Cell type-dependent modulation of senescence features using Weo electrolyzed water.

Electrolyzed-reduced water has powerful antioxidant properties with constituents that scavenge reactive oxygen species (ROS), which are known to be produced by several intrinsic and extrinsic processes. When there is an imbalance between ROS production and antioxidant defenses, oxidative stress occurs. Persistent oxidative stress leads to cellular senescence, an important hallmark of aging, and is involved in several age-related conditions and illnesses. This study aims to investigate whether Weo electrolyzed water (WEW) could modulate the phenotype of senescent cells. We compared normal human lung fibroblasts (BJ) and breast cancer cells (T47D) treated with hydrogen peroxide (H2O2) to induce senescence. We assessed the molecular impact of WEW on markers of cellular senescence, senescence-associated secretory phenotype (SASP) factors, and stress response genes. Treatment with WEW modulated markers of cellular senescence, such as the senescence-associated ß-galactosidase (SA-ß-gal) activity, EdU incorporation and p21 expression, similarly in both cell types. However, WEW modulated the expression of SASP factors and stress response genes in a cell type-dependent and opposite fashion, significantly decreasing them in BJ cells, while stimulating their expression in T47D cells. Reduction in the expression of SASP factors and stress-related genes in BJ cells suggests that WEW acts as a protective factor, thereby reducing oxidative stress in normal cells, while making cancer cells more sensitive to the effects of cellular stress, thus increasing their elimination and consequently reducing their deleterious effects. These findings suggest that, due to its differential effects as a senomorphic factor, WEW could have a positive impact on longevity and age-related diseases.

Optimisation and Validation of a Nutritional Intervention to Enhance Sleep Quality and Quantity.

BACKGROUND: Disturbed sleep may negatively influence physical health, cognitive performance, metabolism, and general wellbeing. Nutritional interventions represent a potential non-pharmacological means to increase sleep quality and quantity. OBJECTIVE: (1) Identify an optimal suite of nutritional ingredients and (2) validate the effects of this suite utilising polysomnography, and cognitive and balance tests. METHODS: The optimal and least optimal combinations of six ingredients were identified utilising 55 male participants and a Box-Behnken predictive model. To validate the model, 18 healthy, male, normal sleepers underwent three trials in a randomised, counterbalanced design: (1) optimal drink, (2) least optimal drink, or (3) placebo were provided before bed in a double-blinded manner. Polysomnography was utilised to measure sleep architecture. Cognitive performance, postural sway, and subjective sleep quality, were assessed 30 min after waking. RESULTS: The optimal drink resulted in a significantly shorter sleep onset latency (9.9 ± 12.3 min) when compared to both the least optimal drink (26.1 ± 37.4 min) and the placebo drink (19.6 ± 32.0 min). No other measures of sleep, cognitive performance, postural sway, and subjective sleep quality were different between trials. CONCLUSION: A combination of ingredients, optimised to enhance sleep, significantly reduced sleep onset latency. No detrimental effects on sleep architecture, subjective sleep quality or next day performance were observed.

Dietary nitrate supplementation improves sprint and high-intensity intermittent running performance.

The influence of dietary nitrate (NO3-) supplementation on indices of maximal sprint and intermittent exercise performance is unclear. PURPOSE: To investigate the effects of NO3- supplementation on sprint running performance, and cognitive function and exercise performance during the sport-specific Yo-Yo Intermittent Recovery level 1 test (IR1). METHODS: In a double-blind, randomized, crossover study, 36 male team-sport players received NO3--rich (BR; 70 mL·day-1; 6.4 mmol of NO3-), and NO3--depleted (PL; 70 mL·day-1; 0.04 mmol NO3-) beetroot juice for 5 days. On day 5 of supplementation, subjects completed a series of maximal 20-m sprints followed by the Yo-Yo IR1. Cognitive tasks were completed prior to, during and immediately following the Yo-Yo IR1. RESULTS: BR improved sprint split times relative to PL at 20 m (1.2%; BR 3.98 ± 0.18 vs. PL 4.03 ± 0.19 s; P < 0.05), 10 m (1.6%; BR 2.53 ± 0.12 vs. PL 2.57 ± 0.19 s; P < 0.05) and 5 m (2.3%; BR 1.73 ± 0.09 vs. PL 1.77 ± 0.09 s; P < 0.05). The distance covered in the Yo-Yo IR1 test improved by 3.9% (BR 1422 ± 502 vs. PL 1369 ± 505 m; P < 0.05). The reaction time to the cognitive tasks was shorter in BR (615 ± 98 ms) than PL (645 ± 120 ms; P < 0.05) at rest but not during the Yo-Yo IR1. There was no difference in response accuracy. CONCLUSIONS: Dietary NO3- supplementation enhances maximal sprint and high-intensity intermittent running performance in competitive team sport players. Our findings suggest that NO3- supplementation has the potential to improve performance in single-sprint or multiple-sprint (team) sports.

Dose-dependent effects of dietary nitrate on the oxygen cost of moderate-intensity exercise: Acute vs. chronic supplementation.

PURPOSE: To investigate whether chronic supplementation with a low or moderate dose of dietary nitrate (NO3(-)) reduces submaximal exercise oxygen uptake (V˙O2) and to assess whether or not this is dependent on acute NO3(-) administration prior to exercise. METHODS: Following baseline tests, 34 healthy subjects were allocated to receive 3 mmol NO3(-), 6 mmol NO3(-) or placebo. Two hours following the first ingestion, and after 7, 28 and 30 days of supplementation, subjects completed two moderate-intensity step exercise tests. On days 28 and 30, subjects in the NO3(-) groups completed the test 2 h post consumption of a NO3(-) dose (CHR + ACU) and a placebo dose (CHR). RESULTS: Plasma nitrite concentration ([NO2(-)]) was elevated in a dose-dependent manner at 2 h, 7 days and 28-30 days on the CHR + ACU visit. Compared to pre-treatment baseline, 6 mmol NO3(-) reduced the steady-state V˙O2 during moderate-intensity exercise by 3% at 2 h (P = 0.06), 7 days and at 28-30 days (both P < 0.05) on the CHR + ACU visit, but was unaffected by 3 mmol NO3(-) at all measurement points. On the CHR visit in the 6 mmol group, plasma [NO2(-)] had returned to pre-treatment baseline, but the steady-state V˙O2 remained reduced. CONCLUSION: Up to ∼4 weeks supplementation with 6 but not 3 mmol NO3(-) can reduce submaximal exercise V˙O2. A comparable reduction in submaximal exercise V˙O2 following chronic supplementation with 6 mmol NO3(-) can be achieved both with and without the acute ingestion of NO3(-) and associated elevation of plasma [NO2(-)].

Pancreatic islet vasculature adapts to insulin resistance through dilation and not angiogenesis.

Pancreatic islets adapt to insulin resistance through a complex set of changes, including ß-cell hyperplasia and hypertrophy. To determine if islet vascularization changes in response to insulin resistance, we investigated three independent models of insulin resistance: ob/ob, GLUT4(+/-), and mice with high-fat diet-induced obesity. Intravital blood vessel labeling and immunocytochemistry revealed a vascular plasticity in which islet vessel area was significantly increased, but intraislet vessel density was decreased as the result of insulin resistance. These vascular changes were independent of islet size and were only observed within the ß-cell core but not in the islet periphery. Intraislet endothelial cell fenestration, proliferation, and islet angiogenic factor/receptor expression were unchanged in insulin-resistant compared with control mice, indicating that islet capillary expansion is mediated by dilation of preexisting vessels and not by angiogenesis. We propose that the islet capillary dilation is modulated by endothelial nitric oxide synthase via complementary signals derived from ß-cells, parasympathetic nerves, and increased islet blood flow. These compensatory changes in islet vascularization may influence whether ß-cells can adequately respond to insulin resistance and prevent the development of diabetes.

Long term effects of high fat or high carbohydrate diets on glucose tolerance in mice with heterozygous carnitine palmitoyltransferase-1a (CPT-1a) deficiency: Diet influences on CPT1a deficient mice.

BACKGROUND: Abnormal fatty acid metabolism is an important feature in the mechanisms of insulin resistance and beta-cell dysfunction. Carnitine palmitoyltransferase-1a (CPT-1a, liver isoform) plays a pivotal role in the regulation of mitochondrial fatty acid oxidation. We investigated the role of CPT-1a in the development of impaired glucose tolerance using a mouse model for CPT-1a deficiency when challenged by either a high-carbohydrate (HCD) or a high-fat diet (HFD) for a total duration of up to 46 weeks. METHODS: Insulin sensitivity and glucose tolerance were assessed in heterozygous CPT-1a deficient (CPT-1a+/-) male mice after being fed either a HCD or a HFD for durations of 28 weeks and 46 weeks. Both glucose and insulin tolerance tests were used to investigate beta-cell function and insulin sensitivity. Differences in islet insulin content and hepatic steatosis were evaluated by morphological analysis. RESULTS: CPT-1a+/- mice were more insulin sensitive than CPT-1a+/+ mice when fed either HCD or HFD. The increased insulin sensitivity was associated with an increased expression of Cpt-1b (muscle isoform) in liver, as well as increased microvesicular hepatic steatosis compared to CPT-1a+/+ mice. CPT-1a+/- mice were more glucose tolerant than CPT-1a+/+ mice when fed the HCD, but there was no significant difference when fed HFD. Moreover, CPT-1a+/- mice fed HFD or HCD had fewer and smaller pancreatic islets than CPT-1a+/+ mice. CONCLUSIONS: CPT-1a deficiency preserved insulin sensitivity when challenged by long term feeding of either diet. Furthermore, CPT-1a deficient mice had distinct phenotypes dependent on the diet fed demonstrating that both diet and genetics collectively play a role in the development of impaired glucose tolerance.

Glucose-dependent blood flow dynamics in murine pancreatic islets in vivo.

Pancreatic islets are highly vascularized and arranged so that regions containing beta-cells are distinct from those containing other cell types. Although islet blood flow has been studied extensively, little is known about the dynamics of islet blood flow during hypoglycemia or hyperglycemia. To investigate changes in islet blood flow as a function of blood glucose level, we clamped blood glucose sequentially at hyperglycemic ( approximately 300 mg/dl or 16.8 mM) and hypoglycemic ( approximately 50 mg/dl or 2.8 mM) levels while simultaneously imaging intraislet blood flow in mouse models that express green fluorescent protein in the beta-cells or yellow fluorescent protein in the alpha-cells. Using line scanning confocal microscopy, in vivo blood flow was assayed after intravenous injection of fluorescent dextran or sulforhodamine-labeled red blood cells. Regardless of the sequence of hypoglycemia and hyperglycemia, islet blood flow is faster during hyperglycemia, and apparent blood volume is greater during hyperglycemia than during hypoglycemia. However, there is no change in the order of perfusion of different islet endocrine cell types in hypoglycemia compared with hyperglycemia, with the islet core of beta-cells usually perfused first. In contrast to the results in islets, there was no significant difference in flow rate in the exocrine pancreas during hyperglycemia compared with hypoglycemia. These results indicate that glucose differentially regulates blood flow in the pancreatic islet vasculature independently of blood flow in the rest of the pancreas.

Real-time, multidimensional in vivo imaging used to investigate blood flow in mouse pancreatic islets.

The pancreatic islets of Langerhans are highly vascularized micro-organs that play a key role in the regulation of blood glucose homeostasis. The specific arrangement of endocrine cell types in islets suggests a coupling between morphology and function within the islet. Here, we established a line-scanning confocal microscopy approach to examine the relationship between blood flow and islet cell type arrangement by real-time in vivo imaging of intra-islet blood flow in mice. These data were used to reconstruct the in vivo 3D architecture of the islet and time-resolved blood flow patterns throughout the islet vascular bed. The results revealed 2 predominant blood flow patterns in mouse islets: inner-to-outer, in which blood perfuses the core of beta cells before the islet perimeter of non-beta cells, and top-to-bottom, in which blood perfuses the islet from one side to the other regardless of cell type. Our approach included both millisecond temporal resolution and submicron spatial resolution, allowing for real-time imaging of islet blood flow within the living mouse, which has not to our knowledge been attainable by other methods.

Homozygous carnitine palmitoyltransferase 1a (liver isoform) deficiency is lethal in the mouse.

To better understand carnitine palmitoyltransferase 1a (liver isoform, gene=Cpt-1a, protein=CPT-1a) deficiency in human disease, we developed a gene knockout mouse model. We used a replacement gene targeting strategy in ES cells that resulted in the deletion of exons 11-18, thus producing a null allele. Homozygous deficient mice (CPT-1a -/-) were not viable. There were no CPT-1a -/- pups, embryos or fetuses detected from day 10 of gestation to term. FISH analysis demonstrated targeting vector recombination at the expected single locus on chromosome 19. The inheritance pattern from heterozygous matings was skewed in both C57BL/6NTac, 129S6/SvEvTac (B6;129 mixed) and 129S6/SvEvTac (129 coisogenic) genetic backgrounds biased toward CPT-1a +/- mice (>80%). There was no sex preference with regard to germ-line transmission of the mutant allele. CPT-1a +/- mice had decreased Cpt-1a mRNA expression in liver, heart, brain, testis, kidney, and white fat. This resulted in 54.7% CPT-1 activity in liver from CPT-1a +/- males but no significant difference in females as compared to CPT-1a +/+ controls. CPT-1a +/- mice showed no fatty change in liver and were cold tolerant. Fasting free fatty acid concentrations were significantly elevated, while blood glucose concentrations were significantly lower in 6-week-old CPT-1a +/- mice compared to controls. Although the homozygous mutants were not viable, we did find some aspects of haploinsufficiency in the CPT-1a +/- mutants, which will make them an important mouse model for studying the role of CPT-1a in human disease.

Estradiol may limit lipid oxidation via Cpt 1 expression and hormonal mechanisms.

OBJECTIVE: Evidence indicates that estrogen depresses hepatic lipid oxidation. We tested the hypothesis that estradiol (E(2)) treatment depresses transcription of carnitine palmitoyltransferase-1 (Cpt 1) mRNA and increases adiposity. RESEARCH METHODS AND PROCEDURES: Six ovariectomized female rats were given a subcutaneous pellet of E(2) (5 mg/d), and six were given placebo. Rats were pair-fed by group for 18 days. Body composition was assessed chemically: mRNA for liver Cpt 1, adipose tissue uncoupling protein-2 (Ucp 2), and quadriceps Ucp 3 by Northern analysis; serum glucose, triglycerides (TGs), and free fatty acids by standard techniques; and serum insulin and glucagon by radioimmunoassay. RESULTS: E(2)-treated rats lost more weight than placebo-treated rats (37.3 +/- 6.0 vs. 16.2 +/- 2.6 g, p < 0.01), but did not differ in final carcass composition (adjusted for eviscerated body mass). E(2)-treated rats had lower liver Cpt 1 (p < 0.001) and skeletal muscle Ucp 3 (p < 0.05) mRNA and lower concentrations of glucose, glucagon, and free fatty acids (p < 0.05). E(2)-treated rats tended to have higher insulin (p = -0.067) and TG (p = 0.097). TG tended to be correlated with Cpt 1 mRNA (r = -0.56 and p = 0.07). DISCUSSION: These results suggest that, although E(2) is likely to suppress lipid oxidation and promote TG synthesis, these effects are not manifested in a relative increase in carcass adiposity after 18 days of treatment, at least under conditions of negative energy balance. The possible role of E(2)-mediated changes in insulin and glucagon secretion on hepatic substrate metabolism warrants further study.