Keigo Yasuda

Leptin and high glucose stimulate cell proliferation in MCF-7 human breast cancer cells: reciprocal involvement of PKC-α and PPAR expression

Glucose concentration may be an important factor in breast cancer cell proliferation, and the prevalence of breast cancer is high in diabetic patients. Leptin may also be an important factor since plasma levels of leptin correlated with TNM staging for breast cancer patients. The effects of glucose and leptin on breast cancer cell proliferation were evaluated by examining cell doubling time, DNA synthesis, levels of cell cycle related proteins, protein kinase C (PKC) isozyme expression, and peroxisome proliferator-activated receptor (PPAR) subtypes were determined following glucose exposure at normal (5.5 mM) and high (25 mM) concentrations with/without leptin in MCF-7 human breast cancer cells. In MCF-7 cells, leptin and high glucose stimulated cell proliferation as demonstrated by the increases in DNA synthesis and expression of cdk2 and cyclin D1. PKC-alpha, PPARgamma, and PPARalpha protein levels were up-regulated following leptin and high glucose treatment in drug-sensitive MCF-7 cells. However, there was no significant effect of leptin and high glucose on cell proliferation, DNA synthesis, levels of cell cycle proteins, PKC isozymes, or PPAR subtypes in multidrug-resistant human breast cancer NCI/ADR-RES cells. These results suggested that hyperglycemia and hyperleptinemia increase breast cancer cell proliferation through accelerated cell cycle progression with up-regulation of cdk2 and cyclin D1 levels. This suggests the involvement of PKC-alpha, PPARalpha, and PPARgamma.

Responses of plasma adrenocortical steroids to low dose ACTH in normal subjects

OBJECTIVE The standard ACTH test in clinical use employs a pharmacological dose of ACTH which assesses the maximum secretory capacity of the adrenal cortex. We have investigated the responses of plasma adrenocortical steroids including cortisol, aldosterone and dehydroepiandrosterone (DHEA) to physiological doses of ACTH (ACTH 1–24, tetracosactide, Cortrosyn) and determined the minimal dose which induces a response equivalent to that induced by a pharmacological dose of ACTH.

Glucose Tolerance and Insulin Secretion in Patients with Parathyroid Disorders: Effect of Serum Calcium on Insulin Release

To evaluate the role of serum calcium in human insulin secretion, insulin responses after a 100-g oral glucose load were studied in nine patients with primary hyperparathyroidism, five with idiopathic hypoparathyroidism, three with pseudohypoparathyroidism and one with normocalcemic secondary hyperparathyroidism. Glucose tolerance values in these disorders were almost normal. Insulin responses in primary hyperparathyroidism were increased, and those in idiopathic hypoparathyroidism and pseudohypoparathyroidism were reduced significantly as compared to normal subjects. Isulin response in secondary hyperparathyroidism was normal. The calculated insulin area during an oral glucose load was significantly correlated with serum calcium (5.1 to 12.2 mg per deciliter), and a linear relation was obtained (y = 1.59x - 3.3, r = 0.81, p less than 0.001), although a relation with the glucose area was not found. These observations indicate that serum calcium has an important effect on insulin secretion in parathyroid disorders.

Increased oxidized form of human serum albumin in patients with diabetes mellitus

High-performance liquid chromatographic (HPLC) analysis of human serum albumin (HSA) on Asahipak GS-520H columns at neutral pH (6.87) showed a clear resolution of human mercaptalbumin (HMA) and nonmercaptalbumin (HNA), which are reduced and oxidized form of HSA, respectively. We studied the conversion of HMA to HNA (mercapt-nonmercapt conversion) as an index of oxidative change of the tissues and organs in 28 normal subjects and in a total of 47 patients with non-insulin dependent diabetes mellitus (NIDDM). Mean (+/- SD) values of the HMA fraction of HSA, f(HMA), [HMA/(HMA + HNA)], was significantly lower in NIDDM patients than in normal subjects (0.63 +/- 0.067 vs 0.75 +/- 0.028, P < 0.001). It was lower in poorly controlled NIDDM patients (0.63 +/- 0.058, n = 20) than in well controlled NIDDM patients (0.67 +/- 0.032, n = 9) (P < 0.05). Plasma glucose values sampled on occasions including overnight fasting and postprandial ones (r = -0.441, n = 47, P < 0.01), but not plasma glucose values sampled on overnight fasting (r = -0.345, n = 29) or postprandial (r = -0.467, n = 18) conditions and HbA1c (r = -0.211, n = 34), negatively correlated with the f(HMA) values, indicating that mercapt-nonmercapt conversion may not be due to cumulative hyperglycemia over a month, but due to short-term alteration in blood glucose level. The presence or absence of diabetic complications including nephropathy, retinopathy and neuropathy did not affect the f(HMA) values. In conclusion, decreased f(HMA) values in the diabetic patients suggested the presence of a rapidly altered oxidative change of albumin due to hyperglycemia.

Prevalence of thyroid hormone autoantibodies in healthy subjects

OBJECTIVE The prevalence of thyroid hormone autoantibodies in patients with thyroid disorders has been well described, although the results have been variable. However, the prevalence of thyroid hormone autoantibodies in apparently healthy subjects remains unknown and its determination was the main objective of this study. SUBJECTS AND METHODS Sera obtained from 880 healthy subjects (365 men and 515 women) were examined for thyroid hormone autoantibodies by immune precipitation using radiolabelled thyroid hormones or thyroid hormone analogues.

Soy Product Intake and Serum Isoflavonoid and Estradiol Concentrations in Relation to Bone Mineral Density in Postmenopausal Japanese Women

To evaluate soy intake and serum concentrations of estradiol and isoflavonoids and their relationship to bone mineral density (BMD) and serum bone-specific alkaline phosphatase (bone ALP) activity, we conducted a cross-sectional study of 87 postmenopausal Japanese women. Soy product and isoflavone intake from soy products and intake of nutrients were assessed with a semiquantitative food-frequency questionnaire. BMD (mg/cm2) was measured by single-energy X-ray absorptiometry at the site of the calcaneus. Serum estradiol (E2) and the sex hormone-binding globulin (SHBG) were measured by radioimmunoassay. Serum genistein and daidzein concentrations were measured by a high-performance liquid chromatography MS/MS method. A statistically significant correlation was observed between the ratio of E2 to SHBG and BMD (Spearman r = 0.38, p = 0.0003) after controlling for age, body mass index, smoking status, age at menarche, and intake of vegetable fat, vitamin C and salt. Soy product and isoflavone intake and serum isoflavones were not significantly correlated with BMD after controlling for the covariates. Serum ALP was not significantly correlated with soy product and isoflavone intake, the E2/SHBG ratio or serum isoflavones. The present study supports the association of BMD with serum estradiol; however, it does not support the association of BMD with soy or isoflavone intake or serum isoflavone levels.

Hot flushes and other menopausal symptoms in relation to soy product intake in Japanese women.

OBJECTIVE To examine the relationships between dietary intake of soy products and hot flushes and other menopausal symptoms. METHODS Subjects were 284 women aged 40-59 years who attended a health check-up program provided by a general hospital in Gifu, Japan. They completed a health questionnaire including the Kupperman test of menopausal distress. Diet was assessed by a semiquantitative food frequency questionnaire. RESULTS Fermented soy product intake but not total soy product intake was significantly negatively correlated with hot flush severity (r = -0.16, p = 0.01) after controlling for age and menopausal status. Neither total soy product intake nor fermented soy product intake was significantly correlated with menopausal index score. Estimated isoflavone intake from total and fermented soy products was significantly lower by 15% (p = 0.02) and 19% (p = 0.01), respectively, in women with hot flushes, compared to those without hot flushes after controlling for covariates. CONCLUSION The data support a hypothesis that intake of fermented soy products alleviates the severity of hot flushes.

DHEA improves glucose uptake via activations of protein kinase C and phosphatidylinositol 3-kinase

We have examined the effect of adrenal androgen, dehydroepiandrosterone (DHEA), on glucose uptake, phosphatidylinositol (PI) 3-kinase, and protein kinase C (PKC) activity in rat adipocytes. DHEA (1 μM) provoked a twofold increase in 2-[3H]deoxyglucose (DG) uptake for 30 min. Pretreatment with DHEA increased insulin-induced 2-[3H]DG uptake without alterations of insulin specific binding and autophosphorylation of insulin receptor. DHEA also stimulated PI 3-kinase activity. [3H]DHEA bound to purified PKC containing PKC-α, -β, and -γ. DHEA provoked the translocation of PKC-β and -ζ from the cytosol to the membrane in rat adipocytes. These results suggest that DHEA stimulates both PI 3-kinase and PKCs and subsequently stimulates glucose uptake. Moreover, to clarify the in vivo effect of DHEA on Goto-Kakizaki (GK) and Otsuka Long-Evans fatty (OLETF) rats, animal models of non-insulin-dependent diabetes mellitus (NIDDM) were treated with 0.4% DHEA for 2 wk. Insulin- and 12- O-tetradecanoyl phorbol-13-acetate-induced 2-[3H]DG uptakes of adipocytes were significantly increased, but there was no significant increase in the soleus muscles in DHEA-treated GK/Wistar or OLETF/Long-Evans Tokushima (LETO) rats when compared with untreated GK/Wistar or OLETF/LETO rats. These results indicate that in vivo DHEA treatment can result in increased insulin-induced glucose uptake in two different NIDDM rat models.We have examined the effect of adrenal androgen, dehydroepiandrosterone (DHEA), on glucose uptake, phosphatidylinositol (PI) 3-kinase, and protein kinase C (PKC) activity in rat adipocytes. DHEA (1 microM) provoked a twofold increase in 2-[3H]deoxyglucose (DG) uptake for 30 min. Pretreatment with DHEA increased insulin-induced 2-[3H]DG uptake without alterations of insulin specific binding and autophosphorylation of insulin receptor. DHEA also stimulated PI 3-kinase activity. [3H]DHEA bound to purified PKC containing PKC-alpha, -beta, and -gamma. DHEA provoked the translocation of PKC-beta and -zeta from the cytosol to the membrane in rat adipocytes. These results suggest that DHEA stimulates both PI 3-kinase and PKCs and subsequently stimulates glucose uptake. Moreover, to clarify the in vivo effect of DHEA on Goto-Kakizaki (GK) and Otsuka Long-Evans fatty (OLETF) rats, animal models of non-insulin-dependent diabetes mellitus (NIDDM) were treated with 0.4% DHEA for 2 wk. Insulin- and 12-O-tetradecanoyl phorbol-13-acetate-induced 2-[3H]DG uptakes of adipocytes were significantly increased, but there was no significant increase in the soleus muscles in DHEA-treated GK/Wistar or OLETF/Long-Evans Tokushima (LETO) rats when compared with untreated GK/Wistar or OLETF/LETO rats. These results indicate that in vivo DHEA treatment can result in increased insulin-induced glucose uptake in two different NIDDM rat models.

Congenitally defective aldosterone biosynthesis in humans: The involvement of point mutations of the P-450C18 gene (CYP11B2) in CMO II deficient patients

The gene for steroid 18-hydroxylase (P-450C18) has been recently assigned to encode corticosterone methyl oxidases Type I and Type II which were previously postulated to catalyze the final two steps in the biosynthesis of aldosterone in humans. Molecular genetic analysis of the P-450C18 gene is three patients from three different families affected with CMO II deficiency has indicated that a point mutation of CGG----TGG (181Arg----Trp) in exon 3 and one of GTG----GCG (386Val----Ala) in exon 7 occur exclusively in the gene of the patients. Analysis of PCR products by restriction enzymes (HapII and HphI) has indicated that the patients are homozygous and the unaffected parent is heterozygous for both mutations, in accordance with the established concept that CMO II deficiency is inherited in an autosomal recessive manner. These data clearly provide the molecular genetic basis for the characteristic biochemical phenotype of CMO II clinical variants.

Association of intakes of fat, dietary fibre, soya isoflavones and alcohol with uterine fibroids in Japanese women.

Certain dietary components which could affect oestrogen may have implications in the aetiology of uterine fibroids. We previously found that soya intake was inversely associated with a subsequent risk of hysterectomy, suggesting a potentially protective effect of soya against uterine fibroids, the major clinical indication for hysterectomy. We cross-sectionally assessed the associations of intakes of fat, soya foods, dietary fibre and alcohol with uterine fibroids. Study subjects were 285 premenopausal Japanese women participating in a health-check up programme, including gynaecological examinations, provided by a general hospital between October 2003 and March 2006. The presence of fibroids was confirmed by transvaginal sonogram. If women had undergone hysterectomy, self-report of fibroids was accepted. Each subjects usual diet, including alcohol, was determined with the use of a validated FFQ. Fifty-four women were identified as prevalent cases of fibroids or having had hysterectomy due to fibroids. The mean alcohol intake was statistically significantly higher among women with fibroids than among those without fibroids after controlling for known or suspected risk factors. For the highest compared with the lowest tertile of alcohol intake, the OR of uterine fibroids was 2.78 (95% CI 1.25, 6.20). There was no significant association of intake of fats, soya isoflavones or dietary fibre with uterine fibroids. The data suggest that higher alcohol intake is associated with a higher prevalence of uterine fibroids. Further studies on diet, especially phyto-oestrogens, and uterine fibroids are needed given the limited data currently available.