cortisol-4-¹⁴C, dexamethasone-1, (2) -³H, and betamethasone-1, 2, 4-³H. Each compound was administered to the rats intramuscularly, and the radioactivities recovered by 75% methanol from plasma, various tissues and feces were measured using a Packard Tricarb Liquid Scintillation Spectrometer. Urinary metabolites were extracted with ethyl acetate, chromatographed in a Frantz Y and Bush B5 system and repurified. The analysis of urinary metabolites was performed using the Porter-Silber reaction, UV absorption, 17-ketonization and the sulphuric acid chromogen spectrum.
The results were as follows :
1) Dexamethasone exhibited longer plasma half-life than betamethasone. The maximum plasma concentration of these two compounds was almost equal.
2) The more potent glucocorticoids, such as dexamethasone and betamethasone, were excreted more slowly in the urine and feces than cortisol.
3) The radioactivities of these two labelled glucocorticoids were recovered more in the liver, kidney and adrenal gland than in the muscle, subcutaneous tissue and brain. And the radioactivities of the more potent dexamethasone were greater in all organs tested than those of the less potent betamethasone.
4) Dexamethasone and betamethasone were less metabolized than cortisol. And the major compounds of their extracts were unchanged materials. But among these two fluorocorticoids, dexamethasone was less metabolized than betamethasone. The main metabolic pathway of the two compounds was as follows : dexamethasone-hydroxylation at C-6, betamethasone-hydroxylation at C-6 and reduction at C-20.
5) In the rat, the more potent dexamethasone exhibits a longer plasma half-life, is excreted more slowly and is less metabolized than the less potent betamethasone.
From the above results it seems logical to assume that the higher concentration of glucocorticoids at the target organs, the greater resistance toward inactivation by the liver, and the slower excretion contribute to the enhancement of the anti-inflammatory activity of synthetic glucocorticoids.

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The effects of acetazolamide which, like the parathyroid hormone, can produce not only bicarbonaturia but also phosphaturia were studied in 4 patients with idiopathic hypoparathyroidism (Id.H.) as well as in a patient with pseudohypoparathyroidism (Ps.H.). All patients were given 250 mg of acetazolamide perorally 3 times a day over a period of 10 days.
Acetazolamide increased serum total calcium concentration (corrected for serum protein), (P<0.01), and decreased serum phosphate concentration (P<0.05) significantly, causing chronic metabolic acidosis in all patients. In addition, Chvostek's and Trousseau's signs became clearly negative.
In all patients, acetazolamide elicited as much excretion of urinary phosphate as that of sodium and potassium but had no significant effect on the urinary excretion of cyclic AMP from the first day of its administration.
In Id.H., in comparison with Ps.H., a slight but significant increase in the urinary excretion of cyclic AMP and calcium began to manifest itself at later periods during the administration of acetazolamide and tended to continue even after it was withdrawn.
In Ps.H., both the increase in serum calcium and the decrease in serum phosphate were greater than in Id.H., in addition to the greater tendency to develop metabolic acidosis. However, little or no increase was observed in the excretion of urinary calcium despite severe chronic metabolic acidosis and high serum calcium levels in Ps.H.
The results suggest that in contrast to the parathyroid hormone, the phosphaturic effect of acetazolamide is not mediated through the renal adenylate cyclase-cyclic AMP system. In Ps.H., acetazolamide could produce an abnormally potent inhibition on the process of bicarbonate reabsorption in proximal tubules, and the inhibition of calcium reabsorption seemed to be hindered by the presence of chronic metabolic acidosis.

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To clarify the function of the renin-angiotensin system in diabetes mellitus with or without complications, the responsiveness of plasma renin activity (PRA) to the administration of furosemide was investigated in 32 patients with diabetes mellitus and in 35 age-matched normal subjects. Ten of these diabetic patients had no detectable complications, and the remaining 22 had one or more of the complications of hypertension, nephropathy and neuropathy.
The complicated diabetic patients had a longer history of diabetes mellitus and a greater increase in fasting blood sugar, as compared with the uncomplicated diabetic patients. Eighty mg of furosemide was orally administrated in the early morning.
Blood for the PRA assay was drawn before and 4 hours after the administration of furosemide, and urine was collected for 4 hours after the administration. PRA was measured by the Skinner method.
The mean baseline PRA in uncomplicated diabetic patients was 1.3±0.3 ng/ml/h, not significantly different from that (1.2±0.1) in normal subjects. On the other hand, the mean baseline PRA in complicated diabetic patients was 0.7±0.1 ng/ml/h, significantly lower than in the normal subjects and the uncomplicated diabetic patients. The means of urine volume and sodium excretion after the furosemide administration were unchanged in uncomplicated diabetic patients and significantly lower in complicated diabetic patients, as compared with the normal subjects. The mean PRA after the furosemide administration in uncomplicated diabetic patients was 3.5±0.7 ng/ml/h, not significantly different from that (3.8±0.5) in normal subjects, while the value in complicated diabetic patients was 1.5±0.2 ng/ml/h, significantly lower than in uncomplicated diabetic patients. When the responses of PRA, urine volume and sodium excretion to the furosemide administration were studied in relation to each complication, the hypertensive group had low levels of baseline PRA, decreased responses of PRA, water diuresis and natriuresis to furosemide administration as compared with the nonhypertensive group. The group with neuropathy also had similar findings, as compared with the group without neuropathy. The group with nephropathy had insignificant decreases in the furosemide-induced diuresis and natriuresis, as compared with the group without nephropathy.
These results suggest that the renin-angiotensin system may function normally in uncomplicated diabetes mellitus but not in complicated diabetes mellitus, and that the blunted responsiveness of the renin-angiotensin system to the furosemide administration in complicated diabetes mellitus may be related to an altered renin release in the juxtaglomerular apparatus and the impairment of diuretic and natriuretic functions in the renal tubules secondary to chronic diabetes mellitus.

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The binding of ³H-corticosterone or ³H-dexamethasone to macromolecules in cytosols of the brain has been studied in vitro in adrenalectomized male rats.
The cytosol fraction was incubated with ³H-corticosterone at 2-4°C for 4 h. Steroid bound to cytosol was separated from the unbound fraction by three methods : gel chromatography of Sephadex G-25, dextran coated charcoal absorption and the protamine precipitation procedure.
The binding properties were analyzed by Scatchard plots. The dissociation constant (K) was. found to be 5.4×10^-9M, and the number of binding sites/mg cytosol protein was 0.49 p moles, indicating a binding with high affinity and limited capacity for ³H-corticosterone.
The ontogeny of the cytosol glucocorticoid binding micromolecules was investigated in the various brain regions of 20, 30 and 65 day-old male. The concentration of corticosterone binding sites was higher at 30 days than at other days. The ³H-corticosterone binding capacity was highest in the hippocampus and septum. Chromatography of the cytosol proteins on Sephadex G-200 showed the presence of two stable corticosterone binding proteins. One (A) was eluted at the void volume fraction, suggesting that its molecular weight is greater than 200,000, and another (B) appeared in the portion corresponding to the elution position of transcortin in serum.
When the same procedure was applied to the study on ³H-dexamethasone binding properties, only one peak (C) was eluted at the void volume fraction. The competitive binding studies of each peak indicated that A and C had a high degree of specificity for corticosterone, cortisol and dexamethasone. After absorption by anti-rat serum using the affinity chromatography, peak B was found to be retained on the column, since it dis appeared when the eluate was further applied on a Sephadex G-200 column. The B binder seems, therefore, to be antigenically similar to transcortin in serum.

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We conducted a medical survey on subjects living in the mountain districts, located at about 400 to 500 m above sea level, near Malang City in the eastern part of Java, the Republic of Indonesia. Serum levels of thyrotropin (TSH) and thyroid hormones were measured in 163 subjects from 9 to 70 years of age, of which 45 were male and 118 were female. A synthesized thyrotropin releasing hormone (TRH) 500pg infusion test was performed on 41 of the subjects from 14 to 70 years of age (7 males, 34 females) and on 15 normal Japanese subjects from 19 to 35 years of age (7 males, 8 females) as controls.
Serum TSH and triiodothyronine (T₃) were measured by means of radioimmunoassay, serum thyroxine (T₄) by competitive protein binding analysis, and serum PBI by the auto-analyser method. The incidence of antithyroglobulin antibody (thyroid test) and antimicrosomal antibody (microsome test) was also studied to assess the role of auto-immunity in goitrogenesis. Goiter size was classified into five grades : from Grade 0 (without goiter) to Grade IV (with huge goiter) by our criteria.
Mean serum PBI values were low in all groups with goiters, especially in subjects with huge goiters, but there was no significant difference between non-goitrous and goitrous groups. In one of the 28 non-goitrous subjects, the serum T₄ level was low while that of the remaining 27 was within the normal range. Serum T₄ values of subjects with big and huge goiters were significantly lower than those of the non-goitrous group. There was no significant difference between the control and non-goitrous groups.
On the other hand, serum T₃ levels were normal in only two of the 28 non-goitrous subjects and were low in the remaining 26. In the group with huge goiters, serum T₃ levels were low in 9, normal in 20, and elevated in only one. T₃ values in the groups with big and huge goiters were significantly higher than those of the non-goitrous group. As mentioned above, there was a negative correlation between serum T₄ and T₃ in subjects with goiters, and relative hypersecretion of T₃ occurred in the goitrous groups in contrast to the non-goitrous group. The result of our study confirms that preferential secretion of T₃ by the thyroid occurs in goitrous subjects and may represent an important homeostatic mechanism to combat against iodine deficiency.
Serum TSH levels in 28 non-goitrous subjects were all within the normal range, and there were elevated serum TSH levels in only 19 out of the 135 goitrous subjects. Although serum log TSH levels do not significantly correlate with the degree of goiter, the mean serum log TSH of the goitrous groups was higher than that of the non-goitrous group, the mean of which was higher than that of the control subjects.
Following TRH infusion, 7 non-goitrous subjects showed normal serum TSH response to TRH. In 34 goitrous subjects, there were various serum TSH responses, which included a normal response in 17, delayed in 6, hypo in 4, non in 5 and hyper in 2. According to our results of the TRH test, the secretion of TSH by the pituitary in goitrous subjects following TRH infusion may be different from that in a normal control. There were two inverse relations between basal TSH and T₄ (r=-0.43, P<0.01) and peak TSH and T₄ (r=-0.44, P<0.01) following TRH infusion. There was, however, no significant relation between serum TSH and T₃. Since serum TSH correlated with serum T₄ in the TRH test, the TSH secretion of the pituitary may be partially controlled by the serum T₄ level. There was no positive case in the thyroid test and only one positive case in the microsome test.
Many of the subjects with goiter have hypothyroidism, if judged by serum T₄, T₃ and PBI.

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This investigation was performed to evaluate the effect in man of the administration of the human growth hormone (HGH) on the levels of blood sugar, serum insulin (IRI), C-peptide (CPR), glucagon (IRG) and free fatty acids (FFA) under hyperglycemic conditions induced by an intravenous glucose infusion.
Twelve healthy, non-obese, and non-diabetic male volunteers, aged 20-24 years, were selected for the investigation. The studies were begun at 8 a.m. after an overnight fast with the subjects in a recumbent position after 30 min of rest.
In the first series of experiments, glucose was infused intravenously at a rate of 14 mg/ kg/min for 180 min in all subjects as the control experiments. In the second series of experiments, HGH (by Raben) at a dose of 1 mg was given according to the following two procedures : Group A : HGH was administered to 8 subjects by a single intravenous injection within 40 min 10 seconds after initiation of the glucose infusion. Group B : HGH in 250 ml physiological saline was given to 4 subjects by a constant intravenous infusion for 30 min, starting 40 min after initiation of the glucose infusion. Blood samples were taken from the antecubital vein at the opposite side of the glucose infusion before and every 10 min after initiation of the glucose infusion.
The results were as follows :
1) In the first series of experiments, the concentration of blood sugar gradually decreased about 60 min after initiation of the glucose infusion, but the so-called “secondary rise” of HGH was not observed during the glucose infusion.
2) The insulin release induced by the glucose infusion was inhibited by the administration of HGH, which continued for 110 min in Group A and 40 min in Group B.
3) In Group A, the concentration of CPR increased 90 min after the administration of HGH but did not increase in Group B.
4) The concentrations of IRG and FFA did not change after the administration of HGH under such a hyperglycemic condition.
These results clearly demonstrated that acute intravenous administration of HGH caused a decrease in IRI release in hyperglycemic subjects. This indicates that HGH manifests a diabetogenic activity even under hyperglycemic conditions. Further investigation should be necessary to elucidate the mechanism of action of HGH.

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