The Toughness-Enhanced Atelocollagen Double-Network Gel for Biomaterials.

A tough gel composed of atelocollagen, which lacks an immunogenetic site, is a promising material for biomedical application. In this study, we created a composite hydrogel composed of atelocollagen gel cross-linked with glutaraldehyde (GA) and poly-(N,N-dimethylacrylamide) gel exhibiting biocompatibility based on the double-network (DN) gel principle. The tensile toughness of atelocollagen gel remained constant regardless of the amount of cross-linker (GA) used. In contrast, tensile tests of the DN gel indicated that mechanical properties, such as fracture stress and toughness, were significantly higher than those of the atelocollagen gel. Moreover, fibroblast cells adhered and spread on the gels, the Schiff bases of which were treated via reductive amination for detoxification from GA. These findings demonstrate the potential of the proposed gel materials as artificial alternative materials to soft tissues with sub-MPa fracture stress.

Upregulation of Thymidylate Synthase Induces Pemetrexed Resistance in Malignant Pleural Mesothelioma.

Malignant pleural mesothelioma (MPM) is an invasive malignancy that develops in the pleural cavity, and antifolates are used as chemotherapeutics for treating. The majority of antifolates, including pemetrexed (PMX), inhibit enzymes involved in purine and pyrimidine synthesis. MPM patients frequently develop drug resistance in clinical practice, however the associated drug-resistance mechanism is not well understood. This study was aimed to elucidate the mechanism underlying resistance to PMX in MPM cell lines. We found that among the differentially expressed genes associated with drug resistance (determined by RNA sequencing), TYMS expression was higher in the established resistant cell lines than in the parental cell lines. Knocking down TYMS expression significantly reduced drug resistance in the resistant cell lines. Conversely, TYMS overexpression significantly increased drug resistance in the parental cells. Metabolomics analysis revealed that the levels of dTMP were higher in the resistant cell lines than in the parental cell lines; however, resistant cells showed no changes in dTTP levels after PMX treatment. We found that the nucleic acid-biosynthetic pathway is important for predicting the efficacy of PMX in MPM cells. The results of chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR) assays suggested that H3K27 acetylation in the 5'-UTR of TYMS may promote its expression in drug-resistant cells. Our findings indicate that the intracellular levels of dTMP are potential biomarkers for the effective treatment of patients with MPM and suggest the importance of regulatory mechanisms of TYMS expression in the disease.

De novo deoxyribonucleotide biosynthesis regulates cell growth and tumor progression in small-cell lung carcinoma.

Deoxyribonucleotide biosynthesis from ribonucleotides supports the growth of active cancer cells by producing building blocks for DNA. Although ribonucleotide reductase (RNR) is known to catalyze the rate-limiting step of de novo deoxyribonucleotide triphosphate (dNTP) synthesis, the biological function of the RNR large subunit (RRM1) in small-cell lung carcinoma (SCLC) remains unclear. In this study, we established siRNA-transfected SCLC cell lines to investigate the anticancer effect of silencing RRM1 gene expression. We found that RRM1 is required for the full growth of SCLC cells both in vitro and in vivo. In particular, the deletion of RRM1 induced a DNA damage response in SCLC cells and decreased the number of cells with S phase cell cycle arrest. We also elucidated the overall changes in the metabolic profile of SCLC cells caused by RRM1 deletion. Together, our findings reveal a relationship between the deoxyribonucleotide biosynthesis axis and key metabolic changes in SCLC, which may indicate a possible link between tumor growth and the regulation of deoxyribonucleotide metabolism in SCLC.

Effect of postprandial moderate-intensity walking for 15-min on glucose homeostasis in type 2 diabetes mellitus patients.

AIM: Diabetes patients usually have a low activity level and complain about lack of time. Therefore, we investigated the effect of short time, postprandial moderate-intensity exercise on glucose homeostasis in type 2 diabetes patients. METHODS: Eleven patients with type 2 diabetes were recruited. Patients spent the first day of the study without exercise (non-exercise day; NE day). In the second day, they walked at moderate-intensity (40% of the maximum heart rate reserve) for 15 min, 30 min after each meal (exercise day; E day). Glucose homeostasis was estimated by a continuous glucose monitor (CGM). All meals during the study were of standard composition. We compared NE day and E day concerning 24-h glucose homeostasis and 3 h postprandial glucose levels by the incremental area under the curve (iAUC) method. Medications were not changed during the study. RESULTS: The number of patients under basal supported oral therapy, intensive insulin therapy and oral hypoglycemic agents (OHA) were 5, 4 and 2, respectively. The blood glucose standard deviation over 24 h and the iAUC for the 24-h glycemic variability (NE day vs. E day; 34,765 [21,424-56,014] vs. 23,205 [15,323-39,779]) were smaller in E day than in NE day. CONCLUSION: These results suggest that postprandial moderate-intensity walking, easily performable in daily life activities, was effective for improving glucose homeostasis. Further study should be performed to clarify the relationship between postprandial walk and drug therapy (insulin and OHA), including insulin secretory ability.

Extraction of Aqueous Metabolites from Cultured Adherent Cells for Metabolomic Analysis by Capillary Electrophoresis-Mass Spectrometry.

Metabolomic analysis is a promising omics approach to not only understand the specific metabolic regulation in cancer cells compared to normal cells but also to identify biomarkers for early-stage cancer detection and prediction of chemotherapy response in cancer patients. Preparation of uniform samples for metabolomic analysis is a critical issue that remains to be addressed. Here, we present an easy and reliable protocol for extracting aqueous metabolites from cultured adherent cells for metabolomic analysis using capillary electrophoresis-mass spectrometry (CE-MS). Aqueous metabolites from cultured cells are analyzed by culturing and washing cells, treating cells with methanol, extracting metabolites, and removing proteins and macromolecules with spin columns for CE-MS analysis. Representative results using lung cancer cell lines treated with diamide, an oxidative reagent, illustrate the clearly observable metabolic shift of cells under oxidative stress. This article would be especially valuable to students and investigators involved in metabolomics research, who are new to harvesting metabolites from cell lines for analysis by CE-MS.

Metabolic Characterization of Antifolate Responsiveness and Non-responsiveness in Malignant Pleural Mesothelioma Cells.

Antifolates are a class of drugs effective for treating malignant pleural mesothelioma (MPM). The majority of antifolates inhibit enzymes involved in purine and pyrimidine synthesis such as dihydrofolate reductase (DHFR), thymidylate synthase (TYMS), and glycinamide ribonucleotide formyltransferase (GART). In order to select the most suitable patients for effective therapy with drugs targeting specific metabolic pathways, there is a need for better predictive metabolic biomarkers. Antifolates can alter global metabolic pathways in MPM cells, yet the metabolic profile of treated cells has not yet been clearly elucidated. Here we found that MPM cell lines could be categorized into two groups according to their sensitivity or resistance to pemetrexed treatment. We show that pemetrexed susceptibility could be reversed and DNA synthesis rescued in drug-treated cells by the exogenous addition of the nucleotide precursors hypoxanthine and thymidine (HT). We observed that the expression of pemetrexed-targeted enzymes in resistant MPM cells was quantitatively lower than that seen in pemetrexed-sensitive cells. Metabolomic analysis revealed that glycine and choline, which are involved in one-carbon metabolism, were altered after drug treatment in pemetrexed-sensitive but not resistant MPM cells. The addition of HT upregulated the concentration of inosine monophosphate (IMP) in pemetrexed-sensitive MPM cells, indicating that the nucleic acid biosynthesis pathway is important for predicting the efficacy of pemetrexed in MPM cells. Our data provide evidence that may link therapeutic response to the regulation of metabolism, and points to potential biomarkers for informing clinical decisions regarding the most effective therapies for patients with MPM.

Determinants of Glycated Hemoglobin in Subjects With Impaired Glucose Tolerance: Subanalysis of the Japan Diabetes Prevention Program.

BACKGROUND: Limited evidence is available about the relationship of lifestyle factors with glycated hemoglobin (HbA1c) in subjects with impaired glucose tolerance. The aim of study was to identify such determinant factors of HbA1c in subjects with impaired glucose tolerance. METHODS: This cross-sectional study included 121 men and 124 women with impaired glucose tolerance, who were diagnosed based on a 75-g oral glucose tolerance test. Demographic and biochemical parameters, including the body mass index (BMI), fasting plasma glucose (FPG), 2-h post-load glucose (2-h PG), and HbA1c, were measured. The pancreatic ß-cell function and insulin resistance were assessed using homeostasis model assessment (HOMA-ß). Dietary intake was assessed by a food frequency questionnaire. RESULTS: The levels of FPG, 2-h PG, and carbohydrate intake were correlated with the HbA1c level in men, while the FPG and 2-h PG levels were correlated with the HbA1c level in women. In multiple regression analyses, BMI, FPG, 2-h PG, and white rice intake were associated with HbA1c levels in men, while BMI, FPG, HOMA-ß, and bread intake were associated with HbA1c levels in women. CONCLUSIONS: The present findings suggest that a substantial portion of HbA1c may be composed of not only glycemic but also several lifestyle factors in men with impaired glucose tolerance. These factors can be taken into consideration as modifiable determinants in assessing the HbA1c level for the diagnosis and therapeutic monitoring of the disease course.

Effects of lifestyle intervention on weight and metabolic parameters in patients with impaired glucose tolerance related to beta-3 adrenergic receptor gene polymorphism Trp64Arg(C/T): Results from the Japan Diabetes Prevention Program.

The beta-3 adrenergic receptor (ADRB3), primarily expressed in adipose tissue, is involved in the regulation of energy metabolism. The present study hypothesized that ADRB3 (Trp64Arg, rs4994) polymorphisms modulate the effects of lifestyle intervention on weight and metabolic parameters in patients with impaired glucose tolerance. Data were analyzed from 112 patients with impaired glucose tolerance in the Japan Diabetes Prevention Program, a lifestyle intervention trial, randomized to either an intensive lifestyle intervention group or usual care group. Changes in weight and metabolic parameters were measured after the 6-month intervention. The ADRB3 polymorphisms were determined using the polymerase chain reaction restriction fragment length polymorphism method. Non-carriers showed a greater weight reduction compared with the carriers in both the lifestyle intervention group and usual care group, and a greater increase of high-density lipoprotein cholesterol levels than the carriers only in the lifestyle intervention group. ADRB3 polymorphisms could influence the effects of lifestyle interventions on weight and lipid parameters in impaired glucose tolerance patients.

Effect of baseline HbA1c level on the development of diabetes by lifestyle intervention in primary healthcare settings: insights from subanalysis of the Japan Diabetes Prevention Program.

OBJECTIVES: To determine the effects of a lifestyle intervention on the development of type 2 diabetes mellitus (T2DM) among participants with impaired glucose tolerance (IGT), in particular in the subgroup with baseline glycated hemoglobin (HbA1c) levels ≥5.7%, in primary healthcare settings. DESIGN: Randomized controlled trial. SETTING: 32 healthcare centers in Japan. PARTICIPANTS: Participants with IGT, aged 30-60 years, were randomly assigned to either an intensive lifestyle intervention group (ILG) or a usual care group (UCG). INTERVENTIONS: During the initial 6 months, participants in the ILG received four group sessions on healthy lifestyles by public health providers. An individual session was further conducted biannually during the 3 years. Participants in the UCG received usual care such as one group session on healthy lifestyles. OUTCOME MEASURES: The primary endpoint was the development of T2DM based on an oral glucose tolerance test. RESULTS: The mean follow-up period was 2.3 years. The annual incidence of T2DM were 2.7 and 5.1/100 person-years of follow-up in the ILG (n=145) and UCG (n=149), respectively. The cumulative incidence of T2DM was significantly lower in the ILG than in the UCG among participants with HbA1c levels ≥5.7% (log-rank=3.52, p=0.06; Breslow=4.05, p=0.04; Tarone-Ware=3.79, p=0.05), while this was not found among participants with HbA1c levels <5.7%. CONCLUSIONS: Intensive lifestyle intervention in primary healthcare setting is effective in preventing the development of T2DM in IGT participants with HbA1c levels ≥5.7%, relative to those with HbA1c levels <5.7%. TRIAL REGISTRATION NUMBER: UMIN000003136.

Current situation of clinical trials in Beijing and Shanghai, China.

OBJECTIVE: Global clinical trials are important because they facilitate rapid delivery of new and effective drugs to patients Assessment of the current situation of clinical trials conducted in Asia is critical for improving performance of global clinical trials. However, review reports from China or other Asian countries are not yet available. Therefore, the purpose of this research was to investigate the current quality of clinical trials conducted in Shanghai, as well as Beijing. METHODS: Questionnaires were distributed to medical doctors attending institutes in Beijing and Shanghai in which clinical trials have been conducted. These questionnaires were delivered and collected from both areas by the Peking University research team of Beijing and the Fudan University research team of Shanghai respectively. Analysis and evaluation were conducted by research teams from both China and Japan. RESULTS: Subjects were randomly selected by the respective research team. A total of 145 questionnaires in Beijing and 162 in Shanghai were administered: all 307 questionnaires were completed. In total, 57.2% and 74.5% of respondents from Beijing and Shanghai, respectively, reported participation in audits and inspections on an annual basis conducted by their own institute. A total of 49.2% and 56.0% of respondents from Beijing and Shanghai, respectively, reported that they received reports after the audits and inspections by an institute. 23.5% and 37.7% of respondents from Beijing and Shanghai, respectively, reported participation in audits conducted annually by external authorities. A total of 18.9% and 29.5% of respondents from Beijing and Shanghai, respectively, reported that they received reports after the audits and inspections by an external authority. CONCLUSIONS: Our research suggests that clinical trials in Shanghai, as well as in Beijing, are conducted vigorously and appropriately monitored by audits and inspections conducted by concerned institutes and/or by an external authority.

Effects of physical exercise prescribed by a medical support team on elderly lower extremity osteoarthritis combined with metabolic syndrome and/or type 2 diabetes.

AIM: The aims of the present study were to investigate the effectiveness of exercise intervention provided by a medical support team specializing in lifestyle-related diseases in the treatment of elderly lower extremity osteoarthritis and to examine the influence of bodyweight decrease on changes in the evaluation indexes. METHODS: Participants were 61 patients (57 women and 4 men, aged 68.3 ± 9.6 years) with lower extremity osteoarthritis (109 total diseased joints) and either one or more of the following diseases: obesity, metabolic syndrome and type 2 diabetes. Indexes relating to metabolic diseases, orthopedic disorders, lifestyle activity level and health-related quality of life (HRQOL) were obtained before and after exercise intervention. RESULTS: The numbers of patients with obesity, metabolic syndrome, type 2 diabetes, gonarthrosis and coxarthrosis were 56, 49, 32, 56 and 9, respectively. The mean intervention period was 4.7 ± 1.6 months (2-10.8 months). Indexes relating to metabolic diseases and orthopedic disorders, activity level and HRQOL were all significantly improved after intervention (P < 0.05). Bodyweight decreased by 10.3% and showed a correlation with other evaluated items. Five explanatory variables were extracted through multiple regression analysis that bodyweight reduction rate was set as the criterion variable: changes of bodyweight, body mass index, percent body fat, glycated hemoglobin and total energy expenditure per bodyweight. CONCLUSION: The exercise intervention provided by our medical support team clearly improved indexes relating to metabolic diseases and orthopedic disorders. In addition, decreased bodyweight was related to improvements in metabolic factors, motor function and HRQOL.

Cancer mortality among atomic bomb survivors exposed as children.

OBJECTIVES: To compare cancer mortality among A-bomb survivors exposed as children with cancer mortality among an unexposed control group (the entire population of Japan, JPCG). METHODS: The subjects were the Hiroshima and Nagasaki A-bomb survivor groups (0-14 years of age in 1945) reported in life span study report 12 (follow-up years were from 1950 to 1990), and a control group consisting of the JPCG. We estimated the expected number of deaths due to all causes and cancers of various causes among the exposed survivors who died in the follow-up interval, if they had died with the same mortality as the JPCG (0-14 years of age in 1945). We calculated the standardized mortality ratio (SMR) of A-bomb survivors in comparison with the JPCG. RESULTS: SMRs were significantly higher in exposed boys overall for all deaths, all cancers, leukemia, and liver cancer, and for exposed girls overall for all cancers, solid cancers, liver cancer, and breast cancer. In boys, SMRs were significantly higher for all deaths and liver cancer even in those exposed to very low doses, and for all cancers, solid cancers, and liver cancer in those exposed to low doses. In girls, SMRs were significantly higher for liver cancer and uterine cancer in those exposed to low doses, and for leukemia, solid cancers, stomach cancer, and breast cancer in those exposed to high doses. CONCLUSIONS: We calculated the SMRs for the A-bomb survivors versus JPCG in childhood and compared them with a true non-exposed group. A notable result was that SMRs in boys exposed to low doses were significantly higher for solid cancer.

Association between improvements in insulin resistance and changes in cognitive function in elderly diabetic patients with normal cognitive function.

AIM: The number of elderly with cognitive dysfunction has been increasing in developed countries. Several studies have shown that diabetes is a risk factor for declines in cognitive function and, recently, numerous studies have demonstrated that exercise improves insulin resistance (IR). However, no studies have been undertaken to examine the relationship between IR and cognitive dysfunction. METHODS: Sixteen elderly diabetic patients participated in this study (mean age 70.9 ± 3.7 years). They were divided into control and training groups, and the training group was instructed to exercise 4 days per week for 12 weeks using horse riding simulation equipment (JOBA). Insulin sensitivity was measured by the euglycemic clamp technique and several tests to assess cognitive function, such as the Mini-Mental State Examination (MMSE), were performed. RESULTS: There were no statistically significant differences between the control and training groups in the baseline data. The differences of glucose infusion rate values and delayed word list scores, between baseline and the follow up, were significantly correlated (P = 0.024). In addition, changes in fasting blood sugar (FBS) and in the Trail Making Test B (TMT-B) score were also correlated (P = 0.035). CONCLUSION: The current study showed that a 12-week exercise intervention program did not significantly improve IR in elderly diabetic patients. However, changes in IR were associated with improvements in memory function, and reduced FBS was associated with improvements in TMT-B.

Prevention of type 2 diabetes in a primary healthcare setting: three-year results of lifestyle intervention in Japanese subjects with impaired glucose tolerance.

BACKGROUND: A randomized control trial was performed to test whether a lifestyle intervention program, carried out in a primary healthcare setting using existing resources, can reduce the incidence of type 2 diabetes in Japanese with impaired glucose tolerance (IGT). The results of 3 years' intervention are summarized. METHODS: Through health checkups in communities and workplaces, 304 middle-aged IGT subjects with a mean body mass index (BMI) of 24.5 kg/m2 were recruited and randomized to the intervention group or control group. The lifestyle intervention was carried out for 3 years by public health nurses using the curriculum and educational materials provided by the study group. RESULTS: After 1 year, the intervention had significantly improved body weight (-1.5 ± 0.7 vs. -0.7 ± 2.5 kg in the control; p = 0.023) and daily non-exercise leisure time energy expenditure (25 ± 113 vs. -3 ± 98 kcal; p = 0.045). Insulin sensitivity assessed by the Matsuda index was improved by the intervention during the 3 years. The 3-year cumulative incidence tended to be lower in the intervention group (14.8% vs.8.2%, log-rank test: p = 0.097). In a sub-analysis for the subjects with a BMI > 22.5 kg/m2, a significant reduction in the cumulative incidence was found (p = 0.027). CONCLUSIONS: The present lifestyle intervention program using existing healthcare resources is beneficial in preventing diabetes in Japanese with IGT. This has important implications for primary healthcare-based diabetes prevention. TRIAL REGISTRATION NUMBER: UMIN000003136.

Effect of branched-chain amino acid supplementation during unloading on regulatory components of protein synthesis in atrophied soleus muscles.

Maintenance of skeletal muscle mass depends on the equilibrium between protein synthesis and protein breakdown; diminished functional demand during unloading breaks this balance and leads to muscle atrophy. The current study analyzed time-course alterations in regulatory genes and proteins in the unloaded soleus muscle and the effects of branched-chain amino acid (BCAA) supplementation on muscle atrophy and abundance of molecules that regulate protein turnover. Short-term (6 days) hindlimb suspension of rats resulted in significant losses of myofibrillar proteins, total RNA, and rRNAs and pronounced atrophy of the soleus muscle. Muscle disuse induced upregulation and increases in the abundance of the eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1), increases in gene and protein amounts of two ubiquitin ligases (muscle RING-finger protein 1 and muscle atrophy F-box protein), and decreases in the expression of cyclin D1, the ribosomal protein S6 kinase 1, the mammalian target of rapamycin (mTOR), and ERK1/2. BCAA addition to the diet did not prevent muscle atrophy and had no apparent effect on regulators of proteasomal protein degradation. However, BCAA supplementation reduced the loss of myofibrillar proteins and RNA, attenuated the increases in 4E-BP1, and partially preserved cyclin D1, mTOR and ERK1 proteins. These results indicate that BCAA supplementation alone does not oppose protein degradation but partly preserves specific signal transduction proteins that act as regulators of protein synthesis and cell growth in the non-weight-bearing soleus muscle.

Effects of daily mechanical horseback riding on insulin sensitivity and resting metabolism in middle-aged type 2 diabetes mellitus patients.

The present study was conducted to investigate the effect of daily passive exercise using a horseback riding machine (Joba) on insulin sensitivity and resting metabolism in middle-aged, diabetic patients. Participants were 24 type 2 diabetes mellitus patients aged 59 +/- 8 years (mean +/- SD; range from 43 to 75 years of age). Patients were randomly divided into control (normal lifestyle) and Joba exercise groups. The latter group was instructed to perform one 30-min session of Joba riding per day, 7 times per week, for 3 months. Compared with baseline values, serum immunoreactive insulin (IRI) concentrations decreased and HOMA-IR was improved by Joba training. In addition, exercise duration per day significantly correlated (r = -0.65) with changes in serum IRI, and 3-month mechanical horseback riding significantly increased the resting metabolic rate of the patients. These results suggest that daily Joba passive exercise is potentially useful as a means to improve insulin sensitivity and resting metabolism in diabetic patients. The Joba fitness equipment can prove especially useful as an alternative exercise therapy for aged individuals incapable of performing independent exercise or for those who suffer from knee-joint disorders.

Goshajinkigan (Chinese herbal medicine niu-che-sen-qi-wan) improves insulin resistance in diabetic rats via the nitric oxide pathway.

Goshajinkigan (GJG), an aqueous extract of a combination of 10 herbal medicines, is widely used for the treatment of diabetic neuropathy in Japan. In this study, the effect of GJG on insulin-induced glucose disposal in normal and streptozotocin (STZ) diabetic rats was analyzed using the euglycemic clamp technique. Male Wistar rats, aged 9 weeks, were randomly assigned to six groups: group NS, normal rats receiving saline; group NG, normal rats receiving GJG (800 mg x kg(-1) x day(-1), p.o.); group NGL, normal rats receiving GJG + N(G)-monomethyl-L-arginine (L-NMMA, 1 mg x kg(-1) x min(-1), i.v.); group DS, diabetic rats receiving saline; group DG, diabetic rats receiving GJG; group DGL, diabetic rats receiving GJG + L-NMMA. After daily oral administrations of saline or GJG for one week, euglycemic clamp experiments were performed. The metabolic clearance rates of glucose (MCR) in the DS, DG, and DGL groups (8.7 +/- 2.9, 18.2 +/- 2.5, and 8.1 +/- 1.8 ml x kg(-1) x min(-1), respectively) were significantly lower than those in the NS, NG, and NGL groups (24.1 +/- 4.5, 24.5 +/- 3.1, and 22.2 +/- 2.1 ml x kg(-1) x min(-1), respectively). In addition, the MCR in the DG group was significantly higher than that in the DS and DGL groups, while no significant difference was detected among the NS, NG, and NGL groups. Furthermore, the amelioration of insulin resistance by GJG in diabetic rats was hampered by L-NMMA infusion. These results suggest that daily GJG administrations ameliorate insulin resistance in STZ-diabetic rats, and that the nitric oxide pathway may mediate the effect of GJG.

Decreased enzyme activity and contents of hepatic branched-chain alpha-keto acid dehydrogenase complex subunits in a rat model for type 2 diabetes mellitus.

The mitochondrial branched-chain alpha-keto acid dehydrogenase complex (BCKDC) is responsible for the committed step in branched-chain amino acid catabolism. In the present study, we examined BCKDC regulation in Otsuka Long-Evans Tokushima Fatty (OLETF) rats both before (8 weeks of age) and after (25 weeks of age) the onset of type 2 diabetes mellitus. Long-Evans Tokushima Otsuka (LETO) rats were used as controls. Plasma branched-chain amino acid and branched-chain alpha-keto acid concentrations were significantly increased in young and middle-aged OLETF rats. Although the hepatic complex was nearly 100% active in all animals, total BCKDC activity and protein abundance of E1alpha, E1beta, and E2 subunits were markedly lower in OLETF than in LETO rats at 8 and 25 weeks of age. In addition, hepatic BCKDC activity and protein amounts were significantly decreased in LETO rats aged 25 weeks than in LETO rats aged 8 weeks. In skeletal muscle, E1beta and E2 proteins were significantly reduced, whereas E1alpha tended to increase in OLETF rats. Taken together, these results suggest that (1) whole-body branched-chain alpha-keto acid oxidation capacity is extremely reduced in OLETF rats independently of diabetes development, (2) the aging process decreases BCKDC activity and protein abundance in the liver of normal rats, and (3) differential posttranscriptional regulation for the subunits of BCKDC may exist in skeletal muscle.