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The fruit of Prunus mume (ume, also known as Japanese apricot) has been used as a functional food in Japan since ancient times. We previously reported that ume stimulates the differentiation of preosteoblastic cells. Osteocalcin (OCN) is secreted by osteoblasts, and there is known association with glucolipid metabolism and cognitive function. This study sought to clarify the relationship between ume extracts and OCN production both in vitro and in vivo. Alkaline phosphatase activity and OCN level in the ethyl acetate extracts of ume-treated extracts were significantly increased in preosteoblast MC3T3-E1 cells compared with the control group. In human study, serum OCN level was significantly higher in the high ume intake group than in the low intake group in community-dwelling participants over 60 years old. These results suggest that ume has the potential to upregulated OCN production both in vitro and in vivo.

Dietary factors are thought to play an important role in the prevention of cognition diseases and depression in late life. In the present study, we compared the effects between the theogallin-rich tea cultivar, "MK5601" and a common Japanese tea cultivar, "Yabukita" on behaviors and hippocampal neurotrophin levels in experimental animals. Middle-aged mice (aged 8 mo) were given either of the tea infusions or water ad libitum for 4 mo. In the novel object location test, the middle-aged mice drinking water or "Yabukita" performed worse than young mice (aged 2-3 mo) although the middle-aged mice drinking "MK5601" retained spatial memory at the same level as the young mice. We also found that the middle-aged mice drinking "MK5601" showed high levels of neurotrophin-3 in the hippocampus. In conclusion, the "MK5601" tea infusion appears to be effective in preventing age-related changes in cognitive function, as compared with a common Japanese tea cultivar.

研究グループは、大阪大学大学院医学研究科臨床遺伝子治療学と共同で、血液中で検出されるアミロイドβ(Aβ)はグルコースやインスリンに感受性のある末梢組織(膵臓、脂肪組織、骨格筋、肝臓など)から分泌されていることを明らかにしました。また、末梢組織で分泌されたAβは膵臓のβ細胞に作用してインスリン分泌を抑える調節因子として働いていることも明らかにしました。本研究結果は、血液Aβレベルが食事によって大きく変動することを示しており、アルツハイマー病(AD)の診断マーカーとして使う際には、空腹時に採血するなど特別の注意が必要であることを示しています。ADはAβが脳内に蓄積することが原因で起こります。Aβは主に脳で産生されますが、血液中でも検出され、脳内のアミロイド病理を反映しているのではないかとの考えから、ADマーカーとしての研究が進められています。しかし、Aβの前駆体APPやAβ産生酵素は脳に限らず多くの末梢組織で発現しており、血液中のAβの由来については解明されていませんでした。また、ヒトやマウスにグルコースやインスリンを投与すると、血液中のAβが一過性に上昇することが報告されていますが、その理由も明らかではありませんでした。そこで本研究グループは、マウスを使用して末梢Aβの産生組織と末梢での生理作用を観察しました。その結果、末梢のAβはグルコース刺激により膵臓のβ細胞からインスリンとともに分泌され、インスリン刺激により脂肪組織、骨格筋、肝臓からそれぞれのオルガノカインとともに分泌されることが明らかになりました。また、分泌されたAβは膵臓のβ細胞に作用し、インスリンの分泌を抑制することにより、血糖の調節に寄与していることも明らかになりました。, Type 2 diabetes mellitus is known to be a risk factor for Alzheimer's disease(AD), but the underlying mechanisms remain unclear. In AD, the cerebral accumulation of amyloid β(Aβ) triggers a pathological cascade leading to neurodegeneration. Plasma Aβ levels are thought to reflect the brain amyloid pathology and currently used as a diagnostic biomarker of AD. However, ...

Zanthoxylum piperitum (ZP, 'Japanese pepper') is a traditional medicine and pepper used in Asian countries such as Japan. Hydroxy-α-sanshool, a pungent-tasting substance contained within ZP, has been reported to slightly suppress immunoglobulin E (IgE)-mediated mast cell degranulation. The current study aims to newly identify anti-allergic compounds derived from ZP. We examine the inhibitory mechanisms behind IgE-mediated mast cell degranulation. By inhibitory effect-guided isolation, we identified degranulation inhibitory compounds derived from ZP fruit: 1-acetoxy-7-hydroxy-3, 7-dimethylocta-2E, 5E-diene (ZP1) and 8-hydroxygeranyl acetate (ZP2). ZP1 and ZP2 inhibited IgE-mediated degranulation and A23187-mediated degranulation in RBL-2H3 mast cells. Our findings suggest the inhibition of degranulation by ZP1 and ZP2 was by inhibition of Lyn phosphorylation, followed by inhibition of intracellular Ca2+ mobilization, protein kinase C alpha phosphorylation, membrane ruffling, and granule-to-plasma membrane fusion. Oral administration of ZP1 or ZP2 attenuated an IgE-mediated passive cutaneous anaphylactic reaction in mice. Histological observation suggests that this effect occurred via inhibition of mast cell degranulation. These findings indicate that ZP1 and ZP2 attenuate allergic reaction via inhibition of IgE-mediated mast cell degranulation.

Matcha has high contents of theanine, caffeine, and epigallocatechin gallate (EGCG). Among these, theanine reduces psychological and physiological stress responses, although this effect is disturbed by caffeine and EGCG. It was reported that the continued ingestion of high-quality matcha, whose molar ratio of caffeine and EGCG to theanine and arginine was kept at less than two, reduces stress responses. However, most matcha on the market has an inadequate ratio. Therefore, we investigated the influence of continued ingestion of matcha with a ratio of over two, on emotional behaviors after stress. Continued ingestion of matcha was suggested to reduce the anxiety-like behaviors induced by psychological and physiological stresses.

Osteoporosis is a complication of diabetes mellitus (DM). The pathology of diabetic osteoporosis is distinct from postmenopausal osteoporosis, and there are no specific treatment guidelines for diabetic osteoporosis. In the current study, this issue was addressed by evaluating the effect of osteoporosis medications, such as the anabolic agent PTH [teriparatide (TPTD)] and the antiresorptive agents calcitonin [elcatonin (ECT)] and bisphosphonate [risedronate (RIS)], on bone metabolism as well as on glucose and lipid metabolism in spontaneously diabetic Torii (SDT) fatty rats, which are a model of type 2 DM (T2DM). The medicines were injected subcutaneously into 8-week-old male SDT fatty rats three times weekly for 8 weeks. TPTD treatment in SDT fatty rats increased the osteoblast number and function on trabecular bone in vertebrae, and increased the trabecular bone mass, bone mineral density (BMD), and mechanical strength of vertebrae. Additionally, TPTD improved cortical bone structure and increased BMD. RIS decreased the osteoclast number and function, which led to an increase in vertebral bone mineral content and BMD in the femoral diaphysis, and mechanical strength was increased in the vertebrae. ECT showed no clear effects on bone mass or metabolism. Similar to diabetic lesions, all of the drugs had no effects on hyperglycemia, pancreas morphology, or serum insulin and glucagon levels. However, triglyceride levels and lipid droplets in fatty liver were decreased in the TPTD group. These results suggest that TPTD may be useful for treating fatty liver in addition to osteoporosis in T2DM.

Locomotive syndrome (LS) is a concept that refers to the condition of people requiring healthcare services because of problems associated with locomotion. Depression is a major psychiatric disease among the elderly, in addition to dementia. The purpose of this study was to determine the association between LS and depression. The study participants were 224 healthy elderly volunteers living in a rural area in Japan. LS was defined as scores ≥ 16 on the 25-question Geriatric Locomotive Function Scale (GLFS-25). Depression was defined as scores ≥ 5 on the 15-item Geriatric Depression Scale (GDS-15). Height and body weight were measured. The prevalence of LS and depression was 13.9% and 24.2%, respectively. Compared with the non-LS group, the LS group was older, was shorter, had a higher BMI, and had higher GDS-15 scores. Logistic regression analysis showed that participants with GDS-15 scores ≥ 6 had higher odds for LS than those with GDS-15 scores < 6 (odds ratio [OR] = 4.22). Conversely, the depression group had higher GLFS-25 scores than the nondepression group. Participants with GLFS-25 scores ≥ 5 had higher odds for depression than those with GLFS-25 scores < 5 (OR = 4.53). These findings suggest that there is a close relationship between LS and depression.

Calcitonin (CT) plays an important role in calcium homeostasis, and its precursor, proCT, is positively associated with the body mass index in the general human population. However, the physiological role of endogenous CT in the regulation of metabolism remains unclear. Knockout mice with gene-targeted deletion of exon 4 of Calca (CT KO) were generated by targeted modification in embryonic stem cells. Male mice were used in all experiments and were fed a slightly higher fat diet than the standard diet. The CT KO mice did not exhibit any abnormal findings in appearance, but exhibited weight loss from 15 months old, i.e., significantly decreased liver, adipose tissue, and kidney weights, compared with wild-type control mice. Furthermore, CT KO mice exhibited significantly decreased fat contents in the liver, lipid droplets in adipose tissues, serum glucose, and lipid levels, and significantly increased insulin sensitivity and serum adiponectin levels. CT significantly promoted 3T3-L1 adipocyte differentiation and suppressed adiponectin release. These results suggested that CT gene deletion prevents obesity, hyperglycemia, and hyperlipidemia in aged male mice. This is the first definitive evidence that CT may contribute to glucose and lipid metabolism in aged male mice, possibly via decreased adiponectin secretion from adipocytes.

The green tea (Camellia sinensis L.) cultivar “Sunrouge” contains anthocyanins, catechins and flavonols. To determine whether ingesting green tea containing anthocyanins improves visual function and blood pressure (BP) in healthy adults, a randomized, double-blind, placebo-controlled study was performed. A total of 120 healthy subjects, aged between 20 and 60 years and with a systolic BP (SBP) value of ≤125 and

To examine the possible benefits of tea flavonols, we compared anti-atherogenic effects between common and flavonol-rich tea cultivars. The tea infusion made from a flavonol-rich cultivar, but not a common cultivar, significantly decreased the plasma oxidized low-density lipoprotein level in mice fed a high-cholesterol diet. The result suggests that tea flavonols have the potential to protect against cardiovascular diseases.

Granulosa cells form ovarian follicles and play important roles in the growth and maturation of oocytes. The protection of granulosa cells from cellular injury caused by oxidative stress is an effective therapy for female infertility. We here investigated an effective bioactive compound derived from Prunus mume seed extract that protects granulosa cells from hydrogen peroxide (H2O2)-induced apoptosis. We detected the bioactive compound, 3,4-dihydroxybenzaldehyde (3,4-DHBA), via bioactivity-guided isolation and found that it inhibited the H2O2-induced apoptosis of granulosa cells. We also showed that 3,4-DHBA promoted estradiol secretion in granulosa cells and enhanced the mRNA expression levels of steroidogenic factor 1, a promoter of key steroidogenic enzymes. These results suggest that P. mume seed extract may have clinical potential for the prevention and treatment of female infertility.

It has been shown that amyloid β (Aβ) secretion regulates cholesterol efflux from cells and that the E693Δ (Osaka) mutation in amyloid precursor protein (APP) promotes intracellular accumulation of Aβ and thus reduces its secretion. These findings led us to speculate that APP with the Osaka mutation (APPOSK ) might have a defect in cholesterol efflux and thus cause cellular malfunction. We therefore examined the effects of this mutation on intracellular cholesterol transport and efflux in cultured cells. Upon cholesterol loading, APPOSK -expressing cells exhibited higher levels of cellular cholesterol than wild-type APP-expressing cells, suggesting impaired cholesterol efflux. It is known that, after its internalization, cholesterol is transported from the endosomes to the endoplasmic reticulum (ER) and Golgi apparatus and then to the plasma membrane. In APPOSK -expressing cells, cholesterol accumulated with Aβ in the ER and Golgi apparatus and alone in endosomes/lysosomes. These results imply that the mutation-induced disturbance of Aβ trafficking from the ER to the plasma membrane affects cholesterol transport to cause cholesterol accumulation in the ER and Golgi apparatus and, consequently, in endosomes. Furthermore, we detected an enhanced mitochondrial accumulation of Aβ and cholesterol in APPOSK -expressing cells, and this was accompanied by an increase in the generation of reactive oxygen species (ROS). The present findings suggest that Aβ trafficking is important for intracellular cholesterol transport and efflux and that the Osaka mutation potentiates cholesterol-dependent exacerbation of intracellular Aβ toxicity, i.e. Aβ-induced ROS generation, by disturbing Aβ-mediated cholesterol efflux from the cell.

We performed proteomics analysis of the cerebrospinal fluid (CSF) of healthy dogs and dogs with meningoencephalitis of unknown etiology (MUE). By comparing two-dimensional electrophoreses (2DE), an upregulated spot was found in MUE dogs. This protein was identified as a neuron-specific enolase (NSE) by analysis with MALDI-TOF mass spectrometry. In comparing dot blots using an antibody against NSE, the NSE levels in the CSF of MUE dogs was significantly higher than that of the controls. NSE is a diagnostic marker of neuroendocrine tumors, brain injury and spinal cord trauma in humans. It seems that the NSE concentration in the CSF is increased by cellular destruction in canine encephalitis. Though elevation of NSE may not be specific in canine encephalitis because the NSE level was increased in other CNS diseases, further study including measurement with serum is necessary.

The molecular mechanism of severe adverse effects of the endocrine disruptor diethylstilbestrol (DES) on reproductive organs is not currently understood. The effects of DES on testicular proteins were studied in adult male rats orally treated with 0.35 and 3.5 mg DES/kg every two days for two weeks before the manifestation of morphological toxicities. Two up-regulated proteins (glutamine synthetase and chaperonin containing TCP1), two down-regulated proteins (thioredoxin-like 1 and testis-specific autoantigen) and two proteins with altered isoelectric points (protein disulfide isomerase [PDI a3] and enolase 1) were identified in DES groups. Carbonylation of PDI a3 was detected. A significant decrease in PDI activity and significant increases in caspase-12 and calpain activities were also found in the group. It is suggested that testicular toxicity by DES was initiated by the down-regulation of thioredoxin-like-1 leading to the cellular redox inbalances, and the resultant oxidative modification of several important proteins involved in protein foldings.

ATP-binding cassette transporter isoform C2 (ABCC2) localizes to the apical plasma membrane in polarized cells. Apical localization of ABCC2 in hepatocytes plays an important role in biliary excretion of endobiotics and xenobiotics, but the mechanism by which ABCC2 localizes to the apical membrane has not been conclusively elucidated. Here, we investigate the role of scaffolding proteins on ABCC2 localization with a focus on the function of PDZK1 (post-synaptic density 95/disk large/zonula occludens-1 domain containing 1) in regulating ABCC2 localization. The C-terminal 77 residues of ABCC2 were used to probe interacting proteins from HepG2 cells. Protein mass fingerprinting identified PDZK1 as a major interacting protein. PDZK1 associated with the plasma membrane, most likely at the apical vacuoles of HepG2 cells. Affinity pull-down assays confirmed that the C-terminal NSTKF of ABCC2 bound to the fourth PDZ domain of PDZK1. Removal of this PDZ-binding motif significantly reduced the normal apical localization of ABCC2. In HepG2 cells, overexpression of this fourth domain overcame endogenous PDZK1 and reduced the ABCC2 localization at the apical membrane with a reciprocal increase of intracellular accumulation of mislocalized ABCC2. These results suggest a possible role for an interaction between ABCC2 and PDZK1 in apical localization of ABCC2 in hepatocytes.

We examined the effects of a 9-week exercise training (TR) in Wistar male rats, beginning at 4 weeks of age, on the density of endothelial cells (ECs) in epididymal white adipose tissue (WAT) and the mRNA expression of angiogenic factors in adipose tissue stromal vascular fraction (SVF) cells. The number of ECs and mRNA expressions were assessed by lectin staining and real-time reverse transcriptase-polymerase chain reaction, respectively. Compared with control (CR) rats, TR rats gained weight more slowly and had significantly lower final weight of WAT due to the reduction in the size and the number of adipocytes. TR significantly increased the number of ECs per square millimeter and per adipocyte (1.37- and 1.23-fold, respectively) in WAT. This is probably because the number of adipocytes is fewer while the number of ECs is constant in the WAT of TR rats, because the regression line of TR rats for adipocyte number-dependent EC number was shifted toward the left without significant differences in the slopes between groups. TR also induced the upregulation of mRNA expression of vascular endothelial growth factor (Vegf)-A and Vegf-receptor-2 in SVF cells, thereby retaining a constant number of ECs in the WAT.

The green tea (Camellia sinensis L.) cultivar “Sunrouge” contains anthocyanins, catechins and flavonols. To determine whether ingesting green tea containing anthocyanins improves visual function and blood pressure (BP) in healthy adults, a randomized, double-blind, placebo-controlled study was performed. A total of 120 healthy subjects, aged between 20 and 60 years and with a systolic BP (SBP) value of ≤125 and

Aims Our aim was to evaluate the effect of exercise training (TR) on adipocyte-size-dependent expression of leptin and adiponectin. Main methods Male Wistar rats were divided into 2 groups, sedentary control (CR) and TR group, and both monitored for 9 weeks. Adipocytes isolated from epididymal, retroperitoneal, and inguinal fat depots were independently separated into 3 fractions of different cell size, and the relationships between adipocyte size and either leptin or adiponectin mRNA were determined by real-time RT-PCR analysis. Key findings In epididymal and inguinal adipose tissue, positive relationships between adipocyte size and both leptin and adiponectin mRNA expression were found. Comparison of TR and CR rats showed no significant effect of TR on the slopes of the linear regression lines of correlation between leptin mRNA and adipocyte size in either adipose tissue, whereas the slopes of the regression line of correlation between adipocyte size and adiponectin mRNA were greater in TR group. Leptin levels per milliliter of plasma were significantly lower in TR than CR rats, whereas leptin levels adjusted to the 3 fat depots did not differ. TR did not affect adiponectin levels in plasma, whereas adiponectin levels adjusted to the 3 fat depots were significantly greater in TR than CR group. Significance TR-induced reduction in leptin mRNA expression was closely associated with smaller adipocyte size. However, TR amplified the adipocyte-size-dependent expression of adiponectin mRNA, suggesting that TR-induced alterations in adiponectin mRNA may also be mediated by factor(s) other than adipocyte size.

Administration of the beta(2)-agonist clenbuterol has been shown to reduce the expression of beta(2)-adrenoceptor (AR) mRNA in fast-twitch fiber-rich (extensor digitorum longus, EDL) muscle without changing that in slow-twitch fiber-rich (soleus, SOL) muscle in rats. However, the regulatory mechanism for muscle fiber type-dependent down-regulation of the expression of beta(2)-AR mRNA induced by clenbuterol is still unclear. Therefore, mRNA expression of transcriptional and post-transcriptional regulatory factors for beta(2)-AR mRNA levels in fast-twitch fiber-rich (EDL and plantaris, PLA) and slow-twitch fiber-rich (SOL) muscles in clenbuterol-administered (1.0 mg/kg body weight/day for 10 days, subcutaneous) rats was studied by real-time reverse transcription-polymerase chain reaction. Administration of clenbuterol significantly reduced expression of beta(2)-AR mRNA in EDL and PLA muscles without changing that in SOL muscle. Administration of clenbuterol also significantly reduced the mRNA expression of transcriptional regulatory factor (glucocorticoid receptor) and mRNA stabilizing factor (Hu antigen R) in EDL and PLA muscles without changing those in SOL muscle. These results suggest that muscle fiber type-dependent effects of clenbuterol on expression of beta(2)-AR mRNA are closely related to the down-regulation of mRNA expression of transcriptional and post-transcriptional regulatory factors for beta(2)-AR mRNA levels.

Glucocorticoids are known to increase the density and mRNA levels of beta-adrenoceptors (beta-AR) via the glucocorticoid receptor (GR) in many tissues. However, the effects of these changes in the skeletal and cardiac muscles remain relatively unknown. We have investigated the effects of dexamethasone on the expression of the beta(1)-, beta(2)-, and beta(3)-AR mRNAs and GR mRNA in fast-twitch fiber-rich extensor digitorum longus (EDL), slow-twitch fiber-rich soleus (SOL), and left ventricle (LV) muscles by real-time quantitative RT-PCR. Male rats were divided into a dexamethasone group and control group. The weight, RNA concentration, and total RNA content of EDL muscle were 0.76-, 0.85-, and 0.65-fold lower, respectively, in the dexamethasone group than in the control group. The weight, RNA concentration, and total RNA content of SOL muscle were 0.92-, 0.87-, and 0.81-fold lower, respectively, in the dexamethasone group than in the control group; these differences were significant. However, the weight/body weight and total RNA content/body weight of LV muscle were 1.38- and 1.39-fold higher, respectively, in the dexamethasone group than in the control group, respectively; these differences were also significant. Dexamethasone significantly decreased GR mRNA expression in EDL muscle without changing the expression of the beta(1)-, beta(2)-, and beta(3)-AR mRNAs. However, dexamethasone significantly decreased the expressions of beta(2)-AR and GR mRNAs in SOL muscle and significantly increased beta(1)-AR mRNA expression in LV muscle-without changing GR mRNA expression. These results suggest that the effects of dexamethasone on the expression of beta(1)- and beta(2)-AR mRNAs and muscle mass depend on the muscle contractile and/or constructive types.

The acute effects of dihydrocapsaicin (DHC) and capsaicin (CAP) on the number of white blood cells (WBCs), neutrophils, eosinophils, basophils, monocytes, lymphocytes, T lymphocytes, B lymphocytes and NK cells, and serum corticosterone levels were studied in rats. Male 7-wk-old SD rats were divided into DHC (3.0 mg/kg BW), CAP (3.0 mg/kg BW) and control (CON) groups. The number of total WBCs was 1.30-1.42 times significantly higher in the DHC group than in the CON group at 6-12 h. The number of neutrophils was 1.62 times significantly higher in the DHC group than in the CON group at 12 h. The number of total WBCs and neutrophils, however, showed no significant changes between the CAP and CON groups. The number of lymphocytes was 0.61 and 0.70 times significantly lower in the DHC and CAP groups than in the CON group at 3 h. The number of T lymphocytes and B lymphocytes was 0.74 and 0.54 times lower in the DHC group than in the CON group, respectively. CAP, however, did not significantly change the number of T lymphocytes or B lymphocytes. No significant changes in the number of NK cells were observed among the three groups. CAP and DHC did not change the number of monocytes, eosinophils or basophils. No significant changes of the serum corticosterone levels were observed among the three groups. In conclusion, capsaicinoids decreased the number of acquired immunity cells, and increased the number of total WBCs and neutrophils without changing the number of monocytes, eosinophils or basophils. The magnitude of these effects was relatively higher in DHC than in CAP.

The aim of the present study was to determine whether the antiobesity effects of tea catechins (TCs) are associated with the expression of uncoupling protein 1 (UCP1) in brown adipose tissue (BAT). Male Sprague-Dawley rats were fed a high-fat (HF; 35% fat) diet for 5 weeks, then divided into four groups and fed an HF, HF with 0.5% TC (HFTC), normal-fat (NF; 5% fat) or NF with 0.5% TC (NFTC) diet for 8 weeks. At the end of the experimental period, perirenal and epididymal white adipose tissues (WATs) and interscapular BAT were isolated. The NFTC group had significantly lower perirenal WAT weights than the NF group (NF: 12.7+/-0.53 g; NFTC: 10.2+/-0.43 g; P

Background and aim. Helicobacter pylori is known to be a major pathogenic factor in the development of gastritis, peptic ulcer disease and gastric cancer. Recently, chicken egg yolk immunoglobulin Y (IgY) has been recognized as an inexpensive antibody source for passive immunization against gastrointestinal infections. The present study was designed to investigate the effect of anti-urease IgY on H. pylori infection in Mongolian gerbils. Methods. H. pylori-infected Mongolian gerbils were administered a diet containing anti-urease IgY, with or without famotidine (F). After 10 weeks, bacterial culture and measurement of the gastric mucosal myeloperoxidase (MPO) activity were performed. In a second experiment, another group of gerbils was started on a diet containing F + IgY a week prior to H. pylori inoculation. After 9 weeks, these animals were examined. Results. In the H. pylori-infected gerbils, there were no significant differences in the level of H. pylori colonization among the different dietary and control groups. However, the MPO activity was significantly decreased in the H. pylori group administered the F + IgY diet compared with that in the H. pylori group administered the IgY, F, or control diet. Furthermore, in the gerbils administered the F + IgY diet prior to the bacterial inoculation, inhibition of H. pylori colonization and suppression of the elevated gastric mucosal MPO activity were observed. Conclusions. Oral administration of urease-specific IgY not only inhibited H. pylori disease activity in H. pylori-infected gerbils, but also prevented H. pylori colonization in those not yet infected. These encouraging results may pave the way for a novel therapeutic and prophylactic approach in the management of H. pylori-associated gastroduodenal disease.

Recently, chicken egg yolk was recognized as an inexpensive antibody source, and the therapeutic usefulness of egg yolk immunoglobulin Y (IgY) in oral passive immunization has been investigated. Although multiple antibiotic treatments eradicate most Helicobacter pylori (H. pylori) infections, therapy fails in 10-15% of cases due to the development of drug resistance. Consequently, it is important that new, more broadly based therapies for the treatment of H. pylori infection should be identified. The present study evaluated the effect, on H. pylori infection, of IgY prepared from egg yolk of hens immunized with H. pylori urease (anti-HpU IgY). Seventeen asymptomatic volunteers diagnosed as H. pylori-positive by the 1 3 C-urea breath test (UBT) were orally administered anti-HpU IgY for 4 weeks. Four weeks later, UBT values were significantly decreased although no case showed H. pylori eradication. An H. pylori-positive 53-year-old female gastritis patient administered anti-HpU IgY plus lansoprazole for 8 weeks showed a decrease in serum pepsinogen (PG) I and UBT values as well as an increase in the PG I/II ratio. In conclusion, anti-HpU IgY may mitigate H. pylori-associated gastritis and partially attenuate gastric urease activity. Furthermore, anti-HpU IgY combined with antacids appears to ameliorate gastric inflammation. These encouraging results may represent a novel approach to the management of H. pylon-associated gastroduodenal disease.

We examined the effects of exercise training (treadmill running over 9 weeks) on the ability of isolated adipocytes to secrete tumor necrosis factor-alpha (TNF-alpha) and type 1 soluble TNF receptor (sTNFR1) in vitro in Wistar rats. We also examined the effects of exercise training on the expression of membrane bound forms of type 1 TNF receptor (mTNFR1) in adipocyte crude membranes of the same rat subjects. Exercise training significantly increased the secretions of TNF-alpha from isolated adipocytes. Treatment with a cyclooxygenase inhibitor, either indomethacin (100 microM) or eicosatetraynoic acid (100 microM), significantly blocked the release of TNF-alpha from adipocytes in both exercise-trained rat group and sedentary control rat group, suggesting that some cyclooxygenase metabolite(s) acts as a ligand in TNF-alpha synthesis. Decreased amounts of TNF-alpha were found to be significantly greater in both exercise-trained rat group than in sedentary control rat group after incubation with inhibitors. Thus, the inhibitory effect of both indomethacin and eicosatetraynoic acid was significantly greater in adipocytes from exercise-trained rats. Both plasma sTNFR1 levels and adipocytes-derived sTNFR1 were found to be significantly less in the exercise-trained rat group. Western blot analysis revealed that exercise training remarkably increased the expressions of mTNFR1 in adipocyte crude membrane. Thus, exercise training enhanced the ability of isolated adipocytes to secrete TNF-alpha with reduced secretion of sTNFR1, and provoked the greater expressions of mTNFR1 in adipocyte crude membrane. These alterations may induce enhanced the autocrine effects of TNF-alpha within adipocytes in exercise-trained rats.

Release process of enzymes from baker's yeast accompanied by inactivation was experimentally investigated by using an agitating bead mill. The enzymes examined were Alkaline phosphatase and alcohol dehydrogenase. As a result, the disruption rate of yeast was directly proportional to the product of the concentration of undisrupted yeast and agitating speed of impeller, and the disruption process of yeast was clarified by this analysis. In order to investigate the inactivation process of enzymes, the activity of enzymes within the yeast was measured using a lytic method, the results of which are not influenced by the mechanical force. In the case where there is no inactivation, the release rate of enzymes was directly proportional to the product of the activity of unreleased enzymes and agitating speed of impeller. In the case of inactivation, the release rate of enzymes was directly proportional to the product of the activity of uninactivated enzymes and agitating speed of impeller. The experimentally analyzed models based on the kinetics were found to describe well the release process of enzymes accompanied by the inactivation from baker's yeast, disrupted by the agitating bead mill.

Circulating concentrations of adipocytokines, such as leptin, tumor necrosis factor-alpha (TNF-alpha), and plasminogen activator inhibitor-1 (PAI-1), vary with exercise training, menopause, or regional variations in adipose tissues. In the present study, the relationships between body fat distributions and some adipocytokines were compared in premenopausal (N = 9) and postmenopausal women (N = 9), before and after exercise training. The training for 10 weeks (that is, 3 days/week) consisted of aqua exercise plus resistance exercise. The training reduced percent fat, body mass index, total fat mass (TFM), subcutaneous fat mass (SFM), and plasma levels of leptin and PAI-1 in both women. Mean value of plasma TNF-alpha tended to increase after training in both women. Plasma leptin levels were lower in postmenopausal than in premenopausal women, independently of training. In premenopausal women, plasma leptin levels correlated well with either TFM or SFM before and after training. Regression analysis of decreases in plasma leptin with a reduced amount of SFM revealed that decreases in plasma leptin depended to a greater extent on a loss of SFM. In postmenopausal women, no significant correlation was found between leptin levels and any of the fat depots. Plasma TNF-alpha levels correlated well with visceral fat mass (VFM) in premenopausal but not in postmenopausal women. Changes in TNF-alpha levels after training correlated well with reduced amount of VFM in premenopausal but not in postmenopausal women. Plasma PAI-1 levels were not different between groups. Moreover, no significant correlation was found between PAI-1 levels and any of the fat depots in both women. Thus, in premenopausal but not in postmenopausal women, changes in plasma concentrations of leptin and TNF-alpha correlate well with specific alterations in relative amount of SFM and VFM after training, respectively. However, no significant relationship between PAI-1 and any of the fat depots was found independently of either menopause or training.

Twenty-five neurons in the region of the nucleus of the solitary tract (NTS) were antidromically activated by electrical stimulation of the subfornical organ (SFO) in male rats under urethane anesthesia. Microiontophoretically applied bicuculline, a gamma-aminobutyric acid (GABA)(A) antagonist, but not phaclofen, a GABA(B) antagonist, attenuated the post-antidromic inhibitory response evoked by SFO stimulation of approximately two-third (n=17) of identified neurons, indicating the existence of recurrent inhibitory systems through GABA(A) receptors. Iontophoretically applied GABA decreased the spontaneous activity of all identified neurons, and the GABA-induced inhibition was prevented by simultaneously applied bicuculline, but not by phaclofen. Activation of peripheral baroreceptors, achieved by rising arterial blood pressure with an intravenous infusions of phenylepherine, suppressed the activity of the majority (n=20) of identified neurons. The inhibitory response of identified neurons (n=7) to baroreceptor activation was partially antagonized by iontophoretically applied bicuculline, but not by phaclofen. These results imply that GABAergic mechanisms may modulate the baroreceptor reflex acting on GABA(A) receptors of NTS neurons with ascending projections to the SFO in the region of the NTS.

It has been shown that amyloid β (Aβ) secretion regulates cholesterol efflux from cells and that the E693Δ (Osaka) mutation in amyloid precursor protein (APP) promotes intracellular accumulation of Aβ and thus reduces its secretion. These findings led us to speculate that APP with the Osaka mutation (APPOSK ) might have a defect in cholesterol efflux and thus cause cellular malfunction. We therefore examined the effects of this mutation on intracellular cholesterol transport and efflux in cultured cells. Upon cholesterol loading, APPOSK -expressing cells exhibited higher levels of cellular cholesterol than wild-type APP-expressing cells, suggesting impaired cholesterol efflux. It is known that, after its internalization, cholesterol is transported from the endosomes to the endoplasmic reticulum (ER) and Golgi apparatus and then to the plasma membrane. In APPOSK -expressing cells, cholesterol accumulated with Aβ in the ER and Golgi apparatus and alone in endosomes/lysosomes. These results imply that the mutation-induced disturbance of Aβ trafficking from the ER to the plasma membrane affects cholesterol transport to cause cholesterol accumulation in the ER and Golgi apparatus and, consequently, in endosomes. Furthermore, we detected an enhanced mitochondrial accumulation of Aβ and cholesterol in APPOSK -expressing cells, and this was accompanied by an increase in the generation of reactive oxygen species (ROS). The present findings suggest that Aβ trafficking is important for intracellular cholesterol transport and efflux and that the Osaka mutation potentiates cholesterol-dependent exacerbation of intracellular Aβ toxicity, i.e. Aβ-induced ROS generation, by disturbing Aβ-mediated cholesterol efflux from the cell.

The actual levels of steroid hormones in organs are vital for endocrine, reproductive and neuronal health and disorders. We developed an accurate method to determine the levels of steroid hormones and steroid conjugates in various organs by an efficient preparation using a solid-phase-extraction cartridge. Each steroid was identified by the precursor ion spectra using liquid chromatography–electrospray ionization time-of-flight mass spectrometry, and the respective steroids were quantitatively analysed in the selected reaction monitoring mode by liquid chromatograph-mass spectrometry/mass spectrometry (LC-MS/MS). The data showed that significant levels of testosterone, corticosterone and precursors of both hormones were detected in all organs except liver. The glucuronide conjugates of steroid hormones and the precursors were detected in all organs except liver, but sulfate conjugates of these steroids were observed only in the target organs of the hormones and kidney. Interestingly, these steroids and the conjugates were not observed in the liver except pregnenolone. In conclusion, an accurate determination of tissue steroids was developed using LC-MS analysis. Biosynthesis of steroid hormones from the precursors was estimated even in the target organs, and the delivery of these steroid conjugates was also suggested via the circulation without any significant hepatic participation.

Aims Hypercholesterolemia is known to be a risk factor for Alzheimer's disease (AD), and diet-induced hypercholesterolemia has been shown to accelerate amyloid pathology in animals. While growing evidence has shown that synaptic and cognitive dysfunction in AD is associated with intraneuronal accumulation of Aβ, the relationships between hypercholesterolemia, memory impairment, and intraneuronal Aβ remains unclear. The present study aims to clarify this association. Main methods Transgenic mice expressing amyloid precursor protein (APP) harboring the Osaka (E693∆) mutation (APPOSK-Tg mice) were used. These mice exhibit intraneuronal Aβ oligomers and memory impairment from 8 months of age. Five-month-old male APPOSK-Tg mice and non-Tg littermates were fed a high-cholesterol diet for 1 month to induce hypercholesterolemia. At 6 months of age, their cognitive function was evaluated by the Morris water maze. Intraneuronal Aβ, synaptic density, and tau phosphorylation were examined by immunohistochemistry. Key findings Serum and brain cholesterol levels were significantly higher in APPOSK-Tg mice and non-Tg littermates that were fed a high-cholesterol diet than in control mice that were fed normal chow, indicating that hypercholesterolemia was successfully induced. Hypercholesterolemic APPOSK-Tg mice, but not control APPOSK-Tg mice or hypercholesterolemic non-Tg littermates, exhibited impaired spatial reference memory, which was accompanied with intraneuronal accumulation of Aβ oligomers, reduced synaptophysin immunoreactivity, and abnormal tau phosphorylation in the hippocampus. Hypercholesterolemia-accelerated accumulation of intraneuronal Aβ oligomers was also observed in another model mouse, Tg2576. Significance Our findings suggest that hypercholesterolemia accelerates intraneuronal accumulation of Aβ oligomers and subsequent synapse loss, resulting in memory impairment.

Effects of hindlimb suspension (HS) and ambulation recovery on hippocampal neurogenesis of newly weaned rats were studied by using immunohistochemical techniques. The number of proliferating cell nuclear antigen-positive (PCNA(+)) cells in the subgranular zone (SGZ) markedly decreased during normal growth. However, neither HS nor subsequent recovery caused additional changes in the number of PCNA(+) cells. The number of doublecortin-positive (DCX(+)) neurons decreased gradually during normal growth. HS resulted in a further decrease in these neurons. However, DCX(+) cell numbers became identical to the levels in age-matched controls after 14 days of recovery. PCNA and DCX-double positive cells in the SGZ were also observed, and their cell numbers were not affected by HS and 14-day ambulation. Thus, HS suppressed the generation of DCX(+) neurons without affecting PCNA(+) cells in the SGZ of weaned rats. Taken together, hippocampal neurogenesis in weaned rats was not severely affected by HS while it decreased significantly as they had grown.

A powdered diet containing 100 or 3 ppm Fe was fed to rats starting at the age of 3 weeks. The voluntary activity level was checked using a wheel in the cage during the 17th week after the beginning of supplementation. Significantly less activity was seen in the 3 ppm Fe group during both light and dark periods. After 20 weeks, the blood and diencephalon were sampled from both groups. Lower hematocrit and blood hemoglobin content was observed in the 3 ppm Fe group. The level of 70 kDa heat shock cognate (HSC70) expression was greater in the diencephalon of the 3 ppm Fe group. In addition, the distribution of HSC70 was determined by proximity ligation assay. More HSC70-positive as well as total cells were noted in several areas of the diencephalon of the iron-deficient rats. The altered expression and distribution of HSC70 might play some role in the neurological changes.

Many cases of encephalitic disease with unknown etiologies have been reported in specific breeds of small dogs. High mobility group box 1 (HMGB1) in neuronal cells was recently found to be a novel cytokine-like mediator that is a marker of neuronal necrosis and inflammation. The aim of this study was to determine whether HMGB1 levels are elevated in the cerebrospinal fluid (CSF) of dogs suspected of having encephalitis. CSF was obtained from 31 dogs that were diagnosed with an encephalitic disease by clinical examinations and magnetic resonance image (MRI) scanning. The CSF samples were analyzed via western blotting (WB) and an enzyme-linked immunosorbent assay (ELISA) with a polyclonal antibody against HMGB1. The mean HMGB1 concentration was significantly higher in the encephalitic dogs than that in the healthy controls. The concentrations of HMGB1 were correlated with the cell counts and total protein concentrations, which are known CSF indicators of the neuronal inflammation associated with encephalitis. These results suggest that HMGB1 protein in CSF confirms the presence of necrosis and inflammation in most cases of canine encephalitis and that HMGB1 will be a new indicator of encephalitis.

The purpose of the present study was to investigate the effect of acute exercise on lipolysis via coordination of hormone-sensitive lipase (HSL) and scaffold proteins, i.e., perilipin A and comparative gene identification-58 (CGI-58), in rat primary adipocytes. Glycerol release was significantly elevated immediately (0 h) and three hours (3 h) after exercise. Both activity and localization to the pellet of HSL were significantly greater in the pellet fraction, which is included in lipid droplet associated-proteins, than in the supernatant fraction. In the pellet fraction, although neither perilipin A nor CGI-58 protein level changed, level of perilipin A/CGI-58 complex was significantly reduced, accompanied by up-regulated association of perilipin A/HSL at 0 h and 3 h after exercise. On the other hand, there were no changes in these molecules at 24 h after exercise, despite a significant decrease in lipolysis that was observed in response to isoproterenol. These findings suggest that acute exercise enhances lipolysis up to at least 3 h after exercise in a manner dependent on modification of HSL and its association with and alteration in scaffold protein.

It has been proposed that mechanically induced tension is the critical factor in the induction of muscle hypertrophy. However, the molecular mechanisms involved in this process are still under investigation. In the present study, the effect of mechanical stretch on intracellular signaling for protein translation initiation and elongation was studied in C2C12 myoblasts. Cells were grown on a silicone elastomer chamber and subjected to 30-min of 5 or 15% constant static or cyclic (60 cycles/min) uniaxial stretch. Western blot analyses revealed that p70 S6 kinase (p70S6K) and eukaryotic elongation factor 2 (eEF2), which are the markers for translation initiation and peptide chain elongation, respectively, were activated by both static and cyclic stretch. The magnitude of activation was greater in response to the 15% cyclic stretch. Cyclic stretch also increased the phosphorylation of MAP kinases (p38 MAPK, ERK1/2 and JNK). However, the pharmacological inhibition of MAP kinases did not block the stretch-induced activation of p70S6K and eEF2. An inhibitor of the mammalian target of rapamycin (mTOR) blocked the stretch-induced phosphorylation of p70S6K but did not affect the eEF2 activation. A broad-range tyrosine kinase inhibitor, genistein, blocked the stretch-induced activation of p70S6K and eEF2, whereas Src tyrosine kinase and Janus kinase (JAK) inhibitors did not. These results suggest that the stretch-induced activation of protein translation initiation and elongation in mouse myoblast cell lines is mediated by tyrosine kinase(s), except for Src kinase or JAK.