Androgen receptor silences thioredoxin-interacting protein and competitively inhibits glucocorticoid receptor-mediated apoptosis in pancreatic ß-Cells.

Androgen receptor (AR) is known to bind to the same cis-element that glucocorticoid receptor (GR) binds to. However, the effects of androgen signaling on glucocorticoid signaling have not yet been elucidated. Here, we investigated the effects of testosterone on dexamethasone (DEX, a synthetic glucocorticoid)-induced apoptosis of pancreatic ß-cells, which might be involved in the pathogenesis of type 2 diabetes mellitus in males. We used INS-1 #6 cells, which were isolated from the INS-1 pancreatic ß-cell line and which express high levels of AR. Testosterone and dihydrotestosterone inhibited apoptosis induced by DEX in INS-1 #6 cells. AR knockdown and the AR antagonist hydroxyflutamide each diminished the anti-apoptotic effects of testosterone. AR was localized in the nucleus of both INS-1 #6 cells and pancreatic ß-cells of male rats. Induction of thioredoxin-interacting protein (TXNIP) is known to cause pro-apoptotic effects in ß-cells. Testosterone suppressed the DEX-induced increase of TXNIP at the transcriptional level. A Chromatin immunoprecipitation assays showed that both AR and GR competitively bound to the TXNIP promoter in ligand-dependent manners. Recombinant DNA-binding domain of AR bound to the same cis-element of the TXNIP promoter that GR binds to. Our results show that AR and GR competitively bind to the same cis-element of TXNIP promoter as a silencer and enhancer, respectively. These results indicate that androgen signaling functionally competes with glucocorticoid signaling in pancreatic ß-cell apoptosis.

Blood pressure reduction by Japanese traditional Miso is associated with increased diuresis and natriuresis through dopamine system in Dahl salt-sensitive rats.

OBJECTIVES: We investigated the antihypertensive mechanism of long-term Miso soup consumption in Dahl salt-sensitive (Dahl S) rats with salt-induced hypertension. MATERIAL AND METHODS: Female Dahl S rats fed a low-salt (0.3% NaCl) diet were divided into three groups: (1) six rats given water, (2) six rats given 0.65% (w/v) saline solution or (3) eight rats given 5% (w/v) Miso soup containing 0.65% (w/v) saline solution. They were followed for 12 weeks. Variables in the plasma or 24-h urine were determined. Systolic blood pressure (SBP) was measured by the tail-cuff method. RESULTS: The SBP increased in an age-dependent manner in Dahl S rats drinking saline solutions. The elevation of SBP was significantly attenuated in Dahl S rats given Miso soup although the ultimate cumulative salt loading was much greater in the Miso group than those given the saline solutions. This SBP reduction in the Miso group was associated with an increase in fractional excretion of Na (FENa) and free water clearance in the kidney. Urinary dopamine excretions were increased in the Miso group compared with that in the saline group. The increase in urinary dopamine excretions was associated with a decrease in brain oxidative stress. Urinary dopamine excretions were an independent predictor of SBP in the Miso group. CONCLUSIONS: Long-term consumption of Miso soup attenuated blood pressure elevation in Dahl salt-sensitive rats with salt-induced hypertension. The blood pressure reduction was due to, at least in part, constituent(s) of the Miso that increase natriuresis and diuresis and enhance dopaminergic nervous activity in the kidney.

Induction of amino acid transporters expression by endurance exercise in rat skeletal muscle.

We here investigated whether an acute bout of endurance exercise would induce the expression of amino acid transporters that regulate leucine transport across plasma and lysosomal membranes in rat skeletal muscle. Rats ran on a motor-driven treadmill at a speed of 28 m/min for 90 min. Immediately after the exercise, we observed that expression of mRNAs encoding L-type amino acid transporter 1 (LAT1) and CD98 was induced in the gastrocnemius, soleus, and extensor digitorum longus (EDL) muscles. Sodium-coupled neutral amino acid transporter 2 (SNAT2) mRNA was also induced by the exercise in those three muscles. Expression of proton-assisted amino acid transporter 1 (PAT1) mRNA was slightly but not significantly induced by a single bout of exercise in soleus and EDL muscles. Exercise-induced mRNA expression of these amino acid transporters appeared to be attenuated by repeated bouts of the exercise. These results suggested that the expression of amino acid transporters for leucine may be induced in response to an increase in the requirement for this amino acid in the cells of working skeletal muscles.

Subcutaneous administration of sodium alginate oligosaccharides prevents salt-induced hypertension in Dahl salt-sensitive rats.

OBJECTIVE: We investigated the mechanism of antihypertensive effects of sodium alginate oligosaccharides, which are enzymatic products of high-molecular-weight natural alginate from seaweeds, in Dahl salt-sensitive (Dahl S) rats. MATERIALS AND METHODS: Dahl S rats fed a high-salt (4% NaCl) diet were subcutaneously administered sodium alginate oligosaccharides (60 mg/day using a continuous osmotic mini-pump) for 14 days. Systolic blood pressure (SBP) was measured using the tail-cuff method, and we determined the influence of the alginate treatment on the metabolism of sodium by measuring sodium excretions in the feces and urine. RESULTS: SBP increased in an age-dependent manner in the untreated Dahl S rats. Sodium alginate oligosaccharide treatment via the subcutaneous route almost completely abolished salt-induced hypertension in Dahl S rats fed a high-salt diet. The level of fecal or urinary sodium excretion did not significantly change during the treatment period with the alginate oligosaccharides. The reduction in SBP rapidly recovered after cessation of the treatment. Moreover, the level of urinary protein excretion was lower in the treated Dahl S rats than in the untreated rats during the experimental period. CONCLUSIONS: Our results suggest that sodium alginate oligosaccharides attenuate salt-induced hypertension in Dahl S rats not through reducing salt absorption, but probably through a direct action on vascular vessels.

Sodium alginate oligosaccharides attenuate hypertension and associated kidney damage in Dahl salt-sensitive rats fed a high-salt diet.

OBJECTIVES: In this article, the antihypertensive effects of sodium alginate oligosaccharides, enzymatic products of high molecular natural alginate from sea weeds, in Dahl salt-sensitive (Dahl S) rats were investigated. MATERIAL AND METHODS: Dahl S rats fed a high-salt (4% NaCl) diet were treated with sodium alginate oligosaccharides (4% or 8% w/w) for 7 weeks. Systolic blood pressure (SBP) was measured by the tail-cuff method, and hypertensive cardiovascular benefits and kidney damage were assessed. Glomerular function and morphological sclerosis were determined. RESULTS: SBP increased in an age-dependent manner in the untreated Dahl S rats. Sodium alginate oligosaccharide treatment attenuated the increase in SBP in a dose-dependent manner. The heart and aortic walls weighed less in the rats treated with sodium alginate oligosaccharides than in the untreated rats. The SBP reduction was associated with a decrease in urinary protein excretion and an increase in the creatinine clearance rate. Sodium alginate oligosaccharides significantly attenuated hypertensive glomerular sclerosis and arterial injury in the kidney. Fractional excretion of sodium (FENa) decreased in low-salt Dahl S rats and increased with a salt challenge. The alginate oligosaccharides decreased FENa in high-salt Dahl S rats. CONCLUSIONS: The results of this study suggest that sodium alginate oligosaccharides attenuate salt-induced hypertension in Dahl S rats. This reduction is associated with decreases in cardiovascular and renal damage.

Sodium alginate oligosaccharides attenuate hypertension in spontaneously hypertensive rats fed a low-salt diet.

We investigated the effects of sodium alginate oligosaccharides (alginate) on the development of spontaneous hypertension in rats. Spontaneous hypertensive rats were treated with alginate for 7 weeks. Systolic blood pressure (SBP) and cardiovascular and kidney damage were assessed. Systolic blood pressure increased in SHRs and this elevation was attenuated with alginate treatment. The heart weight tended to decline. Alginate did not change plasma cholesterol levels or urinary sodium excretions. The slightly higher urinary protein excretion in SHRs was not changed with the treatment; however, morphologic glomerular damage was significantly attenuated. Sodium alginate oligosaccharide attenuates spontaneous hypertension in SHRs, and may help prevent early-stage kidney injury.

Rapid induction of REDD1 expression by endurance exercise in rat skeletal muscle.

An acute bout of exercise induces repression of protein synthesis in skeletal muscle due in part to reduced signaling through the mammalian target of rapamycin complex 1 (mTORC1). Previous studies have shown that upregulated expression of regulated in DNA damage and development (REDD) 1 and 2 is an important mechanism in the regulation of mTORC1 activity in response to a variety of stresses. This study investigated whether induction of REDD1/2 expression occurs in rat skeletal muscle in response to a burst of endurance exercise. In addition, we determined if ingestion of glucose or branched chain amino acids (BCAA) before exercise changes the expression of REDD1/2 in muscle. Rats ran on a motor-driven treadmill at a speed of 28 mmin(-1) for 90 min, and then the gastrocnemius muscle was removed and analyzed for phosphorylation of the eukaryotic initiation factor (eIF) 4E binding protein 1 (4E-BP1) and expression of REDD1/2. Exercise repressed the mTORC1-signaling pathway regardless of the ingestion of nutrients before the exercise, as shown by dephosphorylation of 4E-BP1. In addition, exercise induced the expression of REDD1 mRNA (∼8-fold) and protein (∼3-fold). Exercise-induced expression of REDD1 was not affected by the ingestion of glucose or BCAA. Expression of REDD2 mRNA was not altered by either exercise or nutrients. These findings indicated that enhanced expression of REDD1 may be an important mechanism that could partially explain the downregulation of mTORC1 signaling, and subsequent inhibition of protein synthesis in skeletal muscle during exercise.

Promoting insulin secretion in pancreatic islets by means of bisphenol A and nonylphenol via intracellular estrogen receptors.

In this study, we investigated the effects of endocrine disrupters bisphenol A (BPA) and nonylphenol (NP) on insulin secretion from rat pancreatic islets. Following acute exposure to BPA and NP, neither BPA nor NP (0.1, 1, 10, 100 and 1000 microg/l) affected insulin secretion in concentrations of 16.7 mM glucose. However, insulin secretion following long-term exposure to BPA or NP for 24 h in 16.7 mM glucose was significantly higher than without exposure. To determine whether increased insulin secretion resulting from long-term exposure to BPA and NP is induced via intracellular estrogen receptors, we blocked the cytosolic/nuclear estrogen receptors, using actinomycin-D (Act-D), an inhibitor of RNA synthesis, and ICI 182,780 (ICI), an estrogen receptor inhibitor. Following long-term exposure to BPA (10 microg/l) or NP (10 microg/l), Act-D or ICI treatment eliminated the facilitation of insulin secretion. In conclusion, we have demonstrated for the first time that long-term exposure to endocrine disrupters, such as BPA and NP, promotes in vitro insulin secretion from the pancreatic islets, via cytosolic/nuclear estrogen receptors.

Endurance exercise induces REDD1 expression and transiently decreases mTORC1 signaling in rat skeletal muscle.

Working muscle conserves adenosine triphosphate (ATP) for muscle contraction by attenuating protein synthesis through several different pathways. Regulated in development and DNA damage response 1 (REDD1) is one candidate protein that can itself attenuate muscle protein synthesis during muscle contraction. In this study, we investigated whether endurance exercise induces REDD1 expression in association with decreased mammalian target of rapamycin (mTOR) complex I (mTORC1) signaling and global protein synthesis in rat skeletal muscle. After overnight fasting, rats ran on a treadmill at a speed of 28 m/min for 60 min, and were killed before and immediately, 1, 3, 6, 12, and 24 h after exercise. REDD1 mRNA and corresponding protein levels increased rapidly immediately after exercise, and gradually decreased back to the basal level over a period of 6 h in the gastrocnemius muscle. Phosphorylation of mTOR Ser2448 and S6K1 Thr389 increased with the exercise, but diminished in 1-3 h into the recovery period after cessation of exercise. The rate of protein synthesis, as determined by the surface sensing of translation (SUnSET) method, was not altered by exercise in fasted muscle. These results suggest that REDD1 attenuates exercise-induced mTORC1 signaling. This may be one mechanism responsible for blunting muscle protein synthesis during exercise and in the early postexercise recovery period.

Angiotensinogen gene polymorphisms and food-intake behavior in young, normal female subjects in Japan.

OBJECTIVE: We examined whether angiotensinogen (AGT) gene polymorphisms are associated with food preferences in young, normal female subjects. METHODS: Fifty-two young, normal female subjects (21-22 y old) were recruited. After a 12-h fast, blood samples were obtained to examine the AGT gene polymorphisms (rs699 and rs7079), angiotensin-converting enzyme (ACE) insertion (I)/deletion (D), and adrenergic ß3 receptor (ADRB3) gene polymorphisms (rs4994). A trained dietitian interviewed the participants to determine the portion size and frequency of food eaten for 1 wk by using the established questionnaire FFQg 3.0. RESULTS: The genotypes of the AGT Met235Thr polymorphisms were TT:TC:CC = 2:19:31 (T:C = 0.22:0.78). The genotypes of AGT rs7079 were CC:CA:AA = 26:21:5 (C:A = 0.70:0.30), and those of ACE were DD:DI:II = 5:28:19 (D/I = 0.37:0.63). The genotypes of ADRB3 Trp64Arg were TT:TC:CC = 38:11:3 (T:C = 0.84:0.16). The total caloric intake was greater for those with the MM/MT genotype of AGT Met235Thr than for those with the TT genotype (1993 versus 1698 kcal/d, P < 0.05). The consumption of total lipids, cholesterol, and unsaturated free fatty acids was also higher in those with the MM/MT genotype of AGT Met235Thr than in those with the TT genotype. However, the AGT polymorphism (rs7079) and the ACE I/D were not associated with food preferences. In contrast, the subjects with ADRB3 Trp64 tended to show a high energy intake and preferences for proteins and lipids including fatty acids and cholesterol. They ate more fish and meat. Multiple regression analysis showed that the energy intake in subjects with the MM/MT genotype was independently determined by total lipids (B = 11.7, P < 0.0001) and carbohydrates (B = 4.6, P < 0.0001). CONCLUSIONS: The AGT Met235Thr polymorphism was significantly associated with a higher caloric intake owing to total fat and carbohydrate consumption.

Japanese traditional miso soup attenuates salt-induced hypertension and its organ damage in Dahl salt-sensitive rats.

OBJECTIVE: We investigated the effects of long-term miso soup drinking on salt-induced hypertension in Dahl salt-sensitive (Dahl S) rats. METHODS: Dahl S rats were divided into four groups that consumed 1) water, 2) a 0.9% NaCl solution, 3) a 1.3% sodium NaCl solution, or 4) miso soup containing 1.3% NaCl. They were followed for 8 wk. Systolic blood pressure and hypertensive organ damage were determined. RESULTS: Systolic blood pressure increased in an age- and dose-dependent manner in Dahl S rats drinking salt solutions. The systolic blood pressure increase was significantly less in the Dahl S rats that drank miso soup, although the ultimate cumulative salt loading was greater than that in the Dahl S rats given the 1.3% NaCl solution. This blood pressure decrease was associated with a morphologic attenuation of glomerular sclerosis in the kidney and collagen infiltration in the heart. Urinary protein excretions were less in the miso group than in the rats given the 1.3% NaCl solution. The fractional excretion of sodium was increased and that of potassium was decreased in Dahl S rats given the 1.3% NaCl solution, and these effects were reversed in rats given miso soup toward the values of the control. CONCLUSION: We found that long-term miso soup drinking attenuates the blood pressure increase in salt-induced hypertension with organ damage. This may be caused by a possible retardation of sodium absorption in the gastrointestinal tract or by the direct effects of nutrients in the miso soup from soybeans. The decrease was associated with decreases in cardiovascular and renal damage.

Intestinal expression and metabolic activity of the CYP3A subfamily in female rats.

The intestinal expression of the CYP3A subfamily was investigated in female rats, and the intestinal metabolism of two CYP3A substrates, testosterone and rifabutin, was examined and compared between males and females. CYP3A1/23 and CYP3A2 intestinal expression was barely detected in male and female rats. Although CYP3A9 was predominantly expressed in the female rat liver, its expression in the intestine was not different between the two sexes. The rate of testosterone 6beta-hydroxylation in the female intestine was similar to that for males. Rifabutin was also metabolized at similar rates in both intestines, although the metabolic rate was greater in the female liver. These results indicate that the intestinal drug metabolizing activity of the CYP3A subfamily is similar between males and females, and that CYP3A9 is involved in the intestinal metabolism of CYP3A substrates in both sexes.

Pharmacokinetic characterization of transcellular transport and drug interaction of digoxin in Caco-2 cell monolayers.

To characterize the intestinal absorption of digoxin, its transcellular transport and drug interaction activity was investigated using Caco-2 cell monolayers. We examined digoxin transport in the presence and absence of ouabain to determine whether digoxin binding to Na+,K(+)-ATPase affects its transcellular digoxin transport, and evaluated its influx and efflux clearance by model-dependent pharmacokinetic analysis. Transcellular transport in the basal-to-apical direction was greater than that in the opposite direction. In addition, ouabain decreased the cellular accumulation of digoxin, but it did not alter its transcellular transport profile. The observations for transcellular transport and cellular accumulation in the presence of ouabain were used for the pharmacokinetic analysis, which showed that the efflux clearance of digoxin on the apical side of the monolayer was 15 times greater than that on the basal side. Apical-to-basal transport was increased by carvedilol and pimobendan, and these compounds suppressed the efflux clearance on the apical side and the influx clearance on the basal side. These findings indicate that the intestinal absorption of digoxin is primarily dominated by the efflux process on the luminal side of the intestine, and that carvedilol and pimobendan may vary the rate of intestinal digoxin absorption mainly by inhibiting its exsorptive transport.

T393C polymorphism of GNAS1 associated with the autonomic nervous system in young, healthy Japanese subjects.

1. T393C polymorphism of the gene encoding the Gs-protein alpha-subunit (GNAS1) has been reported recently to be associated with hypertension in which dysfunctions of the autonomic nervous system (ANS) are closely involved. In the present study, the association of this polymorphism with ANS activity was investigated in young, healthy Japanese males. 2. Four hundred and one subjects were genotyped for the T393C polymorphism of GNAS1 by polymerase chain reaction-restriction fragment length polymorphism. Autonomic nervous system activity during supine rest and when standing was assessed in 137 subjects by electrocardiogram R-R interval power spectral analysis. 3. One hundred and fifty-four subjects (38.4%) were homozygous for the T allele (TT), 188 (46.9%) were heterozygous (TC) and 59 (14.7%) were homozygous for the C allele (CC). There were no significant differences as to genotype among the clinical characteristics investigated. In power spectral analysis of heart rate variability, the high-frequency component and parasympathetic nervous system (PNS) index during supine rest were significantly lower in TT and TC carriers than in CC carriers. Furthermore, the increase in heart rate and the responsiveness of sympathetic nervous system index and PNS index to postural change from supine rest to standing were significantly lower in TT and TC carriers than in CC carriers. 4. These observations suggest that the GNAS1 T393C polymorphism is associated with ANS activity in youth, so that it may be useful as a genetic marker for future pathogenesis of hypertension. Follow-up studies are necessary to clarify the prevalence rates of hypertension among 393T allele carriers in the present study.