Male Otsuka Long-Evans Tokushima Research Articles

PDE5 inhibitor potentially improves polyuria and bladder storage and voiding dysfunctions in type 2 diabetic rats.

Bladder dysfunction associated with type 2 diabetes mellitus (T2DM) includes urine storage and voiding disorders. We examined pathological conditions of the bladder wall in a rat T2DM model and evaluated the effects of the phosphodiesterase-5 (PDE-5) inhibitor tadalafil. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats and Long-Evans Tokushima Otsuka (LETO) rats were used as the T2DM and control groups, respectively. Tadalafil was orally administered for 12 weeks. Micturition behavior was monitored using metabolic cages, and bladder function was evaluated by cystometry. Bladder blood flow was evaluated by laser speckle imaging, and an organ bath bladder distention test was used to measure adenosine triphosphate (ATP) release from the bladder urothelium. The expression levels of vesicular nucleotide transporter (VNUT), hypoxia markers, pro-inflammatory cytokines and growth factors in the bladder wall were measured using real-time polymerase chain reaction and enzyme-linked immunosorbent assay. Bladder wall contractions in response to KCl and carbachol were monitored using bladder-strip tests. With aging, OLETF rats had higher micturition frequency and greater urine volume than LETO rats. Although bladder capacity was not significantly different, non-voiding bladder contraction occurred more frequently in OLETF rats than in LETO rats. Bladder blood flow was decreased and ATP release was increased with higher VNUT expression in OLETF rats than in LETO rats. These effects were suppressed by tadalafil administration, with accompanying decreased HIF-1α, 8-OHdG, IL-6, TNF-α, IGF-1, and bFGF expression. The impaired contractile responses of bladder strips to KCl and carbachol in OLETF rats with aging were restored by tadalafil administration. The T2DM rats had polyuria, increased ATP release induced by decreased bladder blood flow and impaired contractile function. PDE5 inhibition improved these changes and may prevent T2DM-associated urinary frequency and bladder storage and voiding dysfunctions.

Detraining after short-term exercise induces hyperphagia and obesity with fatty liver and brown adipose tissue whitening in young male OLETF rats.

This study examined the effects of exercise and detraining at a young age on fat accumulation in various organs. Four-week-old male Otsuka Long-Evans Tokushima Fatty (OLETF) rats were assigned to either the non-exercise sedentary (OLETF Sed) or exercise groups. The exercise group was subdivided into two groups: exercise between 4 and 12 weeks of age (OLETF Ex) and exercise between 4 and 6 weeks of age followed by non-exercise between 6 and 12 weeks of age (OLETF DT). Body weight was significantly lower in the OLETF Ex group than in the OLETF Sed group at 12 weeks of age. Fat accumulation in the epididymal white adipose tissue, liver, and brown adipose tissue was suppressed in the OLETF Ex group. During the exercise period, body weight and food intake in the OLETF DT group were significantly lower than those in the OLETF Sed group. However, food intake was significantly higher in the OLETF DT group than in the OLETF Sed group after exercise cessation, resulting in extreme obesity with fatty liver and brown adipose tissue whitening. Detraining after early-onset exercise promotes hyperphagia, causing extreme obesity. Overeating should be avoided during detraining periods in cases of exercise cessation at a young age.

Exercise training improves obesity-induced inflammatory signaling in rat brown adipose tissue

Chronic inflammation is considered as an etiology of obesity and type 2 diabetes. Brown adipose tissue (BAT) of obese animals shows increased inflammation. Regular exercise has anti-inflammatory effects; however, the effects of exercise training on BAT inflammation in obese animals remain unclear. Thus, this study aimed to investigate the effects of exercise training on inflammation-related signaling in the BAT of obese and diabetic rats. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats, an obese/diabetic rodent model, were randomly divided into either sedentary (n = 11) or exercise training (n = 8) groups. Long-Evans Tokushima Otsuka (LETO; n = 9) rats were used as the nondiabetic sedentary controls. Exercise training using a treadmill was conducted 4 days per week for 20 weeks, starting at 5 weeks old. As a result, exercise training attenuated the phosphorylation levels of p65 and mitogen-activated protein kinases in the BAT of OLETF rats, concurrently with the improvement of obesity and systemic glucose tolerance. Moreover, exercise training decreased oxidative stress and increased the antioxidant and anti-inflammatory protein levels in the BAT. Conversely, exercise training did not alter the expression levels of uncoupling protein-1 and oxidative phosphorylation-related proteins in the BAT, which were lower in the OLETF rats than the LETO rats. In conclusion, our data suggest that exercise training prevents the activation of inflammatory signaling in the BAT of obese/diabetic rats.

Effects of long-term childhood exercise and detraining on lipid accumulation in metabolic-related organs.

The preventive effects of regular exercise on obesity-related health problems are carried over to the non-exercise detraining period, even when physical activity decreases with aging. However, it remains unknown whether regular childhood exercises can be carried over to adulthood. Therefore, this study aimed to investigate the effects of long-term childhood exercise and detraining on lipid accumulation in organs to prevent obesity in adulthood. Four-week-old male Otsuka Long-Evans Tokushima Fatty (OLETF) rats were used as obese animals. OLETF rats were allocated into sedentary and exercise groups: exercise from 4- to 12-week-old and detraining from 12- to 20-week-old. At 12-week-old immediately after the exercise period, regular exercise completely inhibited hyperphagia, obesity, enlarged pancreatic islets, lipid accumulation and lobular inflammation in the liver, hypertrophied adipocytes in the white adipose tissue (WAT), and brown adipose tissue (BAT) whitening in OLETF rats. Additionally, exercise attenuated the decrease in the ratio of muscle wet weight to body weight associated with obesity. Decreased food consumption was maintained during the detraining period, which inhibited obesity and diabetes at 20-week-old after the detraining period. Histologically, childhood exercise inhibited the enlargement of pancreatic islets after the detraining period. In addition, inhibition of lipid accumulation was completely maintained in the WAT and BAT after the detraining period. However, the effectiveness was only partially successful in lipid accumulation and inflammation in the liver. The ratio of muscle wet weight to body weight was maintained after detraining. In conclusion, early long-term regular exercise effectively prevents obesity and diabetes in childhood, and its effectiveness can be tracked later in life. The present study suggests the importance of exercise during childhood and adolescence to inhibit hyperphagia-induced lipid accumulation in metabolic-related organs in adulthood despite exercise cessation.

Phosphodiesterase 5 inhibitor suppresses prostate weight increase in type 2 diabetic rats

Aims: Hyperinsulinemia is an important causative factor of prostate enlargement in type 2 diabetes (T2D), however, clinically prostate weight increases during hypoinsulinemic condition. To investigate the pathogenesis of prostate enlargement and effects of phosphodiesterase 5 inhibitor (PDE5i), male Otsuka Long-Evans Tokushima Fatty (OLETF) and Long-Evans Tokushima Otsuka (LETO) rats were used as T2D and control, respectively.Materials and Methods: OLETF and LETO rats were treated with oral tadalafil (100 μg/kg/day) or vehicle for 12 wks from at the age of 36 wks.Key findings: Prostate weight of OLETF rats was significantly higher than that of LETO at 36 wks, and increased at 48 wks. In OLETF rats, prostate blood flow was significantly lower at 48 wks versus 36 wks. Twelve-week-tadalafil treatment increased prostate blood flow and suppressed prostate weight increase in both strains. This change was inversely correlated with changes in prostate expressions of hypoxia-inducible factor-1 alpha (HIF-1α) and 8-hydroxy-2′-deoxyguanosine (8-OHdG). Increases with age were observed in mRNA and/or protein levels of cytokines interleukin (IL)-6, IL-8, and tumor necrosis factor-alpha (TNF-α) and cell growth factors insulin-like growth factor-1 (IGF-1), basic fibroblast growth factor (bFGF), and transforming growth factor-beta (TGF-β); especially IL-6, TNF-α, IGF-1, bFGF and TGF-β increased with T2D. Tadalafil suppressed these cytokines and growth factors.Significance: These data suggest chronic ischemia caused by T2D leads to oxidative stress, resulting in prostate enlargement through upregulation of several cytokines and growth factors. Treatment with PDE5i improves prostate ischemia and might prevent enlargement via suppression of cytokines and growth factors in T2D.

Impact of Exercise and Detraining during Childhood on Brown Adipose Tissue Whitening in Obesity.

This study aimed to investigate the influence of childhood exercise and detraining on brown adipose tissue (BAT) whitening in obesity. Four-week-old male Long-Evans Tokushima Otsuka (LETO) rats (n = 9) and Otsuka Long-Evans Tokushima Fatty (OLETF) rats (n = 24) were used as non-obese and obese animals, respectively. OLETF rats were divided into non-exercise sedentary (n = 9) and exercise groups. OLETF rats in the exercise group were further divided into subgroups according to the exercise period—exercise from 10- to 12-weeks-old (n = 6); and exercise from 4- to 6-weeks-old, and detraining from 6- to 12-weeks-old (n = 9). At 12-weeks-old, immediately after exercise period, BAT whitening in OLETF rats was inhibited by exercise despite the fact that hypertrophy was not caused in the plantaris muscle. However, the effectiveness was attenuated during the detraining period. Histological BAT whitening and downregulation of uncoupling protein-1 (UCP-1) were found in non-exercise sedentary OLETF rats at 12-weeks-old. The downregulation was not inhibited, even though exercise histologically inhibited BAT whitening in OLETF rats. Childhood exercise decreased BAT whitening in obesity. Detraining attenuated the inhibition of BAT whitening. These results suggest that regular exercise is needed to improve BAT whitening and downregulation of UCP-1 in obesity.

Effects of Aerobic Exercise Training on Mitochondrial Ca2+ Homeostasis and Apoptosis in the Hippocampus of Type 2 Diabetic Rats

PURPOSE: Type 2 diabetes mellitus (T2DM) is a recognized risk factor for neurodegenerative diseases such as Alzheimer’s disease (AD). Mitochondrial dysfunction caused by disturbance of calcium (Ca2+) homeostasis might be a pathophysiological link between T2DM and AD. In the present study, we investigated the effects of aerobic training on the regulation of mitochondrial calcium homeostasis, mitochondrial function (O<sub>2</sub> respiration), and apoptosis in the hippocampus of T2DM rats.METHODS:Thirty male Otsuka Long-Evans Tokushima Fatty rats were divided into three groups: control group (CON, n=10), diabetes control group (DM, n=10), and diabetes+aerobic exercise group (DM+EXE, n=10). The rats in the exercise group were forced to run on a treadmill for 30 min, three times a week for 8 weeks.RESULTS:The DM group showed significantly increased mitochondrial calcium uniporter (MCU) protein and apoptosis-related factors such as Bax and caspase-3 and decreased mitochondrial O<sub>2</sub> respiration when compared with the CON group. However, the hippocampus of rats in the DM+EXE group exhibited a significant decrease in MCU, Bax, and caspase-3 levels and an increase in Bcl-2 and mitochondrial O<sub>2</sub> respiration when compared with the DM group.CONCLUSIONS:These results suggest that regular aerobic exercise alleviates mitochondrial dysfunction and apoptosis by modulating MCU, which is involved in mitochondrial Ca2+ homeostasis in the early stages of T2DM. This might be a potent strategy to prevent neurodegenerative diseases.

Effects of exercise on browning of adipose tissue in OLETF rat

Browning of subcutaneous white adipose tissue (WAT) is induced by stimulation of β3 adrenergic receptor agonist with cold exposure and exercise training. However, it is remained unknown whether browning of adipose tissue equally inducing between obesity and non-obesity. The purpose of the present study was to investigate the effects of exercise on browning of adipose tissue in obesity. Male Otsuka Long-Evans Tokushima fatty (OLETF) rats and Long-Evans Tokushima Otsuka (LETO) rats were used as models of obesity and non-obesity, respectively. The rats were divided into exercise and sedentary groups. The rats of exercise groups were placed in the cages with a running wheel for 12 h (20:00 to 8:00 h) daily from 19 to 21 weeks of age. Food and water were provided ad libitum. There were no significant differences in the amount of voluntary wheel running between animal strains. Although there were no significant differences in the expression level of uncoupling protein 1 (UCP1) of brown adipose tissue (BAT) between the exercise and sedentary groups of LETO rats, the expression levels were significantly higher in the exercise group than the sedentary group of OLETF rats. In the inguinal WAT, the expression levels of UCP1 were below the minimum detectable threshold among all groups in both strains. These results suggest differential sensitivity of activation of UCP1 in BAT between obesity and non-obesity. In addition, one of the reasons for undetectable exercise-induced UCP1 expression in the WAT could be due to the low intensity by voluntary wheel running. This study was approved by the Institutional Animal Care and Use Committee of Hiroshima University (A19-163) and conducted in accordance with the Hiroshima University Regulations for Animal Experimentation. All experiments complied with the National Institute of Health Guidelines for the Care and Use of Laboratory Animals.

Influences of exercise and detraining during childhood on brown adipose tissue in OLETF rats

Brown adipose tissue (BAT) oxidizes lipids to fuel thermogenesis. However, the activity declines in obesity due to brown-to-white conversion in BAT. Because the properties of adipose tissue are determined before adult, childhood exercise could prevent brown-to-white conversion and maintain metabolic activity of BAT in future. The purpose of the present study was to investigate histological influences of childhood exercise and detraining in BAT of genetic predisposition to obesity. Four-week-old male Long-Evans Tokushima Otsuka (LETO) rats and age-matched male Otsuka Long-Evans Tokushima fatty (OLETF) rats were used as non-obesity animals and spontaneous obesity animals, respectively. OLETF rats were divided into the non-exercise sedentary (OLETF Sed) or exercise groups. OLETF rats in the exercise group were further divided into subgroups according to exercise age. OLETF rats in the exercise groups were placed in the cages with a running wheel for 12 h (20:00 to 8:00) daily from 4 to 6 (OLETF Ex 4-6 w) or 10 to 12 (OLETF Ex 10-12 w) weeks of age (i.e. the OLETF Ex 4-6 w group: exercise from 4 to 6-week-old, detraining from 6 to 12-week-old; the OLETF Ex 10-12 w group: non-exercise from 4 to 10-week-old, exercise from 10 to 12-week-old). Food and water were provided ad libitum. At 12-week-old, transverse sections were obtained from interscapular BAT and stained with hematoxylin and eosin for histological observation. The results at 12-week-old were shown below. The wet weights of interscapular BAT were significantly higher in the OLETF Sed group than in LETO rats. The values were significantly lower in the OLETF Ex 10-12 w group than in the OLETF Sed group. There were no significant differences for the weights between the OLETF Ex 4-6 w and OLETF Sed groups. Histological observation revealed conversion of brown adipocytes to white-like unilocular cells in both LETO and OLETF rats. In LETO rats, the white-like unilocular cells were located uniformly in the fat lobule. In OLETF rats, enlarged white-like unilocular cells were also observed in the peripheral zone of the lobule. There were no significant differences for the diameters of white-like unilocular cells located in the central zone of the lobule among the experimental groups. The densities of white-like unilocular cells located in the central zone of the lobule were significantly lower in the OLETF Ex 10-12 w group than in the other groups. There were no significant differences for the densities among LETO rats, the OLETF Sed, and OLETF Ex 4-6 w groups. Childhood Exercise immediately inhibited brown-to-white conversion in the BAT and decreased the mass. However, the effectiveness did not be maintained for a long time. These results suggested that chronic exercise is required to maintain adipose tissue browning in genetic predisposition to obesity.

Influences of childhood exercise on adipose tissue browning in OLETF rats

Exercise promotes mitochondrial uncoupling in brown adipose tissue (BAT), which results in browning and accelerated energy metabolism. Because obesity leads to mitochondrial dysfunction with consequential impaired uncoupling, exercise-induced adipose tissue browning could be inhibited in genetic predisposition to obesity. The purpose of the present study was to investigate the influences of childhood exercise in spontaneous obesity animals focusing on adipose tissue browning. Four-week-old male Otsuka Long-Evans Tokushima fatty (OLETF, 88 ± 4 g body weight) rats and age-matched male Long-Evans Tokushima Otsuka (LETO, 82 ± 4 g body weight) rats were used as spontaneous obesity animals and non-obesity animals, respectively. All rats were divided into the exercise or non-exercise groups. The rats in the exercise groups were placed in the cages with a running wheel for 12 h (20:00 to 8:00) daily from 4 to 6 weeks of age. Food and water were provided ad libitum. Transverse sections were obtained from interscapular BAT and stained with hematoxylin and eosin for histological observation. In addition, the expression levels of uncoupling protein 1 (UCP1) in BAT were measured by western blotting. White adipocytes with unilocular lipid droplet were widely observed in the BAT of OLETF rats. The enlarged white adipocytes were rarely observed in OLETF rats of the exercise group. Although there were no significant differences in the expression level of UCP1 among the experimental groups, the levels were obviously different between OLETF rats in the exercise group. Total food intake during 12 h of voluntary wheel running was lower in the rat with the high UCP1 expression than in the low expression. Total running distance was higher in the rat with high UCP1 expression than in the low expression. Therefore, higher ratio of the total running distance to total food intake was shown in the OLETF rat with higher UCP1 expression. Exercise in childhood shrank the adipocytes in the BAT, suggesting the effectiveness to prevent pediatric obesity. Additionally, exercise-induced UCP1 expression could depend on the ratio of exercise to food intake.

Exercise and Curcumin in Combination Improves Cognitive Function and Attenuates ER Stress in Diabetic Rats.

Type 2 diabetes mellitus (T2DM) is a metabolic disease associated with chronic low-grade inflammation that is mainly associated with lifestyles. Exercise and healthy diet are known to be beneficial for adults with T2DM in terms of maintaining blood glucose control and overall health. We investigated whether a combination of exercise and curcumin supplementation ameliorates diabetes-related cognitive distress by regulating inflammatory response and endoplasmic reticulum (ER) stress. This study was performed using male Otsuka Long-Evans Tokushima Fatty (OLETF) rats (a spontaneous diabetes Type 2 model) and Long-Evans Tokushima Otsuka (LETO) rats (LETO controls) by providing them with exercise alone or exercise and curcumin in combination. OLETF rats were fed either a diet of chow (as OLETF controls) or a diet of chow containing curcumin (5 g/kg diet) for five weeks. OLETF rats exercised with curcumin supplementation exhibited weight loss and improved glucose homeostasis and lipid profiles as compared with OLETF controls or exercised OLETF rats. Next, we examined cognitive functions using a Morris water maze test. Exercise plus curcumin improved escape latency and memory retention compared to OLETF controls. Furthermore, OLETF rats exercised and fed curcumin had lower IL6, TNFα, and IL10 levels (indicators of inflammatory response) and lower levels of ER stress markers (BiP and CHOP) in the intestine than OLETF controls. These observations suggest exercise plus curcumin may offer a means of treating diabetes-related cognitive dysfunction.

Correcting imbalance of sex hormones by a phosphodiesterase 5 inhibitor improves copulatory dysfunction in male rats with type 2 diabetes

IntroductionSexual dysfunction is a common complication in men with type 2 diabetes and is often refractory to treatment. This study investigated the long-term influence of the phosphodiesterase 5 inhibitor (PDE5I)...

Hyperbaric Treatment With Normal Air Prevents The Progression Of Hyperglycemia In Type 2 Diabetes

Previous study reported that hyperbaric treatment with 36% oxygen decreases glucose and insulin level in type 2 diabetes rats, and the improvement could be due to increased oxygen saturation and blood flow during hyperbaric treatment. However, the effects of simple hyperbaric treatment without high oxygen remain unknown. PURPOSE: To investigate the effects of hyperbaric treatment with normal air on hyperglycemia in type 2 diabetes, focusing on skeletal muscle hemodynamics. METHODS: 24-week-old male Otsuka Long-Evans Tokushima fatty (OLETF) rats and Long-Evans Tokushima Otsuka (LETO) rats were used as diabetes model and non-diabetes model, respectively. All rats were assigned to hyperbaric treatment or non-treatment groups. The rats in the hyperbaric treatment group were exposed to hyperbaric chamber at 1.3 ATA with normal air for 8 hours a day for 16 weeks. The oxygen saturation and total-hemoglobin (Hb) changes in the calf muscle during hyperbaric treatment was measured by near-infrared spectroscopy. Oral glucose tolerance test was performed at 40-week-old. RESULTS: Oxygen saturation and total-Hb were significantly increased during hyperbaric treatment from 73.3 to 76.7% and 25.0 to 26.3×104/mm3 in OLETF rats, 71.1 to 74.8% and 20.1 to 22.5×104/mm3 in LETO rats (p<0.05). The glucose and insulin levels were significantly higher in OLETF rats than LETO rats at both fasting and after glucose administration (p<0.05). Among OLETF rats, the glucose levels at 30, 60, 120 min after glucose administration were significantly lower in the hyperbaric treatment group than the non-treatment group (30 min: 325 ± 71 vs. 385 ± 48, 60 min: 332 ± 67 vs. 421 ± 111, 120 min: 216 ± 45 vs. 230 ± 20 mg/dL, p<0.05). Additionally, the fasting insulin level and the levels at 120 min after glucose administration were significantly lower in the hyperbaric treatment group than the non-treatment group (Fasting: 3.6 ± 1.1 vs. 4.3 ± 2.7, 120 min: 4.4 ± 1.6 vs. 5.2 ± 3.9 ng/mL, p<0.05). CONCLUSIONS: The present study demonstrated that hyperbaric treatment with normal air also prevents the progression of hyperglycemia in OLETF rats, and the treatment without high oxygen increases oxygen saturation and blood flow in the skeletal muscle.

SAT-173 Antiproliferative Effect of Teneligliptin on Atherosclerosis in Vascular Smooth Muscle Cell

Background and aims: In vivo and in vitro studies revealed that sitagliptin, linagliptin treatment suppressed proliferation of vascular smooth muscle cells (VSMCs). However, data are lacking regarding the effects of teneligliptin. Here, we demonstrate the effects of teneligliptin on VSMC proliferation and migration according to fluctuating glucose levels. Materials and methods: Primary cultures of male Otsuka Long-Evans Tokushima fatty (OLETF) rat VSMCs were obtained from enzymatically dissociated rat thoracic aorta. VSMCs with teneligliptin (100 μM) were incubated for 72 h with alternating normal (5.5 mmol/L) and high glucose (25 mmol/L) media every 12 h. The proliferation of VSMCs, proliferative and apoptotic molecular pathway and the migration of VSMC were analyzed with teneligliptin. Results: In MTT assay, teneligliptin attenuated the increase in cells pretreated with 100-μM teneligliptin 72 h prior to glucose fluctuation (F+T group) in comparison with fluctuation group (F group) (p-value, 0.005). The protein levels of phospho-MEK1/2 and phospho-ERK1/2 were significantly reduced in F+T group compared with F group (phospho-MEK1/2, p-value, 0.016; phospho-ERK1/2, p-value, 0.001). However no changes in the protein levels of caspase 3, Bcl-2, and phospho-Bad, Bax, Bcl-xL, were observed. In wound healing assay, migrated cell percentage was attenuated in F+T group (p-value, 0.043). phospho-BMK protein levels was also reduced in F+T group (p-value, 0.017). Conclusion: This study has demonstrated that teneligliptin significantly reduces the proliferation and migration of VSMCs in intermittent hyperglycemic condition.

Long-term insulin treatment leads to a change in myosin heavy chain fiber distribution in OLETF rat skeletal muscle.

The objective of this study was to investigate molecular and physiological changes in response to long-term insulin glargine treatment in the skeletal muscle of OLETF rats. Male Otsuka Long-Evans Tokushima Fatty (OLETF) and Long-Evans Tokushima Otsuka (LETO) rats aged 24 weeks were randomly allocated to either treatment with insulin for 24 weeks or no treatment, resulting in three groups. Insulin glargine treatment in OLETF rats (OLETF-G) for 24 weeks resulted in changes in blood glucose levels in intraperitoneal glucose tolerance tests compared with age-matched, untreated OLETF rats (OLETF-C), and the area under the curve was significantly decreased for OLETF-G rats compared with OLETF-C rats (P < 0.05). The protein levels of MHC isoforms were altered in gastrocnemius muscle of OLETF rats, and the proportions of myosin heavy chain type I and II fibers were lower and higher, respectively, in OLETF-G compared with OLETF-C rats. Activation of myokines (IL-6, IL-15, FNDC5, and myostatin) in gastrocnemius muscle was significantly inhibited in OLETF-G compared with OLETF-C rats ( P < 0.05). MyoD and myogenin levels were decreased, while IGF-I and GLUT4 levels were increased, in the skeletal muscle of OLETF-G rats ( P < 0.05). Insulin glargine treatment significantly increased the phosphorylation levels of AMPK, SIRT1, and PGC-1α. Together, our results suggested that changes in the distribution of fiber types by insulin glargine could result in downregulation of myokines and muscle regulatory proteins. The effects were likely associated with activation of the AMPK/SIRT1/PGC-1α signaling pathway. Changes in these proteins may at least partly explain the effect of insulin in skeletal muscle of diabetes mellitus.

Effects of voluntary running exercise on bone histology in type 2 diabetic rats.

The incidence of obesity in children and adolescents, which may lead to type 2 diabetes, is increasing. Exercise is recommended to prevent and improve diabetes. However, little is known about the bone marrow environment at the onset of diabetes in the young, and it is unclear whether exercise training is useful for maintaining bone homeostasis, such as mechanical and histological properties. Thus, this study clarified the histological properties of bone and whether exercise contributes to maintaining bone homeostasis at the onset of type 2 diabetes in rats. Four-week-old male Otsuka Long-Evans Tokushima Fatty (OLETF; n = 21) rats as a diabetic model and Long-Evans Tokushima Otsuka (LETO; n = 18) rats as a control were assigned randomly to four groups: the OLETF sedentary group (O-Sed; n = 11), OLETF exercise group (O-Ex; n = 10), LETO sedentary group (L-Sed; n = 9), and LETO exercise group (L-Ex; n = 9). All rats in the exercise group were allowed free access to a steel running wheel for 20 weeks (5–25 weeks of age). In the glucose tolerance test, blood glucose level was higher in the O-Sed group than that in the L-Sed and L-Ex groups, and was markedly suppressed by the voluntary running exercise of O-Ex rats. The energy to fracture and the two-dimensional bone volume at 25 weeks of age did not differ significantly among the groups, though the maximum breaking force and stiffness were lower in OLETF rats. However, bone marrow fat volume was greater in O-Sed than that in L-Sed and L-Ex rats, and was markedly suppressed by wheel running in the O-Ex rats. Our results indicate that exercise has beneficial effects not only for preventing diabetes but also on normal bone remodeling at an early age.

Impaired endothelium-derived hyperpolarization-type relaxation in superior mesenteric arteries isolated from female Otsuka Long-Evans Tokushima Fatty rats

Endothelium-derived hyperpolarization (EDH) is an important signaling mechanism of endothelium-dependent vasorelaxation, and little attention has been paid to the EDH-type responses in female metabolic syndrome such as that observed with type-2 diabetes. We previously reported that EDH-type relaxation was impaired in superior mesenteric arteries from male Otsuka Long-Evans Tokushima Fatty (OLETF) rat, a model of type-2 diabetes, however, the response was unclear in female OLETF rat. Thus, the aim of this study was to examine if EDH-type relaxation was altered in superior mesenteric arteries isolated from female OLETF rats compared to age-matched, control female Long-Evans Tokushima Otsuka (LETO) rats at age 50–59 weeks. We investigated concentration–relaxation curves for acetylcholine (at age 50–53 weeks), NS309 (an activator of small- and intermediate-conductance calcium-activated potassium channels) (at age 50–53 weeks), and GSK1016790A (an agonist of transient receptor potential vanilloid type 4, TRPV4) (at age 58 or 59 weeks) in the presence of the nitric oxide synthase inhibitor NG-nitro-L-arginine and the cyclooxygenase inhibitor indomethacin to investigate EDH-type responses in the superior mesenteric artery. Obesity, mild hyperglycemia, hyperinsulinemia, and hyperlipidemia (i.e., increased total cholesterol, triglyceride, and non-esterified fatty acids) were more frequent in OLETF rats than in age-matched LETO rats at age 50–53 weeks. Acetylcholine-, NS309-, and GSK1016790A-induced relaxations in arteries from OLETF rats were all significantly reduced compared to those in LETO rats. These results indicated that EDH-type relaxations were impaired in female OLETF rats. This novel experimental model may provide new insights into vascular dysfunction in metabolic syndrome in females.

Combined long-term caffeine intake and exercise inhibits the development of diabetic nephropathy in OLETF rats.

This study was performed to examine the effects of long-term caffeine-intake, with and without exercise, on the progression of diabetic nephropathy (DN) in an obese diabetic rat model. Thirty-two male Otsuka Long-Evans Tokushima fatty (OLETF) rats were assigned to sedentary (OLETF-Sed), exercise (OLETF-Ex), caffeine-intake (OLETF-Caf), and combined (OLETF-Caf + Ex) groups. Caffeine-intake groups were fed rat chow containing caffeine (90.7 ± 4.7 mg/kg/day). The OLETF-Ex and OLETF-Caf + Ex groups were able to run voluntarily at any time using a rotatory wheel. Body weight (BW) and blood pressure (BP) were measured weekly from 24 to 29 wk of age. Pre- and posttreatment serum glucose, insulin, and creatinine concentrations were measured, and a 24 h urine sample was collected for measurement of creatinine clearance (Ccr) and albumin excretion (UEAlb). After treatment, the kidneys were removed for morphological analysis. The OLETF-Caf and OLETF-Caf + Ex groups exhibited no BP increase during the study. Both the caffeine-intake groups exhibited a significant increase in urine volume (UV), electrolyte excretion, and Ccr, and decreased UEAlb, following treatment. Furthermore, no structural damage was observed in the kidneys of rats from either caffeine-intake group, whereas the OLETF-Sed and OLETF-Ex groups exhibited DN progression. This study demonstrates that caffeine-intake alone and/or combined with exercise significantly decreases BW and improves glucose intolerance, without the progression of DN. Further research should be performed to examine whether the quantities of caffeine contained in a normal human daily intake also have a protective effect against kidney damage.NEW & NOTEWORTHY The present study showed that caffeine administration alone and/or combined with exercise results in an improvement of diabetic nephropathy (DN), including an increase in creatinine clearance and urinary Na excretion, a decrease in urinary protein excretion, and in renal morphological findings. To our knowledge, there are no other studies showing that caffeine administration inhibits DN progression.

Augmented Contractility to Noradrenaline in Femoral Arteries from the Otsuka Long-Evans Tokushima Fatty Rat, a Model of Type 2 Diabetes

Although vasculopathies may occur systemically, there are few reports regarding femoral arteries of type 2 diabetes. Here, we investigated whether contractile response to noradrenaline in femoral arteries would change in type 2 diabetic male Otsuka Long-Evans Tokushima Fatty (OLETF) rat at the chronic stage of disease (1 year old) versus age-matched control Long-Evans Tokushima Otsuka (LETO) rat. OLETF rat exhibited hyperglycemia, hypertension, hyperlipidemia, and hypoinsulinemia compared to age-matched LETO rat. Noradrenaline-induced contraction was increased in femoral arteries in OLETF rats compared with LETO rats whereas serotonin- or phenylephrine-induced contractions were similar between these two animals. Acetylcholine- and sodium nitroprusside-induced relaxations were similar between the two groups. Very small relaxations in femoral arteries induced by clonidine and isoprenaline were obtained in LETO but not OLETF group. Noradrenaline-induced contraction was enhanced by treatment with NG-nitro-L-arginine (L-NNA), a nitric oxide synthase (NOS) inhibitor, and the between-group difference of contraction was eliminated by such treatment. Indomethacin, a non-selective cyclooxygenase (COX) inhibitor, reduced noradrenaline-induced contraction in both groups, whereas the contraction was greater in OLETF group versus LETO. Femoral arterial protein expression of endothelial NOS, COX-1, and superoxide dismutases were similar between the two groups, whereas reduction of COX-2 expression was seen in OLETF group compared with LETO. Increased contractile responsiveness to noradrenaline is seen in OLETF rat femoral artery and this may be due to reduction of suppressive effect of NO.

Anti-Proliferative Effects of Rutin on OLETF Rat Vascular Smooth Muscle Cells Stimulated by Glucose Variability

PurposeProliferation of vascular smooth muscle cells (VSMCs) plays a crucial role in atherosclerosis. Rutin is a major representative of the flavonol subclass of flavonoids and has various pharmacological activities. Currently, data are lacking regarding its effects on VSMC proliferation induced by intermittent hyperglycemia. Here, we demonstrate the effects of rutin on VSMC proliferation and migration according to fluctuating glucose levels.Materials and MethodsPrimary cultures of male Otsuka Long-Evans Tokushima Fatty (OLETF) rat VSMCs were obtained from enzymatically dissociated rat thoracic aortas. VSMCs were incubated for 72 h with alternating normal (5.5 mmol/L) and high (25.0 mmol/L) glucose media every 12 h. Proliferation and migration of VSMCs, the proliferative molecular pathway [including p44/42 mitogen-activated protein kinases (MAPK), mitogen-activated protein kinase kinase 1/2 (MEK1/2), p38 MAPK, phosphoinositide 3-kinase (PI3K), c-Jun N-terminal protein kinase (JNK), nuclear factor kappa B (NF-κB), and Akt], the migratory pathway (big MAPK 1, BMK1), reactive oxygen species (ROS), and apoptotic pathway were analyzed.ResultsWe found enhanced proliferation and migration of VSMCs when cells were incubated in intermittent high glucose conditions, compared to normal glucose. These effects were lowered upon rutin treatment. Intermittent treatment with high glucose for 72 h increased the expression of phospho-p44/42 MAPK (extracellular signal regulated kinase 1/2, ERK1/2), phospho-MEK1/2, phospho-PI3K, phospho-NF-κB, phospho-BMK1, and ROS, compared to treatment with normal glucose. These effects were suppressed by rutin. Phospho-p38 MAPK, phospho-Akt, JNK, and apoptotic pathways [B-cell lymphoma (Bcl)-xL, Bcl-2, phospho-Bad, and caspase-3] were not affected by fluctuations in glucose levels.ConclusionFluctuating glucose levels increased proliferation and migration of OLETF rat VSMCs via MAPK (ERK1/2), BMK1, PI3K, and NF-κB pathways. These effects were inhibited by the antioxidant rutin.