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We examined the effects of polymethoxyflavonoids (PMFs) on T helper (Th) 17 cell differentiation in vitro and in vivo. Five different PMFs including nobiletin (NOB), sudachitin (SUD), demethoxysudachitin, heptamethoxyflavone and natsudaidain were used for the in vitro study, and effects of those flavonoids on Th17 responses were investigated. NOB and heptamethoxyflavone significantly suppressed the proliferation response, but SUD, demethoxysudachitin and natsudaidain did not suppress the proliferation response. All of the five flavonoids decreased IL-17A production. Mice with experimentally induced autoimmune encephalomyelitis were used as an in vivo Th17 differentiation model. We focused on two flavonoids, NOB and SUD, and examined the effects of those flavonoids. NOB significantly suppressed Th17 cell proliferation and cytokine responses, but SUD only decreased proliferation responses. The results suggest that the suppressive effect of NOB on Th17 response in vivo is stronger than that of SUD.

The essential oil of Citrus sudachi (sudachi oil) is extracted from the peel of sudachi, a citrus plant. We investigated the effect of sudachi oil on immune function in both in vitro antigen (Ag) induced lymphocyte activation and in vivo Ag-specific immune response. In the in vitro study, the proliferative activity of splenocytes upon Ag-specific and non-specific stimulation was suppressed by treatment with sudachi oil in a dose-dependent manner. In addition, the expression level of Ag-presentation-related molecules and their Ag-presenting function on dendritic cells were suppressed by sudachi oil. To examine how sudachi oil regulates an Ag-specific immune response in vivo, mice were immunized with ovalbumin and the immune response of the mice was investigated. Ag-specific proliferation response of splenocytes from mice treated with sudachi essential oil was significantly suppressed. The results indicate that sudachi oil suppresses T cell and dendritic cell functions in vitro and Ag-specific T cell induction in vivo.

We examined how food choice motives and dietary habits changed during the COVID-19 pandemic.Four hundred elderly Japanese completed an online questionnaire in early May in 2021. Participants were retrospectively asked about their intake of food groups and food choice motives before and during the COVID-19 pandemic. Dietary diversity was determined using the dietary variety score calculated from the food frequency questionnaire with 10 food groups. The importance of each of the nine food choice motives for elderly people was assessed. Each scores ranged from 1 to 5. Changes in food choice motives and dietary behaviors during the COVID-19 pandemic were assessed using the paired t-test and a general linear model.Among the food choice motives, scores for the importance of weight control, physical well-being and economical efficiency significantly increased in both sexes (all p0.05). Dietary diversity score was lower during the COVID-19 pandemic than that before the pandemic in women (p=0.019), but there was no difference in men. In the multivariate adjustment model, physical well-being and economical efficiency were shown to have significant positive associations with the COVID-19 pandemic in women (p=0.034 and 0.009, respectively). In contrast, eating out was shown to have a significant inverse association with the COVID-19 pandemic in women (p=0.009).The findings suggest that the COVID-19 pandemic was associated with an increase in some food choice motives and a decrease in the frequency of eating out among elderly female Japanese.

Caveolin-1 is a major protein of the caveolae structure in vascular endothelial cell membrane. Phosphorylation of caveolin-1 is one of the initial events leading to exacerbation of vascular permeability caused by oxidative stress. Although quercetin is known to be an anti-atherosclerosis factor that acts as a dietary antioxidant, little is known about its role in the regulation of caveolin-1 phosphorylation. In this study, we investigated the inhibitory effect of quercetin on hydrogen peroxide-induced caveolin-1 phosphorylation in human umbilical vein endothelial cells. Quercetin inhibited caveolin-1 phosphorylation in cells pretreated with quercetin for 24 h and then exposed to hydrogen peroxide. However, quercetin 3-O-β-glucuronide, a conjugated metabolite of quercetin, did not exert this inhibitory effect. Exposure to hydrogen peroxide increased vascular permeability and reduced mRNA expression of the intercellular adhesion protein, vascular endothelial cadherin (VE-cadherin). By contrast, pretreatment with quercetin suppressed the increase in vascular permeability and decreased VE-cadherin expression. These results indicate that deconjugated quercetin can play a role in the prevention of altered vascular permeability under oxidative stress by suppressing caveolin-1 phosphorylation. Thus, dietary quercetin may be beneficial for the maintenance of endothelial cell function.

BACKGROUND/OBJECTIVES To clarify the effects of intake of legumes and isoflavones on all-cause mortality in middle-aged and older Japanese. SUBJECT/METHODS The subjects were 2136 randomly chosen community-dwelling Japanese men and women aged 40-79 years who participated in a first-wave survey (1997-2000; baseline). The subjects were followed from baseline to December 31, 2017 (mean period of 16.6 ± 4.2 years). Intake of legumes, soybeans, and isoflavones at baseline was assessed using a 3-day dietary record with photographs. The relationships of intake amounts of legumes and isoflavones to mortality were assessed using Cox proportional hazards regression controlling for age, sex, education, employment, body mass index, smoking habits, medical histories, drinking, and energy intake. RESULTS There were 491 deaths during the follow-up period. We found inverse associations of the intake of total soy products and intake of each of the isoflavones with all-cause mortality (p for trend

Sudachitin is a polymethoxylated flavone found in the peel of Citrus sudachi, a unique specialty citrus fruit in Tokushima Prefecture, Japan. Previous reports have demonstrated that sudachitin has anti‑inflammatory and metabolic regulatory activities. However, to the best of our knowledge, no studies have explored whether sudachitin can act as an antitumor therapeutic agent by regulating metabolic functions in the tumor microenvironment. In the present study, cell proliferation and cytotoxicity assays were used to determine whether sudachitin inhibited the in vitro growth of liver cancer and pancreatic carcinoma, cholangiocarcinoma and colorectal cancer cells and to compare its toxicity against normal fibroblasts and induced cancer‑associated fibroblasts (CAFs). Using lactate assays and reverse transcription‑quantitative PCR, the effects of sudachitin on glycolysis in CAFs were investigated. The effects of CAFs on malignant tumor cells were evaluated in vitro using cell proliferation, wound healing and invasion assays. As result, sudachitin inhibited various types of tumor cells with different half‑maximal inhibitory concentrations. Treatment with 50 µM sudachitin for 48 h suppressed tumor and CAFs proliferation but was not cytotoxic against normal fibroblasts. This dose also inhibited glycolysis in CAFs and, thus, diminished their pro‑tumorigenic activities. Overall, the present study revealed that sudachitin has promise as a safe and widely available natural antitumor adjuvant.

The validity of dietary variety score (DVS) using a short-form questionnaire has not been investigated using dietary diversity based on a quantitative distribution of consumed foods in older Japanese. We examined the association between DVS and objective dietary diversity using a Quantitative Index for Dietary Diversity (QUANTIDD) based on the quantitative distribution of foods consumed by older Japanese community dwellers. The subjects were 65 older Japanese community dwellers aged 60?79 years. We used two kinds of scores for assessment of dietary diversity. At first, dietary diversity was determined using DVS calculated from answers to a questionnaire about frequencies of intake of 10 food groups. Second, dietary intake was assessed using a 3-day dietary record with photographs, and dietary diversity was determined using QUANTIDD. The relationships between DVS and QUANTIDD were assessed using partial correlation coefficients controlling for confounders. The correlation coefficient between DVS and QUANTIDD was moderate (r=0.212-0.458). After controlling for confounders, those correlation coefficient between DVS and QUANTIDD remained moderate. The findings suggest that there was a moderate relationship between DVS and QUANTIDD, and DVS using a short-form questionnaire may be useful for assessing dietary diversity in older Japanese community dwellers. J. Med. Invest. 69 : 31-37, February, 2022.

We examined the initial attachment of E. coli to abiotic surfaces conditioned with cereal extracts. The extracts were water-soluble fractions prepared from flours of barley, quinoa, rice, and wheat. Strains used were E. coli ATCC 8739, E. coli NBRC 3301, E. coli NBRC 3302, E. coli NBRC 13168, E. coli NBRC 13891, and E. coli O157:H7 sakai. When surfaces of glass and stainless steel were conditioned at 25 °C for 30 min with 0.5% cereal extracts, significantly lower numbers of E. coli cells attached to the conditioned surfaces than unconditioned ones, irrespective of strains used. The highest activity in reduction of the number of E. coli cells attached to the abiotic surfaces was found in the wheat extract. The suppressive activity was stable after treatments of the extract by autoclave and enzymatic digestion with α-amylase and proteinase K. We purified the active compound by ammonium sulfate fractionation and gel filtration with HiPrep 16/60 Sephacryl S-200 HR after the enzymatic treatments. The purified compound showed an average molecular mass of about 300 kDa by light-scattering measurements. Analyses of its components indicated that the active compound was arabinoxylan; the molar ratios were 1.0 (arabinose) to 2.46 (xylose). Commercially available arabinoxylan (average molecular mass: 370 kDa) also showed the similar activity. To our knowledge, this is the first report on a dietary fiber from cereals which suppresses the initial attachment of E. coli to abiotic surfaces.

The aim of this study was to clarify the characteristics of lifestyle and health awareness according to dietary diversity in a Japanese worksite population. The participants were 1,312 men and women aged 20 to 63 years who were living in Tokushima Prefecture, Japan during the period 2012-2013. We obtained anthropometric data and information on lifestyle characteristics using a self-administered questionnaire. Dietary intake was assessed using a food frequency questionnaire, and dietary diversity was determined using the Quantitative Index for Dietary Diversity (QUANTIDD). The characteristics of lifestyle and health awareness according to quartiles of the QUANTIDD score were assessed using the chi-square test and a general linear model. The higher the QUANTIDD score was, the larger were the proportions of participants who knew the appropriate amount of dietary intake and participants who referred to nutritional component information when choosing and / or buying food. Among participants with higher QUANTIDD scores, the proportion of participants who considered their current diet was good was high in women, whereas the proportion of participants who wanted to improve their diet in the future was high in men. Those results indicate that higher dietary diversity was related to better characteristics of lifestyle and awareness of health. J. Med. Invest. 67 : 255-264, August, 2020.

We investigated the effects of Polygonum tinctorium Lour (PTL), a plant commonly known as indigo, on biological parameters in an animal model of high-fat diet-induced obesity. Wistar rats fed a high-fat diet and treated with PTL showed lower serum levels of triglycerides and total cholesterol levels and a higher serum levels of HDL cholesterol than those in Wistar rats fed a high-fat diet without PTL treatment. The weight of mesenteric fat in PTL-treated rats was decreased compared to that in control rats not treated with PTL. In addition, energy metabolic rate in the dark period, but not in the light period, in PTL-treated rats was higher than that in control rats. Although a significant difference was not observed, body weight in PTL-treated rats tended to be decreased compared to that in control rats. The results show that PTL improves serum lipid profiles in Wistar rats with high-fat diet-induced obesity. J. Med. Invest. 67 : 158-162, February, 2020.

The aim of this study was to clarify the associations of family members living together with healthrelated behaviors in Japanese young workers. The participants were 300 men and women aged 20-39 years in 2015 who had a job. A web-based self-administered questionnaire on status of partnering and parenting, number of family members living together, dietary habits, drinking habit, smoking habit, self-rated health, employment status, working time and commuting time was conducted through Internet. Multiple logistic regression analysis and general linear models were used to assess the association of family members living together with healthrelated behaviors. The multivariate-adjusted odds ratio (95% confidence interval, p-value) for current drinking in unmarried participants living with their parents compared to unmarried participants living alone was 0.35 (0.13-0.93, p=0.036). The adjusted means of frequency of breakfast skipping and frequency of eating out showed a trend for inverse associations with the presence of a partner and children. However, those associations disappeared after adjustment for age of youngest child. The findings suggest that the presence of parents might affect drinking behavior and that age of youngest child living together might affect the frequency of breakfast skipping in young Japanese workers. J. Med. Invest. 66 : 141-147, February, 2019.

Scdachitin is a polymethoxyflavone (5,7,4'-trihydroxy-6,8,3-trimethoxyflavone) that is found in the peel of Citrus sudachi. We examined the effect of sudachitin on immune response in ovalbumin-immunized BALB/c mice. Treatment with sudachitin increased ovalbumin-specific IL-4 and IL-10 productions. In addition, mice that received sudachitin showed higher levels of ovalbumin-specific IgE, IgG1 and IgG l production than did control mice. The antibody response to the thymus-independent antigen 2,4,6-trinitrophenyl Ficoll was not different in the control and sudachitin groups, suggesting that sudachitin does not directly stimulate antibody production. An in vitro study showed that treatment of sudachitin enhanced the ability of antigen presentation in bone marrow-derived dendritic cells. Furthermore, CD11c+ cells that had been treated with sudachitin showed increased expression of co-stimulatory molecules. The results indicate that sudachitin regulates immune function both in vivo and in vitro.

Data were derived from the National Institute for Longevity Sciences-Longitudinal Study of Aging. Subjects comprised 403 men and 373 women aged 60-81 years at baseline who participated in the follow-up study at least once. Bean, soy product and soy isoflavone intake was assessed using a 3-day dietary record at baseline. Cognitive function was assessed by the Mini-Mental State Examination (MMSE). MMSE scores of ≤23 were used to define cognitive impairment. The relationship between bean, soy product and soy isoflavone intake and cognitive impairment was assessed using a generalized estimating equation. Multivariate-adjusted odds ratios (95% confidence intervals) for cognitive impairment with a 1 s.d. increase in total bean, total soybean and total soy isoflavone intakes were 0.48 (0.28-0.81; p = 0.006), 0.51 (0.32-0.83; p = 0.007), and 0.55 (0.32-0.93; p = 0.026), respectively, in women. Total soybean and soy isoflavone intake might decrease the risk of cognitive impairment in elderly Japanese women.

Epidemiological investigations have shown that consumption of soybeans or soy foods reduces the risk of the development of cardiovascular disease, cancer and osteoporosis. The aim of this study was to determine the associations between different soy foods and inflammatory markers, including high-sensitivity C-reactive protein (hs-CRP), interleukin (IL)-6, and IL-18, in Japanese workers. The cross-sectional study included 1,426 Japanese workers (1,053 men and 373 women) aged 20 to 64 years. Intake of 12 soy foods was estimated by a validated food frequency questionnaire. Associations of total soy foods, fermented soy food, non-fermented soy food, soy isoflavone with hs-CRP, IL-6, and IL-18 levels were examined by general linear model regression analysis. We found that total fermented soy food intake was inversely associated with multivariable-adjusted geometric concentration of IL-6 in men (Q1:1.03 pg/mL, Q5:0.94 pg /mL;P for trend = 0.031). Furthermore, it was shown that IL-6 concentrations were inversely associated with miso intake (β = -0.068;p = 0.034) and soy sauce intake in men (β = -0.074;p = 0.018). This study suggests that intake of total fermented soy food, miso and soy sauce be associated with IL-6 concentrations in Japanese men. J. Med. Invest. 65:74-80, February, 2018.

Citrus Sudachi is the special local product of Tokushima Prefecture, and over 98% of Sudachi consumed in Japan every year is produced in Tokushima Prefecture. In this study, we evaluated the function of sudachi peel extract (SPE) using an animal model of obesity. C57BL/6 mice were fed a high-fat diet containing 1% SPE powder. Treatment with SPE significantly decreased body weight compared to that of mice fed a high-fat diet. A significant difference in body weight was observed between the control and SPE groups from 7 weeks after the start of the experiment, the significant difference continued until the end of the 14-week experiment. Reduction of blood glucose levels following insulin administration in SPE-treated mice was grater than that in control mice. Determination of mRNA expression in adipose tissue showed that the expression level of TNF-α in the SPE group was significantly decreased compared to that on the control group. These results suggest that SPE potentially has the ability to attenuate body weight gain. J. Med. Invest. 64: 20-23, February, 2017.

Propolis is a bee product with various biological properties. C57BL/6 mice were fed a high-fat diet and treated with propolis for 14 weeks. Body weight in mice treated with 2% propolis was less than that in control mice from 3 weeks after the start of treatment until 14 weeks except for the 7th week. Mice treated with propolis showed significantly lower epididymal fat weight and subcutaneous fat weight. Infiltration of epididymal fat by macrophages and T cells was reduced in the propolis group. Supplementation of propolis increased feces weight and fat content in feces, suggesting that mechanisms of weight reduction by propolis partly include a laxative effect and inhibition of fat absorption.

The aim of this study was to determine the associations of dietary diversity with prevalences of allergic diseases.The participants were 1,317 men and women aged 20 to 63 years who were living in Tokushima Prefecture, Japan during the period 2012-2013. We obtained anthropometric data and information on lifestyle characteristics and current medical histories of allergic diseases using a self-administered questionnaire. Dietary intake was assessed using a food frequency questionnaire, and dietary diversity was determined using the Quantitative Index for Dietary Diversity (QUANTIDD). The ORs and 95% CIs for each of the allergic diseases with a 1 standard deviation (SD) increase in the QUANTIDD score were estimated, controlling for age, family history of allergic diseases, education, smoking, drinking, physical activity, energy intake and BMI.Higher dietary diversity showed significant inverse dose-response relationships with allergic diseases and allergic rhinitis in women. Multivariate-adjusted ORs (95% CI) for allergic diseases and allergic rhinitis with 1 SD increase in the QUANTIDD score were 0.77 (95% CI: 0.60-0.98, p=0.037) and 0.69 (95% CI: 0.53-0.90, p=0.007), respectively, in women. There were no significant associations between dietary diversity and allergic diseases in men.The results indicate that there is an inverse association between higher dietary diversity and allergic rhinitis in Japanese female workers.

We examined the effect of nobiletin (5,6,7,8,3',4'-hexamethoxyflavone) on immune response in ovalbumin (OVA)-immunized mice. Treatment with nobiletin increased OVA-specific IL-4 and IL-10 production. In addition, mice that received nobiletin showed higher levels of OVA-specific IgE, IgG and IgG1 production than did control mice. The antibody response to the thymus-independent antigen 2,4,6-trinitrophenyl-Ficoll was not different in the control and nobiletin groups, suggesting that nobiletin does not directly stimulate antibody production. An in vitro study showed that treatment with nobiletin enhanced the ability of antigen presentation of bone marrow-derived dendritic cells. The in vivo and in vitro results indicate that nobiletin regulates immune function.

Adipocytokines arebioactive substances synthesizedand secretedbyfat cell. Previous studies have reported an association between weight loss and adipocytokines. However, these studies are inconsistent and they have not clarified the relationship between weight regain and changes in circulating levels of adipocytokines. In this study, we analyzed the relationship between weight and fat volume changes and adipocytokines. The subjects were 235obese people recruited in the Saku Control Obesity Program (SCOP). Participants were randomly assigned to either immediate (Group A) or delayed (Group B, control group). Group A participants were followed for another two years after completion of the one year intervention. As controls, Group B participants received the same intervention as Group A after a delay of one year. Then they were followed up for one year. The intervention consisted in a one-year lifestyle program to induce weight loss, based on a cognitive-behavioral approach. After the first year of the study, body weight, BMI, body fat and abdominal fat areas were significantly lower in group A participants, compared to controls. After the intervention, leptin levels were significantly lowered both in men and women. After one year follow-up, both men and women re-gained about 1.5 kg body weight on average. BMI, waist circumference, fat areas by CT and HBA1c significantly increased during the follow-up period. The change of adipocytokine levels by analysis of the quartile of body weight decrease and regain revealed that increased adiponectin and decreased leptin was noteworthy for weight reduction, while increase of leptin influenced the weight regain. In conclusion, our results suggest that leptin could have broad effects on the distribution off at tissues and on lipid metabolism. Leptin inversely associated with adiponectin, which in turn was necessary to decrease body weight. In particular, leptin decreased remarkably in the process of weight reduction, and its increase seemed to be related in weight regain. The observed increase of adiponectin seemed to be induced by reduction in fat volume.

The aim of this study was to determine the associations of intake of soy products and isoflavones with allergic diseases.We conducted a cross-sectional study in 1437 participants (aged 20-64 years) who were living in Tokushima Prefecture, Japan during the period 2010- 2011. We obtained anthropometric data and information on life style characteristics including dietary intake and current medical histories of allergic diseases using a structural self-administered questionnaire. Multiple logistic regression models were used to assess the associations of soy products and isoflavones with allergic diseases after controlling for age, family history of allergic diseases, smoking, drinking, physical activity, energy intake, BMI and dietary factors.Intake of soy products showed significant inverse dose-response relationships with allergic rhinitis. The third quartile for soy products had an adjusted OR of 0.56 (95% CI: 0.35-0.91) compared to the reference group (first quartile), though intake of soy products showed no dose-response relationship with atopic dermatitis. Intake of soy isoflavones showed a significant inverse dose-response relationship with atopic dermatitis, though the association between intake of soy isoflavones and atopic dermatitis was U-shaped after adjustments for potential confounders. On the other hand, the associations between intake of soy isoflavones and other allergic diseases were not significant.The results indicate that higher intake of soy products is associated with reduced risk of allergic rhinitis in Japanese workers. Furthermore, moderate intake amounts of soy products and soy isoflavones are associated with inverse risk of atopic dermatitis.

Objectives/hypothesis: We aimed to test the hypothesis that chemotherapy changes the gene expression of taste receptors in the tongue to induce dysgeusia in patients with head and neck cancer. Study design: Prospective observation study. Methods: We enrolled 21 patients who received chemoradiotherapy and five patients who underwent radiotherapy for head and neck cancer. The messenger RNA (mRNA) levels of the taste receptor subunits T1R1, T1R2, T1R3, and T2R5 were measured in lingual mucosa scrapings obtained with a small spatula. The perception thresholds of umami, sweet, and bitter tastes were assessed by the whole mouth gustatory test. Results: In four patients with severe stomatitis induced by chemoradiotherapy, the mRNA levels of T1R1, T1R2, T1R3, and T2R5 in the lingual mucosa were significantly decreased. However, in 17 patients with mild/moderate stomatitis, the mRNA levels of T1R3 were significantly and transiently decreased, whereas those of T1R1 and T1R2 remained unchanged and those of T2R5 mRNA were significantly and transiently increased after chemotherapy. There was a significant negative correlation between the perception thresholds of umami or sweet tastes and lingual mRNA levels of T1R3 in patients with mild/moderate stomatitis after chemotherapy. Although the perception threshold of bitter taste remained unchanged, lingual mRNA levels of T2R5 were significantly increased in patients who complained of phantogeusia after chemotherapy. Conclusion: Chemotherapy specifically changed the gene expression of T1R3 and T2R5 in head and neck cancer patients with mild/moderate stomatitis, resulting in both dysgeusia of umami and sweet tastes as well as phantogeusia. Level of evidence: 4. Laryngoscope, 126:E103-E109, 2016.

Excessive dietary phosphorus (P) has been speculated to be a risk factor for cardiovascular disease (CVD). Here, we performed a double-blinded crossover study to investigate the time- and dose-dependent effects of dietary P intake on endothelial function in healthy subjects. Sixteen healthy male volunteers were given meals containing 400, 800, and 1,200 mg P (P400, P800, and P1200 meals, respectively) with at least 7 days between doses. There were no differences in nutritional composition among the experimental diets except for P content. Blood biochemistry data and flow-mediated dilation (%FMD) of the brachial artery were measured while fasted, at 0 h, 1 h, 2 h, and 4 h after meal ingestion, and the next morning while fasted. The P800 and P1200 meals significantly increased serum P levels at 1-4 h after ingestion. A significant decrease in %FMD was observed between 1-4 h,while the P400 meal did not affect %FMD. We observed no differences among meals in serum P levels or %FMD the next morning. A significant negative correlation was observed between %FMD and serum P. These results indicate that excessive dietary P intake can acutely impair endothelial function in healthy people.

Recent evidence suggests that immune cells play an important role in differentiation of inflammatory macrophages in adipose tissue, which contributes to systemic chronic inflammation. Dietary ribonucleic acid (RNA) has been shown to modulate immune function. We hypothesized that RNA affects immune cell function in adipose tissue and then improves inflammatory response in adipose tissue. C57/BL6 mice and recombination activating gene-1 (RAG-1) knockout mice on a C57BL/6 mice background were fed a high-fat diet containing 1% RNA for 12 wk. An oral glucose tolerance test was performed. Supplementation of dietary RNA in C57BL/6 mice fed a high-fat diet resulted in a smaller area under the curve (AUC) after oral glucose administration than that for control mice. The mRNA expression levels of inflammation-related cytokines in adipose tissue and serum interleukin-6 levels were reduced by dietary RNA supplementation. Interestingly, reduction of the AUC value by RNA supplementation was abolished in T and B cell-deficient RAG-1 knockout mice. These results indicate that RNA improves inflammation in adipose tissue and reduces the AUC value following oral glucose administration in a T and B cell-dependent manner.

Citrus Sudachi is the special local product of Tokushima Prefecture, and over 98% of Sudachi consumed in Japan every year is produced in Tokushima Prefecture. In this study, we evaluated the function of sudachi peel extract (SPE) using an animal model of obesity. C57BL/6 mice were fed a high-fat diet containing 1% SPE powder. Treatment with SPE significantly decreased body weight compared to that of mice fed a high-fat diet. A significant difference in body weight was observed between the control and SPE groups from 7 weeks after the start of the experiment, the significant difference continued until the end of the 14-week experiment. Reduction of blood glucose levels following insulin administration in SPE-treated mice was grater than that in control mice. Determination of mRNA expression in adipose tissue showed that the expression level of TNF-α in the SPE group was significantly decreased compared to that on the control group. These results suggest that SPE potentially has the ability to attenuate body weight gain. J. Med. Invest. 64: 20-23, February, 2017.

ObjectiveThe purpose of the present study was to examine the association between soya food consumption and insulin resistance using baseline data of the Japan Multi-Institutional Collaborative Cohort (J-MICC) Study in Tokushima, Japan.DesignThis cross-sectional study included 1274 subjects, aged 34–70 years at baseline, living in Tokushima Prefecture between 2008 and 2013. Fasting blood samples were collected and information on lifestyle characteristics including soya food intake and medical history were obtained using a structured self-administered questionnaire. The homeostasis model assessment of insulin resistance (HOMA-IR) was measured and those with HOMA-IR≥2·5 were defined as having insulin resistance. Multiple logistic regression models were used to analyse the association between soya product intake and the prevalence of insulin resistance.SettingRural communities located in Tokushima Prefecture, Japan, between 2008 and 2013.SubjectsA total of 1148 adults (565 men and 583 women), aged 34–70 years.ResultsThe frequency of intake of miso soup, total non-fried soya products and total soya products showed significant inverse dose–response relationships with insulin resistance, after adjustments for potential confounders. When soya product intake was calculated as soya protein and isoflavone, the odds ratios of insulin resistance decreased significantly as the estimated intake of soya protein increased. Furthermore, significant inverse dose–response relationships were observed for total non-fried soya products and total soya products, after adjustment for total vegetable or total fibre consumption.ConclusionsThe present results indicate that the intake of soya products and non-fried soya products is associated with reduced insulin resistance in the Japanese population.

Buckwheat powder or protein has been shown to decrease the total serum cholesterol level in non-diabetic mice or rats. However, the lipid-lowering effect of buckwheat bran extract (BBE) in diabetic mice has not been fully elucidated. KK-Ay mice that received six-week treatment with BBE showed decreased body weight and liver weight compared to those of control (vehicle) mice. However, there was no significant difference in food intake. BBE treatments prevented liver triglyceride accumulation and decreased the serum level of triglycerides. In addition, mRNA expression levels lipogenic enzyme genes, fatty acid synthase, acetyl-coenzyme a oxidase and stearyl-coenzyme a desaturase 1, but not those of β-oxidized enzyme genes, were decreased in BBE-treated mice. Level of transcription factors ChREBP and SREBP1c, transcripts of lipogenic genes, were also decreased in BBE-treated mice. These results suggest that chronic treatment with BBE derivatives could have beneficial effects on hypertriglycemia in patients with type 2 diabetes mellitus.

Background: Multiplex immunofluorescence (mIF) is a key tool for cancer immunoprofiling in tissue samples. These techniques typically require an investigator to select regions of interest (ROI) within the tumor for digital image analysis. However, tumor heterogeneity and sampling bias may result in data that may not be representative of the whole tumor. Our goal was to compare immunoprofiling data from ROI analysis versus whole tumor in lung, colorectal, and bladder carcinomas using mIF and multispectral image analysis. Methods: 9 lung adenocarcinomas, 11 lung squamous cell carcinomas, 10 colorectal adenocarcinomas (CRC), and 10 bladder urothelial carcinomas (BC) were stained with a 6-marker mIF panel (PDL1, CD8, Ki67, CD68, AE1/AE3, PD1) using the Opal technique, and imaged using a Polaris multispectral scanner. Two pathologists independently selected 5 ROI (0.64 mm2) within the tumor for each case. To address the possibility of human sampling bias, 2 additional sets of 5 ROI were randomly generated in the tumor by a computer. HALO software was used to analyze the ROIs and the whole tumor area. The data was compared using Spearman’s ranked correlation coefficients. Results: 6 cell populations were assessed in the tumor epithelium and stroma: cytotoxic T-lymphocytes (CTL, CD8+), antigen-experienced CTL (CD8+/PD1+), proliferating CTL (CD8+/Ki67+), macrophages (CD68+), PDL1+ tumor cells (AE1AE3+/PDL1+), and proliferating tumor cells (AE1AE3+/Ki67+). These immunophenotypes were assessed in each set of 5 ROI and compared with data from the whole tumor. Spearman’s correlation coefficients (r) ranged from -0.25 to 1.00 depending on markers and tumor type. The best correlations across all tumor types were percentage of PDL1+ tumor cells (r=0.82 to 0.89), while cell densities of CTL and PD1+CTL showed the lowest correlations (r=0.35). Among tumor types, lung tumors and BC showed overall good correlation (r=0.82 to 0.83), depending on the marker. CRC showed the highest variability (average r=0.68), with correlation coefficients as low as 0.35 (PD1+CTL) and 0.46 (PDL1+macrophages). Computer selected ROI were not significantly more or less concordant with whole tumor analysis as compared to ROI selected by pathologists. Conclusion: Our results suggest that for immunoprofiling data, particularly proportion-based endpoints such as percent of PDL1+ tumor cells in lung and BC, ROI analysis has acceptable correlation with data from the whole tumor. However, CRC showed high variability, particularly in PD1+CTL, Ki67+CTL, and PDL1+macrophages, suggesting that CRC has higher tumor immunological heterogeneity compared with lung cancer and BC. These results suggest that investigators should analyze the whole tumor areas, otherwise, special attention should be paid to the image analysis strategy and its validation. Citation Format: Jaime Rodriguez-Canales, Michael Surace, Jennifer Cann. Tumor heterogeneity and its impact on immunoprofiling data: Whole tumor vs regions of interest (ROI) analysis of multiplex immunofluorescence in four carcinoma types [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 491.

The search for a sustainable land management has become a universal issue. It is especially necessary to discuss sustainable land management and to secure a site with enough feed supply to improve the lives of the farmers in the Ethiopian Highlands. This research studied the Adi Zaboy watershed in Tigray in order to reveal the changes in land management, assess how the different forms of land management affected the vegetation through unsupervised classification and normalized difference vegetation index (NDVI) analysis with geographic information system (GIS) 10.5 using a WorldView-2 satellite image taken in September 2016 and field investigation, and consider how to allow both environmental preservation and sustainable use of feed resources. The land management types at the research site were classified as &ldquo, seasonally-closed grazing land&rdquo, &ldquo, prohibited grazing and protected forest land&rdquo, and &ldquo, free grazing land&rdquo, On comparing the NDVI of each type of land management, it was found that the seasonally-closed grazing land makes it highly possible to secure and supply feed resources by limiting the grazing period. The expansion of the prohibited grazing and protected forest land is likely to tighten the restriction on the use of resources. Therefore, sustainable land management to secure feed resources may be possible by securing and actively using seasonally-closed grazing land, securing feed by a cut-and-carry, and using satellite images and GIS.

The aim of the present study was to determine the effect of oral administration of Lactobacillus plantarum OLL2712 (L. plantarum OLL2712) on glucose and lipid metabolism in mice with high-fat diet-induced obesity. Mice that had been administered 10(9) cfu heat-killed L. plantarum OLL2712 for 12 wk showed significant reduction of blood glucose levels in response to insulin. Furthermore, mRNA expression of interleukin-1β in adipose tissue and serum levels of nonesterified fatty acids in mice administered L. plantarum OLL2712 were significantly lower than those in control mice. These results indicate that L. plantarum OLL2712 regulates glucose metabolism.

Previous studies reported that exposure to dioxins was associated with an increased risk of various diseases in general populations. The aim of this study was to examine the association between levels of dioxins in blood and allergic and other diseases. We conducted a cross-sectional study on 1,063 men and 1,201 women (aged 15–76 years), who were living throughout Japan and not occupationally exposed to dioxins, during 2002–2010. In fasting blood samples, polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and dioxin-like PCBs (DL-PCBs) were analyzed by isotope dilution high-resolution gas chromatography/mass spectrometry. We obtained information on life style and self-reported history of diseases using a questionnaire. Blood pressure, blood levels of hemoglobin A1c, and serum lipids were also measured. Multiple logistic regression models were used to analyze the association between dioxin levels in blood and various diseases. Toxic equivalents of PCDDs/PCDFs and total dioxins showed significant inverse dose–response relationships with atopic dermatitis, after adjustments for potential confounders. The highest quartile for total dioxins had an adjusted odds ratio of 0.26 (95 % confidence interval 0.08–0.70) compared to the reference group (first quartile). The odds ratios for hypertension, diabetes mellitus, hyperlipidemia, gout in men, and gynecologic diseases in women significantly increased with increasing toxic equivalents of PCDDs/PCDFs, DL-PCBs, and total dioxins in blood. The present findings suggest that background exposure to dioxins was associated with reduced risk of atopic dermatitis. The results also support the idea that low-level exposure to dioxins is associated with an increased risk of diabetes, hypertension, and hyperlipidemia.

NC/Nga (NC) mice are an animal model for human atopic dermatitis. We found that induction of antigen (Ag)-specific T cell response is diminished in ovalbumin (OVA)- immunized NC mice. Ability of Ag presentation in NC mouse dendritic cells (DCs) was significantly weaker than that in BALB/c and DBA/2 mouse DCs. Expression levels of MHC class II molecules and cathepsin E in NC mouse DCs were significantly lower that those in BALB/c and DBA/2 mouse DCs. These results indicate that low expression levels of MHC class II and cathepsin E might contribute to the defect in induction of Ag-specific T cells in NC mice.

Buckwheat has been shown to have various health benefits such as reduction of hypertension and improvement of hypercholesterolemia; however, its effect on diabetes has not been fully elucidated. In this study, buckwheat bran extracts (BBE) inhibited sucrase activity in vitro more effectively than buckwheat. Balb/c mice pretreated with BBE showed dose-dependent reductions of blood glucose, greater than those observed with control mice, within 60 min following oral sucrose administration. Blood glucose levels in mice pretreated with buckwheat extracts were also significantly lower compared to those in control mice within 30 min following oral administration of sucrose. However, rutin, one of the abundant polyphenols of BBE, did not lower blood glucose level. Our data indicate that components of BBE other than rutin have inhibitory activity against sucrase in vivo. These results suggest that BBE could have beneficial effects on diabetes.

It has been shown that dietary nucleotides modulate immune response. Due to their unique properties in immune responses, nucleotides are used as immunonutrition in the field of clinical nutrition.In this study, we examined the effect of dietary deoxynucleic acid (DNA) on antigen (Ag)-specific immune response in ovalbumin (OVA)-immunized BALB/c mice and determined the mechanism using toll-like receptor 9 (TLR9) knock-out (KO) mice.BALB/c or TLR9 KO mice were fed control and 1% DNA diets and immunized with OVA. Spleen cells from OVA-immunized mice were stimulated with OVA in vitro, and the contents of IFN-γ and IL-4 in supernatants were measured by an ex vivo system. CD11c(+) dendritic cells were purified, and ability of cytokine induction to CD4(+) cells was examined.The level of OVA-specific IL-4 production in the DNA group was significantly higher than that in the control group. In contrast, the level of OVA-specific IFN-γ production in the DNA group was lower than that in the control group. The DNA diet decreased Ag-specific IL-4 production and enhanced Ag-specific IFN-γ production in TLR9 KO mice. CD11c(+) DCs from mice fed the DNA diet had a greater ability than CD11c(+) DCs from mice fed the control diet to induce the production of IL-4 from DO11.10 CD4(+) T cells.Dietary DNA increases Ag-specific IL-4 production and decreases IFN-γ production through a TLR9-dependent pathway. CD11c(+) dendritic cells are target cells in dietary DNA-induced immune regulation.

Hyperbaric air (HBA) is used to improve healing of wounds including diabetic ulcer. The aim of this study was to clarify the effects of HBA exposure on lipid and glu- cose metabolism in db/db mice. HBA did not influence the weight of db/db mice. Serum levels of free fatty acid and triglyceride, but not glucose and insulin, were significantly decreased after 6 weeks of treatment with HBA. The mRNA expressions of CPT-1, PPARα and PGC-1α genes, which are related to lipid metabolism, were significantly up-regulated in the muscle and liver. Increases in TNF α and MCP1 mRNA, which impaired lipid me- tabolism, were also attenuated by HBA treatment. These results suggest that exposure of HBA could have beneficial effects on lipid metabolism in patients with type 2 diabetes mellitus. J. Med. Invest. 57 : 224-231, August, 2010

Exendin-4 (Ex-4) is a glucagon-like peptide-1 receptor (GLP-1R) agonist that has been used as a drug injected subcutaneously for treatment of type 2 diabetes. Many studies have revealed molecular targets of Ex-4, but its influence on adipokines has not been determined. Our study showed that Ex-4 induced secretion of adiponectin into the culture medium of 3T3-L1 adipocytes. This effect of Ex-4 is due to increased adiponectin mRNA level through the GLP-1R. Both forskolin and 3-isobutyl-1-methylxanthine (IBMX), which may finally elevate cyclic adenosine monophosphate (cAMP) concentration, prevented the induction of adiponectin expression by Ex-4. Moreover, H89, a protein kinase A inhibitor, blocked the effect of Ex-4 on adiponectin. On the other hand, Ex-4 decreased the mRNA levels of inflammatory adipokines. The results indicate that Ex-4 directly promotes adiponectin secretion via the protein kinase A pathway in 3T3-L1 adipocytes and may ameliorate insulin resistance.

Problem statement: NC/Nga (NC) mice produced high levels of ovalbumin (OVA)- specific IgG, IgG1 and IgG2a. We previously found deletion polymorphisms in the promoter region of fcgr2b in NC mice. To determine whether this mutation causes a hyper-humoral immune response, we generated congenic BALB/c mice carrying the NC-type fcgr2b allele (NC fcgr2b) and analyzed humoral immune response and Fc γRIIB on germinal center (GC) B cells. Approach: BALB/c, NC and BALB/c- NC fcgr2b congenic mice were immunized with OVA 2 times at a 2-week-interval. Levels of OVA-specific IgG, IgG1 and IgG2a in serum were determined by ELISA. Four-color (anti- B220, anti-IgD, 2.4G2 and PNA) flow cytometry analysis was performed on splenocytes obtained from OVA-immunized mice and levels of Fc γRIIB in GC (IgD-PNA high ) and non-GC (IgD+PNA low ) B cells were analyzed. Results: Although perturbed up-regulation of Fc γRIIB on GC B was observed in congenic mice, levels of OVA-specific Abs were comparable to those in BALB/c mice. Conclusion: NC fcgr2b affects the level of Fc γRIIB in GC B cells but that the reduced Fc γRIIB expression is not related to enhanced Ag-specific A b responses in NC mice.

Although several studies have shown that the type and degree of unsaturation of fatty acids alter the production of proinflammatory mediators, there have been few studies in which the effects of trans isomers on allergic responses were evaluated. NC/Nga (NC) mice raised in conventional conditions develop spontaneous atopic dermatitis-like lesions with IgE hyperproduction. We used this model to evaluate the effect of dietary trans fatty acids (TFAs) on the development of spontaneous dermatitis. NC mice were fed a 4 g TFAs/kg diet (4 g TFA), a 8 g TFAs/kg diet (8 g TFA) or a control diet with no TFA content for 10 wk. The dermatitis condition improved with the increasing intake of TFAs, and there was a significant difference from week 5 of the experimental period between the dermatitis conditions in 8 g TFA-fed mice and control mice. Production of total IgE was also suppressed in mice fed TFAs compared with that in control mice, correlating with the skin lesions. IFN-gamma, but not IL-4, production was significantly greater in TFA-fed mice that in control mice. Our results suggest that intake of TFAs suppresses the development of atopic dermatitis-like lesions in NC mice. Even though we observed a decreased production of IgE levels, mechanisms involved in the development of dermatitis-like lesions seem to be IL-4-independent.

The amount of soy products consumed in Japan is much greater than that in Western countries. Recent evidence indicates that soy isoflavones play a beneficial role in obesity, cancer, osteoporosis, and cardiovascular disease. The soybean isoflavone genistein is present at high levels in soy products. Genistein is structurally similar to 17beta-estradiol (E2), and genistein has been suggested to be act as E2 or an antagonist against E2. Genistein suppresses antigen-specific immune response in vivo and lymphocyte proliferation response in vitro. However, genistein enhances the cytotoxic response mediated by NK and cytotoxic T cells and the cytokine production from T cells. Thus, the effect of genistein on immunity is immune cell-dependent. Due to its unique effect on immune function, genistein has been used for the treatment of the diseases in animal models and it has been found that genistein inhibits allergic inflammatory responses. In this review, we summarize current studies related to the effect of isoflavone genistein on the immune system.

The Long-Evans Cinnamon (LEC) rat has a spontaneous mutation, T helper immunodeficiency (thid), which causes a markedly reduced CD4(+) thymocyte population. Here we positionally clone the locus and identify a deletion in the gene encoding a receptor-like protein tyrosine phosphatase kappa (Ptprk) that led to complete loss of the transcript. The rat Ptprk gene exhibits 98% identity with the human and mouse counterparts and is expressed most abundantly in the CD4(-)CD8(-) double-negative stage. The downregulation of Ptprk in mouse immature thymocytes by RNA interference mimicked the thid phenotype. These results indicate that thid maps to the Ptprk locus and that functional Ptprk is crucial for lineage commitment or progression of CD4(+) T cells. We also found that Ptprk appears to function in parallel with or downstream of Th-POK/cKrox (also known as ZBTB7B), a master regulator of T cell lineage decision.

Oral tolerance is a phenomenon of induction of systemic unresponsiveness to antigens ingested by the oral route and loss of immune response. Studies have shown the importance of vitamin A in oral tolerance in vitro but not in an in vivo experimental model. Therefore, we carried out experiments to determine how vitamin A deficiency affects tolerance induction and the ability of mesenteric lymph node (MLN) CD11c(+) cells to induce regulatory T cells (Tregs). Immunological tolerance was induced by oral ovalbumin (OVA) administration in vitamin A-sufficient mice. OVA-specific antibody and cytokine production were significantly reduced. On the other hand, in vitamin A-deficient mice, both OVA-specific antibody and cytokine production were not suppressed by oral OVA administration. Regarding induction of Tregs, the conversion rate of Foxp3(+) cells from naive CD4(+) cell by CD11c(+) cells was decreased in vitamin A-deficient mice. Our study indicates that vitamin A deficiency causes the breakdown of oral tolerance in vivo.

We aimed to test the hypothesis that chemotherapy changes the gene expression of taste receptors in the tongue to induce dysgeusia in patients with head and neck cancer.Prospective observation study.We enrolled 21 patients who received chemoradiotherapy and five patients who underwent radiotherapy for head and neck cancer. The messenger RNA (mRNA) levels of the taste receptor subunits T1R1, T1R2, T1R3, and T2R5 were measured in lingual mucosa scrapings obtained with a small spatula. The perception thresholds of umami, sweet, and bitter tastes were assessed by the whole mouth gustatory test.In four patients with severe stomatitis induced by chemoradiotherapy, the mRNA levels of T1R1, T1R2, T1R3, and T2R5 in the lingual mucosa were significantly decreased. However, in 17 patients with mild/moderate stomatitis, the mRNA levels of T1R3 were significantly and transiently decreased, whereas those of T1R1 and T1R2 remained unchanged and those of T2R5 mRNA were significantly and transiently increased after chemotherapy. There was a significant negative correlation between the perception thresholds of umami or sweet tastes and lingual mRNA levels of T1R3 in patients with mild/moderate stomatitis after chemotherapy. Although the perception threshold of bitter taste remained unchanged, lingual mRNA levels of T2R5 were significantly increased in patients who complained of phantogeusia after chemotherapy.Chemotherapy specifically changed the gene expression of T1R3 and T2R5 in head and neck cancer patients with mild/moderate stomatitis, resulting in both dysgeusia of umami and sweet tastes as well as phantogeusia.4. Laryngoscope, 126:E103-E109, 2016.