別府 英博

Obesity markedly increases the risk of colorectal cancer. Recently, the preventive effects of edible mushrooms on triglyceride elevation and visceral fat accumulation have been reported. Here, the effects of Pleurotus eryngii (Eringi) and Hypsizygus marmoreus (Bunashimeji) on azoxymethane (AOM)-induced aberrant crypt foci (ACF precancerous lesions) in the colorectums of mice fed a high-fat diet were examined. Eringi (ER) and Bunashimeji (BU) mushroom powder samples were used. Six-week-old male C57BL/6J mice received an intraperitoneal injection of AOM (10 mg/kg) once a week for two weeks, and were sacrificed and dissected at 6 weeks after the start of the experiment. After the initiation of the experiment, they received a normal diet (ND), high-fat diet (HFD), HFD + ER (1 or 5% of diet), or HFD + BU (1 or 5% of diet). As a result, body and fat weights were significantly lower in the 5% ER and BU groups than in the HFD group. Liver triglyceride levels were also significantly lower in the 5% ER and BU groups. Total liver cholesterol levels were significantly lower in the 5% ER group. The numbers of ACF (especially large ACF) showed strong inhibitory effects in both ER and BU groups. Measurement of the cell proliferation marker Ki-67 labeling index in the colonic mucosa demonstrated more significant suppression in both ER and BU groups than in the HFD group. These results suggest that the simultaneous intake of ER and BU may inhibit colorectal tumorigenesis in HFD-fed mice.

Aloe-emodin (1, 8-dihydroxy-3-hydroxyl-methylanthraquinone AE) and emodin (1,3,8-trihydroxy-6-methylanthraquinone EM) are anthraquinone derivatives that have been detected in some medical plants and share similar anthraquinone structures. AE and EM have been shown to exhibit anticancer activities in various cancer cell lines however, the inhibitory effects of these derivatives on the growth of cancer cells were previously reported to be different. Gastric cancer is the second most common cause of cancer cell death worldwide. In the present study, we examined the inhibitory effects of 0.05 mM AE and 0.05 mM EM on the proliferation of the MKN45 human gastric cancer cell line. The proliferation of MKN45 cells was significantly inhibited in AE- and EM-treated groups 24 h and 48 h after treatment. Furthermore, the inhibitory effects of EM were stronger than those of AE. The cell cycle of MKN45 cells were arrested in G0/G1 phase or G0/G1 and G2/M phases by AE and EM, respectively. However, an analysis of intracellular polyamine levels and DNA fragmentation revealed that the mechanisms underlying cell death following cell arrest induced by AE and EM differed.

Aloe vera gel exhibits protective effects against insulin resistance as well as lipid-lowering and anti-diabetic effects. The anti-diabetic compounds in this gel were identified as Aloe-sterols. Aloe vera gel extract (AVGE) containing Aloe-sterols has recently been produced using a new procedure. We previously reported that AVGE reduced large-sized intestinal polyps in Apc-deficient Min mice fed a high fat diet (HFD), suggesting that Aloe vera gel may protect against colorectal cancer. In the present study, we examined the effects of Aloe vera gel powder (AVGP) and AVGE on azoxymethane-induced colorectal preneoplastic aberrant crypt foci (ACF) in mice fed a HFD. Male C57BL/6J mice were given a normal diet (ND), HFD, HFD containing 0.5% carboxymethyl cellulose solution, which was used as a solvent for AVGE (HFDC), HFD containing 3% or 1% AVGP, and HFDC containing 0.0125% (H-) or 0.00375% (L-) AVGE. The number of ACF was significantly lower in mice given 3% AVGP and H-AVGE than in those given HFD or HFDC alone. Moreover, 3% AVGP, H-AVGE and L-AVGE significantly decreased the mean Ki-67 labeling index, assessed as a measure of cell proliferation in the colonic mucosa. In addition, hepatic phase II enzyme glutathione S-transferase mRNA levels were higher in the H-AVGE group than in the HFDC group. These results suggest that both AVGP and AVGE may have chemopreventive effects on colorectal carcinogenesis under the HFD condition. Furthermore, the concentration of Aloe-sterols was similar between 3% AVGP and H-AVGE, suggesting that Aloe-sterols were the main active ingredients in this experiment.

Predicting recovery from hemiparesis after stroke is important for rehabilitation. A few recent studies reported that the levels of some growth factors shortly after stroke were positively correlated with the clinical outcomes during the chronic phase. The aim of this study was to examine the relationships between the serum levels of growth factors (vascular endothelial growth factor [VEGF], insulin-like growth factor-I [IGF-I], and hepatocyte growth factor [HGF]) and improvement in hemiparesis in stroke patients who received rehabilitation in a postacute rehabilitation hospital. Subjects were 32 stroke patients (cerebral infarction: 21 and intracerebral hemorrhage [ICH]: 11). We measured serum levels of VEGF, IGF-I, and HGF and 5 items of the Stroke Impairment Assessment Set (SIAS) for hemiparesis on admission and at discharge. Age-matched healthy subjects (n 5 15) served as controls. Serum levels of VEGF and HGF in cerebral infarct patients on admission were higher than those in control subjects, and the serum levels of IGF-I in stroke patients were lower than those in controls. The level of HGF in ICH patients on admission was negatively correlated with gains in SIAS, and higher outliers in HGF concentration were correlated with lower gains in SIAS. Focusing on the extremely high levels of these factors may be a predictor of the low recovery from hemiparesis after stroke.

Aloe-emodin (AE), a natural anthraquinone compound, has been reported to exhibit anticancer activity in various cancer cell lines and anti-inflammatory effects in murine macrophages. In the present study, we investigated the cancer chemopreventive effects of AE in an Apc-deficient Min mouse model. In the first experiment, male Min mice were fed a basal diet or diets containing 5 ppm AE and 10 ppm AE for 12 weeks. The dietary administration of 5 ppm AE significantly reduced the number of colorectal tumors. In a second experiment, we investigated the effects of AE on colitis-related colon carcinogenesis in Min mouse treated with dextran sodium sulfate (DSS). Female Min mice were administered 1% DSS in their drinking water for 7 days. AE was given to mice in their diet at a dose of 5 or 50 ppm for 5 weeks. Feeding with AE significantly reduced the number of colorectal tumors. When proliferation of cells in normal-appearing colonic mucosa was assessed by monoclonal anti-rat Ki-67 antibody (MIB-5) immunohistochemistry in experiments 1 and 2, the AE treatment significantly decreased the mean MIB-5-labeling index. These results suggest that the dietary administration of low-dose AE may have chemopreventive effects against development of colorectal tumors in Min mice, possibly in part by reducing cell proliferation in colorectal mucosa.

Background: Although different gait analysis methods such as Walking Track Analysis exist, they cannot be used to demonstrate the physical condition of mice with specific gait disorder characteristic. Therefore, we developed a new method for the gait analysis of such mice to accurately assess hind limb angle based on the pelvic axis. New method: We established and verified a gait analysis method capable of pelvic axis-based limb angle measurement by video-recording the gait of a control mice group (C57BL/6J(B6)) and three ataxic mice (ataxic B6-wob/t, Parkinson's disease model (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treated (MPTP)), and cerebellum hypoplasia (cytosine-beta-D-arabinofuranoside treated)) from the ventral side. Results: The assessed hind limb angles of B6-wob/t and MPTP-treated mice were significantly wider than B6 mice (p<0.01). Moreover, we could draw separating lines with slopes of minus one that could separate the data of each group in-the scatter plot of the normalized hind limb step width and angle. Comparison with existing methods: We found no significance when we applied the already existing nose-tail method for the analysis of the hind limb angles of B6 and B6-woblt mice. In the nose-tail method, since the whole body axis of the trunk varies while the trunk of the mouse is laterally bent changing the hind limb angle, B6 and B6-wob/t mice could not be differentiated. However, the two mice groups could be differentiated by the pelvic axis-based gait analysis method. Conclusion: The pelvic axis-based gait analysis method is promising and valid for mice with gait disorder. (C) 2013 Elsevier B.V. All rights reserved.

Aloe vera gel supercritical CO2 extract (AVGE) has been shown to contain five phytosterols, reduce visceral fat accumulation, and influence the metabolism of glucose and lipids in animal model experiments. Recent epidemiologic studies have shown that obesity is an established risk factor for several cancers including colorectal cancer. Therefore, we examined the effects of AVGE on intestinal polyp formation in Apc-deficient Min mice fed a high-fat diet. Male Min mice were divided into normal diet (ND), high fat diet (HFD), low dose AVGE (HFD+LAVGE) and high dose AVGE (HFD+HAVGE) groups. The ND group received AIN-93G diet and the latter 3 groups were given modified high-fat AIN-93G diet (HFD) for 7 weeks. AVGE was suspended in 0.5% carboxymethyl cellulose (CMC) and administered orally to mice in HFD+LAVGE and HFD+HAVGE groups every day (except on Sunday) for 7 weeks at a dose of 3.75 and 12.5 mg/kg body weight, respectively. ND and HFD groups received 0.5% CMC alone. Between weeks 4 and 7, body weights in the HFD and HFD+LAVGE groups were reduced more than those in the ND group. However, body weights were not reduced in the HFD+HAVGE group. Mice were sacrificed at the end of the experiment and their intestines were scored for polyps. No significant differences were observed in either the incidence and multiplicity of intestinal polyps (≤0.5 mm in a diameter) among the three groups fed HFD. However, when intestinal polyps were categorized by their size into 0.5-1.4, 1.5-2.4, or ≥2.5 mm, the incidence and multiplicity of large polyps (≥2.5 mm) in the intestine in the HFD+HAVGE group were significantly lower than those in the HFD group. We measured plasma lipid (triglycerides and total cholesterol) and adipocytokine [interleukin-6 and high molecular weight (HMW) adiponectin] levels as possible indicators of mechanisms of inhibition. The results showed that HMW adiponectin levels in the HFD group were significantly lower than those in the ND group. However, the levels in the HFD+HAVGE group were significantly higher than those in the HFD group. These results indicate that HAVGE reduced large-sized intestinal polyps and ameliorated reduction in plasma HMW adiponectin levels in Min mice fed HFD.

High temperature- and pressure-treated garlic (HTPG) has been shown to have enhanced antioxidative activity and polyphenol contents. Previously, we reported that HTPG inhibited 1,2-dimethylhydrazine-induced mucin depleted foci (premalignant lesions) and O-6-methylguanine DNA adduct formation in the rat colorectum. In the present study, we investigated the modifying effects of HTPG on N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG)-induced pyloric stomach and small intestinal carcinogenesis in mice. Male C57BL/6 mice were given ENNG (100 mg/l) in drinking water for the first 4 weeks, then a basal diet or diet containing 2% or 5% HTPG for 30 weeks. The incidence and multiplicity of pyloric stomach and small intestinal (duodenal and jejunal) tumors in the 2% HTPG group (but not in the 5% HTPG group) were significantly lower than those in the control group. Cell proliferation of normal-appearing duodenal mucosa was assessed by MIB-5 immunohistochemistry and shown to be significantly lower with 2% HTPG (but again not 5% HTPG) than in controls. These results in dicate that HTPG, at 2% in the diet, inhibited ENNG-induced pyloric stomach and small intestinal (especially duodenal) tumorigenesis in mice, associated with suppression of cell proliferation.

Identification of functional molecules in the brain related to improvement of the degree of paralysis or increase of activities will contribute to establishing a new treatment strategy for stroke rehabilitation. Hence, protein expression changes in the cerebral cortex of rat groups with/without voluntary exercise using a running wheel after cerebral infarction were examined in this study. Motor performance measured by the accelerated rotarod test and alteration of protein expression using antibody microarray analysis comprised 725 different antibodies in the cerebral cortex adjacent to infarction area were examined. In behavioral evaluation, the mean latency until falling from the rotating rod in the group with voluntary exercise for five days was significantly longer than that in the group without voluntary exercise. In protein expression profile, fifteen proteins showed significant quantitative changes after voluntary exercise for five days compared to rats without exercise. Up-regulated proteins were involved in protein phosphorylation, stress response, cell structure and motility, DNA replication and neurogenesis (11 proteins). In contrast, down-regulated proteins were related to apoptosis, cell adhesion and proteolysis (4 proteins). Additional protein expression analysis showed that both growth-associated protein 43 (GAP43) and phosphorylated serine41 GAP43 (pSer41-GAP43) were significantly increased. These protein expression changes may be related to the underlying mechanisms of exercise-induced paralysis recovery, that is, neurite formation, and remodeling of synaptic connections may be through the interaction of NGF, calmodulin, PKC and GAP43. In the present study at least some of the participation of modulators associated with the improvement of paralysis might be detected. (C) 2011 Elsevier B.V. All rights reserved.

Background: Agaricus blazei Murrill (ABM) has been shown to exhibit immunostimulatory and anti-cancer activities; however, its mechanism of action is poorly understood. We recently found that the diffusible fraction of hot-water extract of ABM exhibits anti-tumor activity toward leukemic cells, and identified it as agaritine, a hydrazine-containing compound. In the present study, we examined the morphological and cytochemical effects of agaritine on U937 cells to elucidate the tumoricidal mechanism of agaritine. Methods: Surface expression of phosphatidylserine (evaluated by annexin V binding), Fas antigen, DNA cleavage using TUNEL staining, changes in caspase activities and cytochrome c release, before and after treatment with agaritine, were examined using U937 cells. Results: Nuclear damage, DNA fragmentation, was observed by Wright-Giemsa. TUNEL staining and agarose gel electrophoresis when U937 cells were incubated with 10 mu g/mL of agaritine for 48 h. Flow cytometric analysis indicated that agaritine augments the proportion of annexin V-positive U937 cells without significant change in Fas antigen expression. Activities of caspase-3, -8 and -9 were gradually increased after the addition of agaritine. In the presence of caspase-3 or granzyme B inhibitor, except for the caspase-8 inhibitor, annexin V expression was significantly decreased, suggesting that mainly caspase-3 and -9 participate in the apoptotic pathway. Furthermore, cytochrome c release was detected by western blotting analysis after agaritine treatment. Conclusions: These results strongly suggest that the ABM constituent agaritine moderately induces apoptosis in U937 leukemic cells via caspase activation through cytochrome c release from mitochondria. General significance: This is the first report suggesting that the anti-tumor effect of agaritine is mediated through apoptosis. The present results might provide helpful suggestions for the design of anti-tumor drugs toward leukemia patients. (C) 2011 Elsevier B.V. All rights reserved.

Background: Mushrooms of the genus Agaricus are a common folk remedy against carcinoma. The active ingredients, polysaccharides and protein-polysaccharide complexes containing beta-glucan, have been isolated and shown to have indirect tumor-suppressing activity via an immunological activation. Methods: The diffusible fraction of a hot-water extract of Agaricus blazei Murrill (ABM) powder was fractionated by HPLC based on the anti-tumor activity against leukemic cells in vitro. The structure of the anti-tumor substance was determined by NMR and MS analyses. Results: We purified a tumorcidal substance from the diffusible fraction of ABM and identified it as agaritine, beta-N-(gamma-L(+)-glutamyl)-4-(hydroxymethyl) phenylhydrazine, having a molecular mass of 267 Da. This compound inhibited the proliferation of leukemic cell lines such as U937, MOLT4, HL60 and K562 with IC(50) values of 2.7, 9.4, 13.0, and 16.0 mu g/mL, respectively, but showed no significant effect on normal lymphatic cells at concentrations up to 40 mu g/mL. Although agaritine has been suspected of having genotoxic or carcinogenic properties, agaritine did not activate the umu gene of Salmonella, which reacts to carcinogens. General significance: The results indicate that agaritine from ABM has direct anti-tumor activity against leukemic tumor cells in vitro. This is in contrast to the carcinogenic activity previously ascribed to this compound. Our results also show that this activity is distinct from that of beta-glucan, which indirectly suppresses proliferation of tumor cells. (C) 2010 Elsevier B.V. All rights reserved.

The scavenging capacity of reactive oxygen species, such as hydroxyl radicals, is reported not to decrease in boiled garlic (an odorless garlic preparation). We therefore examined the modifying effect of boiled garlic powder (BGP) on 1,2-dimethylhydrazine-induced mucin-depleted foci (MDF) and aberrant crypt foci (ACF), preneoplastic lesions, in the rat colorectum. Male F344 rats (5 weeks old) were fed a basal diet, or experimental diets containing 5% or 1% BGP for 5 weeks. One week later, all rats were injected s.c. with DMH (40 mg/kg, once weekly for 2 weeks). At 10 weeks of age, all the rats were sacrificed, and the colorectum was evaluated for MDF and ACF. In rats given DMH and the 5% or 1% BGP diets (Groups 2 and 3), the numbers of MDF decreased significantly in a dose-dependent manner, compared with the DMH and basal diet value (Group 1) (p < 0.01). The numbers of ACF in Group 2, but not Group 3, showed a non-significant tendency to decrease. Next, the effects of BGP on the formation of DMH-induced O-6-methylguanine (O-6-MeG) DNA adducts in rats were studied. Male F344 rats (5 weeks old) were fed the basal diet, or 10% BGP diet for 5 weeks. All rats were injected i.p. once with 40 mg/kg DMH at the end of week 5. The animals were sacrificed 6 hours after DMH injection to analyze the O-6-MeG DNA adducts in the colorectal mucosa. Dietary administration of BGP significantly inhibited the O-6-MeG DNA adduct levels in the colorectal mucosa, compared with the controls (p < 0.01). These results suggested that BGP may exert chemopreventive effects against colon carcinogenesis at least in the initiation stage.

サイクロデキストリン(CD)がコエンザイムQ10(CoQ10)の水への溶解性，融解熱，ヒトにおける吸収性に及ぼす影響を調べるため，CoQ10を20-24重量%含むCoQ10-CD複合体粉末を調製した．CoQ10の水溶性は，α-CD，デキストリンである程度の改善がみられた．一方，示差走査熱量計分析では，CoQ10の吸熱ピークがβ-CD，γ-CD，β-Iso^®の存在でほぼ消失したことから，β-CD，γ-CD，β-Iso^®は，粉末中のCoQ10をほぼ包接した不溶性複合体を形成することが確認された．吸収試験では，健康な成人女性20名を4群に分け，CoQ10原末，CoQ10-β-CD複合体粉末，CoQ10-γ-CD複合体粉末またはCoQ10-β-Iso^®複合体粉末のCoQ10として0.30 g相当量が絶食下で単回摂取され，摂取前(0時間)および摂取2，4，6，8，24時間後の血漿中総CoQ10濃度をHPLCで測定した．CoQ10摂取後の血漿中総CoQ10濃度から摂取前の濃度を引いた血漿中外因性総CoQ10濃度を求めて吸収性を評価したとき，CoQ10-β-CD複合体摂取群，CoQ10-γ-CD複合体摂取群，CoQ10-β-Iso^®複合体摂取群の摂取0-8時間における血漿中外因性総CoQ10濃度-時間曲線下面積は，CoQ10原末摂取群と比較して有意(p < 0.01)な増加が認められ，β-CD，γ-CD，β-Iso^®がヒトにおけるCoQ10の吸収を促進することが示された．

We carried out three experimental trials to determine antidiabetic effects of Aloe arborescens Miller components. Firstly, ICR mice which received frequent injections of streptozotocin (Sz) in small doses (low-dose Sz-induced diabetes mice) were fed ad libitum with basal diets supplemented with components of Aloe arborescens Miller var. natalensis Berger (Kidachi aloe) and Aloe vera Linne from 31 days before to 73 days after the Sz C, injections. Variation in blood glucose levels, incidence rates of insulitis and blood insulin levels were examined during the trial. As a result, groups receiving diets supplemented at the rate of 2% with whole leaf of Kidachi aloe and 10 KDa fraction powder (a fraction with less than 10 KDa molecular weight derived from Kidachi aloe leaf skin juice by ultra filtration) significantly suppressed the elevation of blood sugar as compared to a control group receiving basal diet. In contrast, there was no significant effect with Aloe vera leaf pulp powder. Insulitis emerged at the rate of 87% in the basal diet group. On the contrary, the whole aloe leaf and 10 KDa fraction groups significantly decreased the incidence of insulitis and incidence rates of whole aloe leaf and 10 KDa fraction powder were 51 and 38%, respectively. While insulin levels in the basal diet group averaged at 0.05 ng, more than four times the insulin level was observed in the 10 KDa group relative to the basal diet group. Secondary, the inhibitory effects of test materials on intestinal glucose absorption were observed using the jejunum of rats. A strong inhibitory action on intestinal glucose absorption was observed in the 10 KDa fraction powder group. Thirdly, phenol compounds derived from aloe in the blood serum and organs were quantitatively measured by a HPLC following forced administration of aloe components to rats to determine absorption kinetics of aloe components inside the body. The primary component of aloe phenol Compounds is the same component of the 10 KDa fraction powder and it was found in the pancreas and liver in addition to in the blood serum. The above results indicate that fore and aft when Sz injections could cause selective toxicity to B cells of islets, the dietary administration of 10 KDa fraction powder to mice would lead to the persistence of aloe phenol compound having an antioxidant activity in the pancreas and blood, which could protect islets of Langerhans from the destruction caused by methyl radical derived from Sz. The results also suggested the possibility of the 10 KDa fraction powder to alleviate the burden of insulin secretion as it has an inhibitory action on glucose absorption in the jejunum of rats. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

The protective actions of components isolated from Aloe arborescens Miller var. natalensis Berger (Kidachi aloe in Japanese) on streptozotocin (Sz)-induced necrosis of B cells in the pancreatic islets of the mouse were investigated to clarify its action mechanism involved in anti-diabetic effects. In this experiment, phenol low molecular weight components of aloin and alolin A that were anti-oxidants and derived from the leaf skin or pulp extract, an aloe carboxypeptidase fraction that is a inhibitor of enhanced vascular permeability and a glycoprotein component that decreases blood glucose were tested with mice precedently administered with Sz which is known as a cytotoxin specific to B cells. The results showed that the treatment group receiving Sz followed by the aloe carboxypeptidase fraction increased the inhibition of dye leakage by 75.8% (p &lt; 0.001) in the extract of whole pancreas in comparison to the control group and the aloe carboxypeptidase fraction group also increased the inhibition effect by 68.4% (P &lt; 0.001) in the extract of pancreatic islets as compared to the control group. The carboxypeptidase is an aloe-derived protease known to inhibit the acetic acid-related enhancement of intraperitoneal vascular permeability in mice. Further, the elevation of blood glucose in Sz-induced diabetic mice intraperitoneally given the aloe carboxypeptitase fraction was significantly (p &lt; 0.01-0.001) restrained at 3, 7 and 14 days after the injection as compared to the control group given solvent only. The results of this experiment suggested that the inhibitory effect on the enhancement of vascular permeability related to the vascular acute inflammatory response at Sz-induced lesions of pancreatic islets was involved in the action mechanism of this enzyme. (c) 2005 Elsevier GmbH. All rights reserved.

The protective actions of components isolated from Aloe arborescens Miller var. natalensis Berger (Kidachi aloe in Japanese) on streptozotocin (Sz)-induced necrosis of B cells in the pancreatic islets of the mouse were investigated to clarify its action mechanism involved in anti-diabetic effects. In this experiment, phenol low molecular weight components of aloin and alolin A that were anti-oxidants and derived from the leaf skin or pulp extract, an aloe carboxypeptidase fraction that is a inhibitor of enhanced vascular permeability and a glycoprotein component that decreases blood glucose were tested with mice precedently administered with Sz which is known as a cytotoxin specific to B cells. The results showed that the treatment group receiving Sz followed by the aloe carboxypeptidase fraction increased the inhibition of dye leakage by 75.8% (p &lt; 0.001) in the extract of whole pancreas in comparison to the control group and the aloe carboxypeptidase fraction group also increased the inhibition effect by 68.4% (P &lt; 0.001) in the extract of pancreatic islets as compared to the control group. The carboxypeptidase is an aloe-derived protease known to inhibit the acetic acid-related enhancement of intraperitoneal vascular permeability in mice. Further, the elevation of blood glucose in Sz-induced diabetic mice intraperitoneally given the aloe carboxypeptitase fraction was significantly (p &lt; 0.01-0.001) restrained at 3, 7 and 14 days after the injection as compared to the control group given solvent only. The results of this experiment suggested that the inhibitory effect on the enhancement of vascular permeability related to the vascular acute inflammatory response at Sz-induced lesions of pancreatic islets was involved in the action mechanism of this enzyme. (c) 2005 Elsevier GmbH. All rights reserved.