Impact of Rice Bran Oil Emulsified Formulation on Digestion and Glycemic Response to Japonica Rice: An In Vitro Test and a Clinical Trial in Adult Men.

To assess the effect of rice bran oil emulsified formulation (EMF) on cooked rice, a single-arm open clinical trial and in vitro testing for digestion and glycemic response were performed. Fifteen Japanese men consumed 200 g of packed rice, cooked with or without EMF. Blood samples were collected 0, 30, 60, and 120 min post-consumption and analyzed for glucose, insulin, and triglyceride levels. Continuous glucose monitoring (CGM) and sensory evaluation were also performed. A two-step in vitro digestion test, simulating gastric and small intestinal digestion was conducted. EMF-added rice group showed higher insulin response levels at 60 min than the placebo group. Stratification of participants with HbA1c ≥ 5.6 or an insulinogenic index ≤ 0.4 revealed a significant reduction in Cmax glucose levels. A significant correlation was observed between venous and CGM blood glucose levels and no significant sensory differences were observed. The in vitro test revealed significantly lower C∞, equilibrium starch concentrations, with EMF. Clinical trial suggests that EMF may stimulate insulin secretion and reduce blood glucose levels in participants with lower insulin responses. In vitro tests suggest that EMF inhibits glycemic digestion. This trial was registered at the UMIN Center (UMIN000053495; registered 31 January 2024).

Assessing the Postprandial Glycemic Response to Japonica Rice (Oryza sativa L. cv. Koshihikari) with a Small Amount of Lysolecithin and Canola Oil in Japanese Adult Men: a Double-blind, Placebo-controlled, Crossover Study.

A double-blind, placebo-controlled, crossover trial was performed to analyze the effects of a small amount of lysolecithin and canola oil on blood glucose levels after consuming japonica rice. Overall, 17 Japanese adult men were assigned to consume 150 g of normally cooked japonica rice (placebo group) and 150 g of japonica rice cooked with 18 mg of lysolecithin and 1.8 g of canola oil (treatment group); these lipids were added as emulsified formulation (EMF) for stability and uniformity. Subsequently, blood samples were collected before and 30, 45, 60, 90, and 120 min after consuming test foods. There was no significant difference in blood glucose, insulin, and triglyceride levels between the groups. However, a stratified analysis of 11 subjects with body mass index (BMI) ≥ 22 revealed that blood glucose levels were significantly lower after 30 min in the treatment group than in the placebo group (p = 0.041). Through in vitro digestibility test, the rice sample of the treatment group was observed to release significantly less glucose within 20 min than that in the placebo group rice. These results suggest that the combination of a small amount of lysolecithin and canola oil modulated the increase in postprandial blood glucose levels induced by the intake of cooked japonica rice in adult men with BMI ≥ 22. This clinical trial was registered with the University Hospital Medical Information Network (UMIN) Center, (UMIN000045744; registered on 15/10/2021).

Effects of egg yolk choline intake on cognitive functions and plasma choline levels in healthy middle-aged and older Japanese: a randomized double-blinded placebo-controlled parallel-group study.

BACKGROUND: Choline, as a neurotransmitter acetylcholine precursor, is reportedly associated with cognitive function. Although there are several cohort and animal studies on choline-containing foods and cognitive function, only a few interventional studies were reported. Egg yolk is a rich source of different choline-containing chemical forms, such as phosphatidylcholine (PC), lysophosphatidylcholine (LPC), and α-glycerophosphocholine (α-GPC). This study aimed to investigate the effect of consuming 300 mg of egg yolk choline per day on cognitive function of Japanese adults. METHODS: A 12-week, randomized, double-blind, placebo-controlled, parallel-group study was conducted in 41 middle-aged and elderly males and females (43.9% female) aged ≥ 60 years and ≤ 80 years without dementia. Participants were randomly assigned to placebo and choline groups. The choline group received a supplement containing egg yolk choline (300 mg/day), and the placebo group received an egg yolk supplement free from choline for 12 weeks. Assessments of Cognitrax, Trail Making Tests (TMT) part A and B, the MOS 36-Item Short-Form Health Survey (SF-36), the Simplified Japanese Version of the WHO-Five Well-Being Index (WHO-5), and plasma choline levels were performed before and 6 and 12 weeks after supplement intake. In the present study, 19 subjects (9 in the placebo group and 10 in the choline group) were excluded due to the violation of the discontinuation criteria or participant compliance, and 41 subjects were analyzed. RESULTS: The change amount of verbal memory scores and verbal memory test-correct hit (delay) was significantly higher in the choline group than in the placebo group at baseline-6 and baseline-12 weeks. The plasma free choline level was significantly higher in the choline group compared with the placebo group at 6 weeks. Conversely, the choline group showed significantly lower Cognitrax processing speed scores, symbol digit coding testing correct responses, and SF-36 physical quality of life summary scores compared to the placebo group at 6 weeks. CONCLUSIONS: The results suggested that continued 300 mg/day intake of egg yolk choline improved verbal memory, which is a part of cognitive functions. To confirm the observed effects of egg yolk choline, more well-designed and large-scale studies are warranted. TRIAL REGISTRATION: Study protocols were pre-registered in the Clinical Trials Registration System (UMIN-CTR) (UMIN 000045050).

Connecting the dots in the associations between diet, obesity, cancer, and microRNAs.

The prevalence of obesity has reached pandemic levels worldwide, leading to a lower quality of life and higher health costs. Obesity is a major risk factor for noncommunicable diseases, including cancer, although obesity is one of the major preventable causes of cancer. Lifestyle factors, such as dietary quality and patterns, are also closely related to the onset and development of obesity and cancer. However, the mechanisms underlying the complex association between diet, obesity, and cancer remain unclear. In the past few decades, microRNAs (miRNAs), a class of small non-coding RNAs, have been demonstrated to play critical roles in biological processes such as cell differentiation, proliferation, and metabolism, highlighting their importance in disease development and suppression and as therapeutic targets. miRNA expression levels can be modulated by diet and are involved in cancer and obesity-related diseases. Circulating miRNAs can also mediate cell-to-cell communications. These multiple aspects of miRNAs present challenges in understanding and integrating their mechanism of action. Here, we introduce a general consideration of the associations between diet, obesity, and cancer and review the current knowledge of the molecular functions of miRNA in each context. A comprehensive understanding of the interplay between diet, obesity, and cancer could be valuable for the development of effective preventive and therapeutic strategies in future.

Non-amplification nucleic acid detection with thio-NAD cycling.

The PCR technique is indispensable in biology and medicine, but some difficulties are associated with its use, including false positive or false negative amplifications. To avoid these issues, a non-amplification nucleic acid detection protocol is needed. In the present study, we propose a method in which nucleic-acid probe hybridization is combined with thio-NAD cycling to detect nucleic acids without amplification. We report our application of this method for the detection of the gene of MPT64 in Mycobacterium tuberculosis. Two different cDNA probes targeted the mpt64 gene: the first probe was used to immobilize the mpt64 gene, and the second probe, linked with alkaline phosphatase (ALP), was hybridized to a target sequence in the mpt64 gene. A substrate was then hydrolyzed by ALP, and a cycling reaction was conducted by a dehydrogenase with its co-factors (thio-NAD and NADH). The single-stranded DNA, double-stranded DNA, plasmid DNA for the mpt64 gene, and whole genome of M. tuberculosis var. BCG were detected at the level of 105-106 copies/assay, whereas the non-tuberculosis mycobacteria (e.g., M. avium, M. intracellulare, M. kansasii, and M. abscessus) were below the limits of detection. The present method enables us to avoid the errors inherent in nucleic acid amplification methods.

Ultrasensitive ELISA Developed for Diagnosis.

For the diagnosis of disease, the ability to quantitatively detect trace amounts of the causal proteins from bacteria/viruses as biomarkers in patient specimens is highly desirable. Here we introduce a simple, rapid, and colorimetric assay as a de novo, ultrasensitive detection method. This ultrasensitive assay consists of a sandwich enzyme-linked immunosorbent assay (ELISA) and thionicotinamide-adenine dinucleotide (thio-NAD) cycling, forming an ultrasensitive ELISA, in which the signal substrate (i.e., thio-NADH) accumulates in a triangular manner, and the accumulated thio-NADH is measured at its maximum absorption wavelength of 405 nm. We have successfully achieved a limit of detection of ca. 10-18 moles/assay for a target protein. As an example of infectious disease detection, HIV-1 p24 could be measured at 0.0065 IU/assay (i.e., 10-18 moles/assay), and as a marker for a lifestyle-related disease, adiponectin could be detected at 2.3 × 10-19 moles/assay. In particular, despite the long-held belief that the trace amounts of adiponectin in urine can only be detected using a radioisotope, our ultrasensitive ELISA was able to detect urinary adiponectin. This method is highly versatile because simply changing the antibody enables the detection of various proteins. This assay system requires only the measurement of absorbance, thus it requires equipment that is easily obtained by medical facilities, which facilitates diagnosis in hospitals and clinics. Moreover, we describe an expansion of our ultrasensitive ELISA to a non-amplification nucleic acid detection method for nucleic acids using hybridization. These de novo methods will enable simple, rapid, and accurate diagnosis.

Activation of TRPV3 Regulates Inflammatory Actions of Human Epidermal Keratinocytes.

Transient receptor potential (TRP) ion channels were first characterized on neurons, where they are classically implicated in sensory functions; however, research in recent decades has shown that many of these channels are also expressed on nonneuronal cell types. Emerging findings have highlighted the role of TRP channels in the skin, where they have been shown to be important in numerous cutaneous functions. Of particular interest is TRPV3, which was first described on keratinocytes. Its functional importance was supported when its gain-of-function mutation was linked to Olmsted syndrome, which is characterized by palmoplantar keratoderma, periorifacial hyperkeratosis, diffuse hypotrichosis and alopecia, and itch. Despite these exciting results, we have no information about the role and functionality of TRPV3 on keratinocytes at the cellular level. In this study, we identified TRPV3 expression both on human skin and cultured epidermal keratinocytes. TRPV3 stimulation was found to function as a Ca2+-permeable ion channel that suppresses proliferation of epidermal keratinocytes and induces cell death. Stimulation of the channel also triggers a strong proinflammatory response via the NF-κB pathway. Collectively, our data show that TRPV3 is functionally expressed on human epidermal keratinocytes and that it plays a role in cutaneous inflammatory processes.

Combination therapy with ONO-KK1-300-01, a cathepsin K inhibitor, and parathyroid hormone results in additive beneficial effect on bone mineral density in ovariectomized rats.

This study examined the effects of a novel cathepsin K inhibitor, ONO-KK1-300-01 (KK1-300), used concurrently with parathyroid hormone (PTH) in ovariectomized (OVX) rats. KK1-300 (3 mg/kg, twice daily), alendronate (1 mg/kg, once daily) or vehicle were orally administered to OVX rats for 56 days, starting the day after ovariectomy, followed by combination treatment with or without PTH (3 µg/kg, subcutaneously three times a week) for another 28 days. OVX control animals exhibited a significant increase in both bone resorption (urinary deoxypyridinoline; DPD) and formation markers (serum osteocalcin) as well as microstructural changes associated with decreased bone mineral density (BMD). Combination treatment with KK1-300 and PTH significantly decreased urinary DPD and increased serum osteocalcin, indicating a sustained beneficial effect compared to the effect of each mono-therapy. On the other hand, combination therapy with alendronate and PTH weakened the PTH-induced increase in osteocalcin. In proximal tibia, combination treatment with KK1-300 and PTH increased BMD to a level significantly higher than that achieved following single treatment with KK1-300 or PTH alone. On the other hand, combination treatment with alendronate and PTH failed to produce any significant additive effect on BMD following single treatment with alendronate or PTH alone. Microstructural analysis revealed that the PTH-induced increase in bone formation (MS/BS and BFR/BS) was fully maintained following combination treatment with KK1-300 and PTH, but not following combination treatment with alendronate and PTH. These findings indicate that KK1-300, unlike alendronate, has an additive effect on the preventive action of PTH on bone loss in OVX rats.

Effect of intermittent and daily regimens of minodronic Acid on bone metabolism in an ovariectomized rat model of osteoporosis.

The goal of the study was to compare the effects of minodronic acid on bone mineral density (BMD) and bone turnover in a rat ovariectomized (OVX) osteoporosis model, using two intermittent treatment regimens (weekly and 4 continuous days every 4 weeks) and a daily regimen. Female F344 rats (age 14 weeks) underwent ovariectomy or a sham operation. Minodronic acid was orally administered at 0.042, 0.21, and 1.05 mg/kg in the intermittent regimens, and at 0.03 and 0.15 mg/kg in the daily regimen for 12 weeks from the day after surgery. Minodronic acid dose-dependently ameliorated the decreases in areal BMD of the lumbar vertebrae and femur, and volumetric BMD of total and trabecular bone in the distal femur. Minodronic acid also suppressed the increase in urinary deoxypyridinoline levels and reduced serum osteocalcin levels. In bone histomorphometry, all three minodronic acid regimens suppressed OVX-induced increases in bone turnover at the tissue level and ameliorated all structural indices, except that an effect on trabecular thickness only occurred with daily treatment. In conclusion, minodronic acid administered weekly or for 4 continuous days every 4 weeks suppressed increased bone resorption and BMD to a similar extent to that of a similar total dose given daily in a rat OVX model.

Effects of eight-month treatment with ONO-5334, a cathepsin K inhibitor, on bone metabolism, strength and microstructure in ovariectomized cynomolgus monkeys.

This study examined the effect of ONO-5334, a cathepsin K inhibitor, on bone turnover, mineral density (BMD), mechanical strength and microstructure in ovariectomized (OVX) cynomolgus monkeys. Vehicle, ONO-5334 (3, 10 or 30 mg/kg) or alendronate (0.5 mg/kg) was orally administered for eight months to sham- and OVX-operated monkeys. ONO-5334 dose-dependently suppressed OVX-induced increase in bone turnover markers (urinary C-terminal cross-linking telopeptide of type I collagen (CTX) and serum osteocalcin). At the dose of 30 mg/kg, ONO-5334 maintained urinary CTX at nearly zero level and kept serum osteocalcin around the level of the sham animals. Marker levels in the alendronate-treated animals were similar to those in the sham animals throughout the study. ONO-5334 dose-dependently reversed the effect of OVX on vertebral BMD as measured by dual-energy X-ray absorptiometry (DXA) with improvement of bone mechanical strength. Both ONO-5334 and alendronate suppressed OVX-induced changes in vertebral microstructure and turnover state. In the femoral neck, peripheral quantitative computed tomography (pQCT) analysis showed that ONO-5334 increased total and cortical BMD. In particular, ONO-5334 significantly increased cortical BMD with improvement of bone mechanical strength. In microstructural analysis, alendronate suppressed OVX-induced increase in femoral mid-shaft osteonal bone formation rate (BFR) to a level below that recorded in the sham group, whereas ONO-5334 at 30 mg/kg did not suppress periosteal, osteonal and endocortical BFR. This finding supports the significant effect of ONO-5334 on cortical BMD and mechanical strength in the femoral neck. The results of this study suggest that ONO-5334 has good therapeutic potential for the treatment of osteoporosis.

ONO-5334, a cathepsin K inhibitor, improves bone strength by preferentially increasing cortical bone mass in ovariectomized rats.

This study compared the effects of ONO-5334, a cathepsin K inhibitor, with those of alendronate on bone mass and strength in ovariectomized rats. Ovariectomy resulted in significant elevation in urinary deoxypyridinoline and plasma C-terminal cross-linking telopeptide of type I collagen (CTX) 8 weeks after surgery. Peripheral quantitative computed tomography analysis showed that total, trabecular, and cortical bone mineral content (BMC) decreased in the proximal tibia, which was paralleled with a significant decline in bone strength. Treatment with ONO-5334 (0.12, 0.6, 3 or 15 mg/kg) once daily for 8 weeks dose-dependently restored the decrease in total BMC and bone mineral density (BMD) in the proximal tibia and suppressed urinary deoxypyridinoline and plasma CTX levels. Alendronate (1 mg/kg, once daily) also fully restored these bone mass parameters. Separate analysis of trabecular and cortical bones, however, showed that ONO-5334 only partially restored trabecular BMD and BMC at 15 mg/kg, whereas alendronate fully restored these parameters. On the other hand, ONO-5334 increased both cortical BMD and BMC with an effect more potent than that of alendronate. Bone geometric analysis indicated that ONO-5334 at 15 mg/kg decreased endosteal circumference without affecting periosteal circumference, resulting in marked increase in cortical thickness. Interestingly, the effects of ONO-5334 on bone strength parameters were more prominent than those of alendronate, although the two test compounds had a similar effect on total BMC. Taken together, our results indicate that ONO-5334 has pharmacological characteristics different from those of alendronate and may offer a unique therapy for patients with osteoporosis.

Effects of ONO-5334, a novel orally-active inhibitor of cathepsin K, on bone metabolism.

In the present study, we examined the in vitro and in vivo pharmacological effects of ONO-5334, a novel inhibitor of cathepsin K, on bone metabolism. In vitro experiments indicated that ONO-5334 is a potent inhibitor of cathepsin K with Ki value of 0.1 nM. Although this compound inhibited other cysteine proteases, such as cathepsin S, L and B, its inhibitory activity for these enzymes was 8 to 320 fold lower than that for cathepsin K. ONO-5334 also inhibited human osteoclasts bone resorption in vitro at a concentration more than 100 fold lower than that of alendronate, a bisphosphonate. While alendronate disrupted actin ring and induced pyknotic nuclei in osteoclasts, ONO-5334 did not have such effects, suggesting that this compound does not affect osteoclasts viability. In in vivo experiments, oral administration of ONO-5334 dose-dependently reduced plasma calcium level increased by parathyroid hormone related peptide in thyroparathyroidectomized rats. Furthermore, in vivo experiment using normal monkeys demonstrated that ONO-5334 decreases serum and urine C-telopeptide of type I collagen level, a bone resorption marker, soon after oral dosing. These levels were consistently decreased below pre-dose levels by repeated oral dosing with ONO-5334 for 7 days. ONO-5334 on the other hand did not affect bone formation markers, serum osteocalcin and bone specific alkaline phosphatase. These findings indicate that ONO-5334 is a specific inhibitor for cathepsin K and thus may be a novel therapeutic agent for metabolic bone diseases.

Long-term minodronic acid (ONO-5920/YM529) treatment suppresses increased bone turnover, plus prevents reduction in bone mass and bone strength in ovariectomized rats with established osteopenia.

The present study examined the effect of the highly potent nitrogen-containing bisphosphonate, minodronic acid (ONO-5920/YM529), on bone mineral density (BMD), bone turnover, bone histomorphometry and bone strength in ovariectomized (OVX) rats. Female F344/DuCrj rats, aged 14 weeks, were OVX or sham operated. After 3 months, the OVX rats showed an increase in bone turnover, and a decrease in bone mass and bone strength. Minodronic acid was administered orally once a day for 12 months at doses of 0, 0.006, 0.03 and 0.15 mg/kg from 3 months after OVX. Minodronic acid dose-dependently inhibited the decrease in BMD of lumbar vertebrae and femur. In the femur, treatment with 0.15 mg/kg minodronic acid increased the BMD of distal and mid sites to sham levels. Minodronic acid dose-dependently suppressed OVX-induced increase in urinary deoxypyridinoline, a bone resorption marker, after a month of treatment and these effects were maintained for 12 months of treatment. Minodronic acid also decreased serum osteocalcin, a bone formation marker. In bone histomorphometric analysis after 12 months of treatment, OVX rats showed an increase in bone resorption (Oc.S/BS and N.Oc/BS) and bone formation (MS/BS and BFR/BV) at lumbar vertebral bodies. Minodronic acid suppressed the OVX-induced increase in bone turnover at tissue level. Trabecular bone volume, trabecular thickness and trabecular number of lumbar vertebral bodies were decreased after OVX. Minodronic acid increased these structural indices, indicating that it prevented the deterioration in trabecular architecture. In a mechanical test at 12 months of treatment, ultimate load of lumbar vertebral bodies and mid femur in the OVX-control group was decreased compared to the sham group. Minodronic acid prevented the reduction in bone strength at both sites. In particular, in the mid femur, treatment with 0.03 and 0.15 mg/kg minodronic acid increased bone strength to sham levels or greater. In conclusion, minodronic acid suppressed increased bone turnover, plus prevented the decrease in BMD, deterioration of bone microarchitecture and reduction in bone strength in OVX rats with established osteopenia. These results suggest that minodronic acid may be clinically useful for treatment of osteoporosis.

Production of optically active 1,2,4-butanetriol from corresponding racemate by Microbial stereoinversion.

Sterigmatomyces elviae DSM 70852 produced 12 g/l (S)-1,2,4-butanetriol (enantiomeric excess >99.9%) from 20 g/l racemate in 82 h. From the results of the inversion of an (R)-isomer to an (S)-isomer and GC-MS, it was suggested that (R)-1,2,4-butanetriol is oxidized to 1,4-dihydroxy-2-butanone, which is reduced to an (S)-isomer.

Effect of antiallergic drugs on interleukin 5-induced eosinophil infiltration of rat airways.

Interleukin (IL)-5 is thought to play important roles in asthma and to be a potential therapeutic target. An intratracheal injection of murine recombinant IL-5 (3-30 microg/animal) induced a dose-dependent increase in the number of eosinophils in the bronchoalveolar lavage fluid of Brown Norway (BN) rats 24 h after administration. Bovine serum albumin (30pg/animal), used as reference material, did not cause any change. The reaction was not observed in F344 rats. The increase in the number of eosinophils did not accompany bronchial hyperreactivity in BN or F344 rats. Prednisolone (3-10 mg/kg, i.p.) and emedastine (30 mg/kg, p.o.) reduced the increased number of eosinophils induced by the IL-5 challenge. These results suggest that IL-5 is a potent inducer of eosinophils in the airway of BN rats. Prednisolone and emedastine are effective against IL-5-induced eosinophilia.