佐藤 稲子

Background: Maternal diet and sociodemographic factors influence xanthophyll concentration and composition in human milk. However, the importance of dietary patterns regarding the intake of fruits, vegetables, and xanthophylls remains unclear. Objective: The aim was to determine the composition of xanthophylls in the human milk of Japanese mothers and explore associations of xanthophylls with dietary and sociodemographic factors. Methods: This cross-sectional study was conducted in the early phase of the Japanese Human Milk Study. Xanthophyll content was measured using liquid chromatography at 30-36 d postpartum. Maternal intake of foods, nutrients, and dietary supplements was estimated using a food-frequency questionnaire. Linear regression models were established using xanthophylls, maternal diet, and sociodemographic factors. Results: Xanthophyll concentrations were measured in human milk from 118 mothers. The xanthophyll concentration varied among individuals. The median (IQR) concentrations of lutein, zeaxanthin, and β-cryptoxanthin were 65.6 ng/mL (51.6-103.4 ng/mL), 18.6 ng/mL (12.9-25.8 ng/mL), and 15.6 ng/mL (9.0-26.0 ng/mL), respectively. In multivariate models, the lutein concentration was associated independently with dietary green vegetables, exclusive breastfeeding, and education (r 2 = 0.153 for the model; β ± SE: 0.468 ± 0.198, 25.048 ± 10.222, and 13.460 ± 6.774; standardized β = 0.210, 0.217, and 0.175; P = 0.019, 0.016, and 0.049 for dietary green vegetables, exclusive breastfeeding, and education, respectively). For zeaxanthin, exclusive breastfeeding was the most appropriate predictor (r 2 = 0.085; β ± SE: 7.811 ± 3.300; standardized β = 0.218; P = 0.020). The highest predictive power for human milk β-cryptoxanthin was obtained with dietary β-cryptoxanthin (r 2 = 0.258; β ± SE: 0.089 ± 0.015; standardized β = 0.468; P < 0.001), attributed to maternal citrus intake. Conclusions: β-Cryptoxanthin in human milk was the xanthophyll most influenced by the maternal diet in Japanese women. The β-cryptoxanthin concentration in human milk was reflected by the maternal β-cryptoxanthin intake, mainly attributed to Japanese citrus consumption. This trial was registered in the Japanese Clinical Trials Registry (https://center6.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000017649) as UMIN000015494.

BACKGROUND: Sodium-glucose cotransporter 2 (SGLT2) inhibitors are widely used to reduce hyperglycemia. The present study investigated the effects of a SGLT2 inhibitor, empagliflozin, on hyperglycemia in a novel rat model of non-obesity type 2 diabetes with enlarged kidney (DEK). METHODS: Male DEK rats with non-fasting blood glucose concentrations ≤300 mg/dl and >300 mg/dl were classified as nondiabetic and diabetic, respectively. Groups of nondiabetic (control) and diabetic (DM-cont) rats were fed standard chow for 12 weeks, whereas another group of diabetic (DM-empa) rats was fed standard chow containing empagliflozin (300 mg/kg/day) for 12 weeks. Blood glucose, body weight, glucose tolerance, food and water intake, urinary volume, plasma and urinary biochemical parameters, and bone mineral density were measured, and their kidneys and pancreas histologically analyzed. RESULTS: Treatment with empagliflozin reduced blood glucose concentration and food intake in diabetic rats, but inhibited loss of adeps renis and led to body weight gain. Empagliflozin attenuated polyuria and polydipsia but increased plasma concentrations of total cholesterol, sodium and total protein toward normal level. Empagliflozin also significantly reduced urinary excretion of proteins and electrolytes and restored bone mineral density and plasma concentrations of valine and isoleucine to normal levels. Moreover, dilation of renal tubules and kidney enlargement were not attenuated in the DM-empa group. CONCLUSION: The response of DEK rats to empagliflozin differed from that of other diabetic animal models, suggesting that DEK rats have unique characters for studying and evaluating the multiple biological effects of SGLT2 inhibitors. These findings also indicted that empagliflozin could ameliorate systemic metabolism and improve renal tubule function in diabetic condition.

Information regarding the pharmacokinetics of oral sildenafil in dogs with pulmonary hypertension is limited. In this study, we examined the pharmacokinetics of oral sildenafil in a canine model of chronic embolic pulmonary hypertension (CEPH). The CEPH model was developed by repeatedly injecting microspheres into the pulmonary arteries. The pharmacokinetics of oral sildenafil at 1, 2 and 4 mg/kg was evaluated using four dogs with pulmonary hypertension in the fasted state. The plasma concentrations of sildenafil were determined using high-performance liquid chromatography, and pharmacokinetic parameters were calculated using a noncompartmental analysis. Sildenafil was well tolerated in this study. Proportional increments in the maximum plasma concentration and area under the curve extrapolated to infinity at drug doses of 1, 2 and 4 mg/kg were detected using a power model analysis. No significant differences were observed among the three doses in the time to maximum plasma concentration. The mean residence time and elimination half-life were slightly but significantly higher at a dose of 4 mg/kg than at a dose of 1 mg/kg.

Cancer consists of heterogeneous cells that contain a small population of cells that possess stem cell properties; these cells, referred to as cancer stem cells (CSCs) or tumor-initiating cells, are involved in tumor progression and metastasis. Using a sphere-forming assay, canine mammary CSCs were found to be similar to human breast CSCs. Metabolic reprogramming has been recognized as a hallmark of various cancers. However, the significance of cellular metabolism in CSCs remains unclear. The aim of this study was to define the metabolic characteristics of CSCs derived from canine mammary tumors and gain an understanding of the maintenance of stemness. We identified metabolite profiles of canine mammary adenocarcinoma cell lines using gas chromatography-mass spectrometry. Metabolites were extracted from both adherent and sphere-forming cells derived from three cell lines. Sphere-forming cells were separated from adherent cells using an orthogonal, partial least-squares discriminant analysis. Sphere-forming cells were found to contain high levels of the amino acids alanine, glycine and proline compared with adherent cells. They also had high levels of palmitoleate, palmitate and dihomo-gamma-linolenic acid compared with adherent cells. In a sphere-forming assay, palmitate increased the number of spheres for all cell lines. These results indicate that the sphere-forming cells derived from canine mammary adenocarcinoma cell lines have specific metabolic profiles that may be useful for the development of CSC-specific therapies targeting metabolic pathways and potential stemness biomarkers; these results also clarify the maintenance of stemness in canine mammary CSCs.

Basic information related to the pharmacokinetics of sildenafil in dogs is scarce. This study aimed to describe the pharmacokinetic properties of oral sildenafil and determine the effect of feeding and dose proportionality. The effect of feeding on pharmacokinetics of sildenafil (1 mg/kg) was investigated using a crossover study with six dogs. In addition, the dose proportionality of sildenafil ranging 1–4 mg/kg was evaluated using five dogs in the fasted states. The plasma concentrations of sildenafil were determined using high-performance liquid chromatography, and pharmacokinetic parameters were calculated using a noncompartmental analysis. Sildenafil administrations were well tolerated in all studies. Feeding reduced the area under the curve extrapolated to infinity (AUCinf) and the maximum plasma concentration (Cmax) significantly. The elimination half-life (T1/2) did not differ between the fasted and the fed states. For dose proportionality, nonproportional increases in AUCinf and Cmax at 1–4 mg/kg doses were detected by a power model analysis.

In dogs, hyperadrenocorticism (HAC) is associated with insulin resistance and diabetes does progress with HAC. There are significant differences in the transcriptomic and proteomic patterns of activated T cells, which parallel the findings in muscle tissues. The aim of this study was to assess how glucocorticoids affect intracellular metabolites in canine peripheral blood mononuclear cells (CnPBMCs) using dexamethasone. A total of 96 metabolites were identified by capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS). After incubation with dexamethasone, the metabolites glucose 1-phosphate, glucose 6-phosphate, fructose 6-phosphate, sedoheptulose 7-phosphate and acetyl-CoA were significantly increased. However, ATP, CTP, dATP, pyruvic acid and NADP(+) were significantly decreased. These results show that a glucocorticoid reduces the catabolic reaction of glucose and accordingly decreases the glucose requirements of CnPBMCs. (C) 2015 Elsevier Ltd. All rights reserved.

Background: Obesity and overweight have been frequently observed in dogs and cats in recent years as in humans. The compositions of fatty acids (FAs) in the accumulated lipids in tissues of obese animals may have important roles in the process and mechanisms related to the onset of metabolic disorders. The purpose of this study was to evaluate the effects of a high fat (HF) diet, which contained a higher proportion of saturated FAs, on FA metabolism and distribution in obese cats. Cats (N = 12) were divided into control diet group (crude fat; 16.0 %) (n = 4) or a high fat (HF) diet group (crude fat; 23.9 %) (n = 8). The HF diet contained up to 60 % of calories from fat and was rich in stearic acid. Blood samples were collected at 0, 2, 4 and 6 weeks after the feeding. Adipose and liver tissues were collected at the 6th week after feeding. We performed analysis of histological findings and fatty acid composition in serum and tissues. Results: Body weights of the cats significantly increased in the HF group. The increased activities of hepatic enzymes and the accumulation of lipid droplets were found in hepatocytes in the HF group at the 6th week after feeding. In this study, the stearic acid (C18:0)-rich HF diet contained less oleic acid (C18:1n-9) and more linoleic acid (C18:2n-6) than the control. However, the composition of oleic acid in the liver was higher, and those of stearic acid and linoleic acid were lower in the HF group at the 6th week after feeding. The higher oleic acid: stearic acid ratio suggests an increase in the conversion from saturated FA to mono-unsaturated FAs, which may reflect the hepatic storage of FAs as a relatively harmless form. Conclusion: The stearic acid-rich HF diet increased hepatic lipid accumulation accompanied by the increased of hepatic oleic acid, increased serum oleic acid and activation of hepatic enzymes. These findings could be an important sign of early stages of dyslipidemia and hepatic damage. Also, the higher oleic acid: stearic acid ratio might be related to the increased activity of SCD-1, which suggests that the stearic acid-rich HF diet evoked hepatic lipogenesis in the feline liver.

Simple liquid chromatographymass spectrometry (LC-MS) was applied to non-targeted metabolic analyses to discover new metabolic markers in animal plasma. Principle component analysis (PCA) and partial least squaresdiscriminate analysis (PLS-DA) were used to analyse LC-MS multivariate data. PCA clearly generated two separate clusters for artificially induced diabetic mice and healthy control mice. PLS-DA of time-course changes in plasma metabolites of chicks after feeding generated three clusters (pre- and immediately after feeding, 0.53 h after feeding and 4 h after feeding). Two separate clusters were also generated for plasma metabolites of pregnant Angus heifers with differing live-weight change profiles (gaining or losing). The accompanying PLS-DA loading plot detailed the metabolites that contribute the most to the cluster separation. In each case, the same highly hydrophilic metabolite was strongly correlated to the group separation. The metabolite was identified as betaine by LC-MS/MS. This result indicates that betaine and its metabolic precursor, choline, may be useful biomarkers to evaluate the nutritional and metabolic status of animals.

Hyperlipidemia refers to increase of triglyceride (TG) and/or total cholesterol (T-cho) in blood. Fatty Acids (FAs) have important roles in the lipid metabolism. The aim of this study was to determine the FA composition of plasma lipid fractions in dogs with hyperlipidemia and to evaluate the FA composition as a new diagnostic marker for obesity at early stage. Thirty-nine dogs were classified into healthy or hyperlipidemia based on the criteria to diagnose hyperlipidemia. The blood biochemical values, such as TG, T-cho, glucose, insulin, adiponectin and Non-Esterified Fatty Acid (NEFA) were measured. FA composition profile was performed on GC/MS system. The values of plasma TG, insulin and NEFA of the hyperlipidemia group were significantly higher than that of control group. Hyperlipidemia group tended to show lower concentration of adiponectin. It was found that only the levels of TG and NEFA, but not T-cho increased significantly in early stage of hyperlipidemia. In hyperlipidemia group, percentages of myristic acid (C14:0), parmitoleic acid (C16:1) and oleic acid (C18:1) increased in total FAs. And the percentage of C18:1 increased in NEFA. Indeed, the higher level of insulin and lower adiponectin concentration were seen in hyperlipidemia group. These results suggest that appearance of insulin resistance may be the result of increases of certain FAs in early stage of insulin resistance. © 2013 Academic Journals Inc.

The establishment of a classification system for domestic animals on consumed feed stuff is thought to be important from both a hygiene and market point of view. We collected plasma samples of Romney lambs (Ovis aries) which were fed one of the following: a herb-clover mix (n=10) which included chicory, red clover, white clover and plantain; a plant-grass mix (n=10) which included plantain, ryegrass and white clover; or a grass mix (n=10) which included ryegrass and white clover. A total of 20 elements in plasma samples obtained from the lambs were analyzed using inductively coupled plasma mass spectrometry. The data were then analyzed by principal component analysis. The lambs were divided into three groups on a score plot depending on the different feed conditions. Furthermore, discriminant analyses of the elements were examined, using linear discriminant analysis with forward stepwise regression. This discriminant function correctly classified the samples from each group. The accuracy of classification of each group, as shown by 10-fold cross-validation, proved the effectiveness of the established discriminant function. It is concluded that using linear discriminant analysis might be a useful tool for the validation of elements from plasma in lambs grown in different conditions.

1,5-anhydro-D-glucitol (1,5AG) is a pyranoid polyol compound found in human circulating blood. Myo-inositol (MI) is a stereoisomer of inositol and serves as a precursor of inositol phospholipids. I,5AG and MI are filtered by the glomerulus and almost completely reabsorbed through the renal tubules. However, under hyperglycemic conditions, reabsorption through the renal tubules is competitively inhibited because the structures of I,5AG and MI resemble that of glucose. In this study, we investigated the kinetics of serum and urine 1,5AG and MI levels in healthy dogs. We demonstrated that 1,5AG and MI exist in canine serum and urine by gas chromatography-mass spectrometry. Under continuous hyperglycemic conditions, the serum 1,5AG concentration in healthy dogs decreased while the serum MI concentration remained unchanged. Urinary excretion of 1,5AG and MI increased significantly after blood glucose concentrations reached 200 to 220 mg/d/. A significant negative correlation was observed between serum 1,5AG and glucose concentrations during hyperglycemia. However, no significant correlation was observed between serum MI and glucose concentrations. In this study, we demonstrated that serum and urine 1,5AG and MI levels were changed by blood glucose concentrations. The serum I,5AG concentration was decreased by continuous hyperglycemia. However, the serum MI concentration does not reflect hyperglycemia.

1. The objective of this study was to determine the effects of the gluconeogenesis inhibitor metformin on 21-d old chickens. The following parameters were measured in the liver and kidney: plasma glucose, plasma mannose, enzyme activities and mRNA expression levels of glucose-6-phosphatase (G6Pase), and phosphoenolpyruvate carboxykinase (PEPCK). 2. Chickens were divided into two groups, and received either metformin (300 mg/kg body weight) or water. Plasma glucose and mannose concentrations were analysed by high performance liquid chromatography (HPLC). G6Pase and PEPCK activities were determined by glucose 6-phosphate and malic acid substrate methods, respectively. The expression levels of mRNA were determined by real-time PCR. 3. Plasma glucose and mannose reached their lowest concentrations 1 h after metformin administration. At 0 center dot 5 h-1 h after metformin administration, the enzyme activities and mRNA expression levels of G6Pase and PEPCK reached their lowest point in the kidney and their highest point in the liver. The decrease observed in the kidney may have been associated with reductions in both plasma glucose and mannose concentrations. 4. In conclusion, the effect of metformin on the kidney of chickens is similar to its effect in mammals. In contrast, no suppression of enzyme activity or mRNA expression was observed in chicken liver. Therefore, the mode of action of metformin, via AMPK activation, may be different in the chicken liver.

Measurements of serum fructosamine, glycated hemoglobin, and glycated albumin (GA) complement serum glucose concentration for better management of diabetes mellitus (DM). Especially, the serum fructosamine test has long been used for diagnosing and monitoring the effect of treatment of DM in dogs. However, fructosamine tests are currently not performed in veterinary medicine in Japan. GA and fructoasmine levels have been shown to strongly correlate. However, the clinical implications of using GA remain to be elucidated. Therefore, the purpose of the current study was threefold: 1) Determine whether GA% is altered by acute hyperglycemia in normal dogs, simulating stress induced hyperglycemia; 2) Demonstrate that GA% does not dynamically change with diurnal variation of blood glucose concentration in diabetic dogs; and 3) Investigate whether GA% is capable of providing an index of glycemic control for 1-3 weeks in diabetic dogs as is the case with diabetic human patients. Our study demonstrated that serum GA% remains very stable and unaltered under acute hyperglycemic conditions (intravenous glucose injection) and in spite of diurnal variation of blood glucose concentration. Furthermore, serum GA% can reflect long-term changes (almost 1-3 weeks) in blood glucose concentration and the effect of injected insulin in diabetic dogs.

Circulating levels of monosaccharides can act as a reflection of systemic glucose/ energy metabolism. Characteristic changes observed in these levels can be seen in patients with diabetes and other metabolic disorders. There have been a few reports describing the significance of mannose metabolism as an energy source under physiological and pathological conditions. However, the relationship between circulating levels of mannose and the pathophysiology of diabetes mellitus are unknown in dogs. This study examined circulating levels of mannose between healthy control and diabetic dogs and evaluated the clinical significance of mannose levels in dogs. Diabetic dogs demonstrated a higher circulating level of mannose in comparison to normal healthy control dogs. Plasma mannose was positively correlated with plasma glucose and fructosamine, respectively. Interestingly, plasma mannose levels were affected by plasma insulin levels. In the context of feeding and glucose tolerance tests, plasma mannose levels responded to changes in circulating insulin levels. Circulating plasma mannose levels decreased after feeding in both control and diabetic animals in spite of observed insulin level differences. However, when glucose tolerance tests were given, a positive correlation between mannose levels and insulin levels was observed. Therefore, plasma mannose levels obtained via glucose tolerance testing may be used as a new diagnostic method for evaluating insulin resistance or deficiency in diabetic dogs.

Serum mannose and glucose concentrations in dogs before and after eating a meal were determined simultaneously with a recently established HPLC method combined with a UV and fluorescence detection system of p-aminobenzoic acid ethyl ester (ABEE)-derivatized monosaccharides. In this newly established HPLC method, detection limits were 0.09 mu mol/L for mannose and 0.04 mmol/L for glucose. Linearity of peak areas vs. amounts of mannose and glucose in the range of 0.27-320 mu mol/L and 0.13-64 mmol/L were observed, respectively. The value of the glucose content measured by the HPLC method was in good agreement with that of the commonly used enzymatic method (control). Serum glucose concentrations in dogs 90 min after the meal were almost the same as those before the meal, whereas serum mannose concentrations decreased significantly after the meal. This HPLC method may be useful for determination of monosaccharides in animal blood. (C) 2007 Elsevier Ltd. All rights reserved.

1. The oral administration of glucose or dietary glucose reduces fasting plasma mannose concentrations in mammals. On the other hand, there have been no reports on plasma mannose levels in birds. We have analysed chicken plasma mannose and glucose by an original high-performance liquid chromatography (HPLC) method, together with plasma non-esterified fatty acid (NEFA) concentrations in chickens. 2. Plasma glucose concentrations of chickens did not differ among three different age groups (0, 18 and 150 d). However, the plasma mannose concentrations of chicks at the age of 0 d were higher than those of chickens at the ages of 18 and 150 d. 3. At the age of 18 and 150 d, plasma glucose concentrations were elevated and plasma mannose and NEFA concentrations were decreased after regular feeding, compared to fasting levels.