Return to Sports Activity After Microfracture for Osteochondral Lesion of the Talus in Skeletally Immature Children.

BACKGROUND: Osteochondral lesions of the talus (OLTs) are among the common sports-related injuries. However, there are few reports on the return to sports after OLT surgery in skeletally immature children. This study was performed to evaluate the return to sports after microfracture for OLTs in skeletally immature children. METHODS: This study involved 17 ankles of 16 patients (mean age, 13.2 years; range, 10-16 years) with open tibial epiphyses on magnetic resonance imaging (MRI) who underwent microfracture for OLTs <10 mm in diameter and confirmation of lesion instability under arthroscopy. Nine of 17 ankles had additional lateral ankle ligament stabilization. All patients were participating in some form of sports. The Japanese Society for Surgery of the Foot (JSSF) score, Ankle Activity Score (AAS), return to sports rate, lesion size, grade of subchondral bone marrow edema, and Magnetic Resonance Observation of Cartilage Repair Tissue (MOCART) score on MRI were evaluated after microfracture. The mean postoperative follow-up period was 53.5 months. RESULTS: The mean JSSF score significantly improved from 76.1 points preoperatively to 94.9 points at the final follow-up (P < .01). The mean AAS showed no change from preoperative state to final follow-up. The return to sports rate was 100%. The lesion size significantly decreased from 76.3 to 56.7 mm2 in area (P = .02) and from 283.2 to 185.6 mm3 in volume (P = .05). The bone marrow edema grade decreased in 8 of 17 ankles. The total MOCART score showed a significant improvement from 6 months to 1 year postoperatively (P = .05). CONCLUSION: All skeletally immature children who underwent microfracture for OLTs in this study were able to return to sports activity and showed improvements in clinical scores and MRI parameters. Microfracture may be considered an effective first-line treatment for OLTs <10 mm in diameter in skeletally immature athletes. LEVEL OF EVIDENCE: Level IV, retrospective case-control study.

Natural variation of diterpenoid phytoalexins in rice: Aromatic diterpenoid phytoalexins in specific cultivars.

Rice (Oryza sativa L.) plants accumulate antimicrobial compounds known as phytoalexins in response to pathogen attack. To date, more than 20 compounds have been isolated as phytoalexins from rice, mostly diterpenoids. However, the quantitative analysis of diterpenoid phytoalexins in various cultivars has revealed that the cultivar 'Jinguoyin' does not accumulate these compounds at detectable concentrations. Therefore, in this study, we attempted to detect a new class of phytoalexins from Bipolaris oryzae infected leaves of 'Jinguoyin'. We detected five compounds in the leaves of the target cultivar, whereas these compounds were not detected in the leaves of 'Nipponbare' or 'Kasalath', which are representative cultivars of the japonica and indica subspecies. Subsequently, we isolated these compounds from ultraviolet (UV)-light-irradiated leaves and determined their structures by spectroscopic analysis and the crystalline sponge method. All the compounds were diterpenoids containing a benzene ring and were detected from the pathogen-infected rice leaves for the first time. Because the compounds showed antifungal activity against B. oryzae and Pyricularia oryzae, we propose that they function as phytoalexins in rice and named them abietoryzins A-E. The abietoryzins tended to accumulate at high concentrations in cultivars that accumulated low levels of known diterpenoid phytoalexins after UV-light irradiation. Of the total of 69 cultivars in the WRC, 30 cultivars accumulated at least one of the abietoryzins, and, in 15 cultivars, the amounts of some abietoryzins were the highest among those of the analyzed phytoalexins. Therefore, abietoryzins are a major phytoalexin group in rice, although their presence has, to date, been overlooked.

Comprehensive Structural Analysis of the Bitter Components in Beer by the HPLC-Assisted Crystalline Sponge Method.

Trans-iso-α-acid is one of the main contributors to the bitter taste of fresh beer and is known to transform into various derivatives during beer aging. However, structural characterization of the derivatives has been a challenging task because of the formation of too many components. Herein, we report that most of the transformation products of trans-iso-α-acid, isolated in this study in only small quantities by HPLC, can be structurally analyzed with the crystalline sponge method. Thirteen compounds, including eight that were previously unreported, have been successfully isolated and analyzed with complete assignment of their absolute configuration. This provides an improved understanding of the chemical transformations that occur during beer aging.

Correction: Crystalline sponge X-ray analysis coupled with supercritical fluid chromatography: a novel analytical platform for the rapid separation, isolation, and characterization of analytes.

Correction for 'Crystalline sponge X-ray analysis coupled with supercritical fluid chromatography: a novel analytical platform for the rapid separation, isolation, and characterization of analytes' by Yoshimasa Taniguchi et al., Analyst, 2021, 146, 5230-5235, DOI: 10.1039/D1AN00948F.

Crystalline sponge X-ray analysis coupled with supercritical fluid chromatography: a novel analytical platform for the rapid separation, isolation, and characterization of analytes.

Crystalline sponge (CS) based X-ray diffraction (XRD) analysis allows for the observation of the structure of an analyte, including its absolute configuration. Herein we report a powerful analytical platform for the separation, isolation, and structural elucidation of a target analyte in a seamless way by coupling supercritical fluid chromatography (SFC) with CS-based XRD analysis (SFC-CSXRD). The efficacy of this methodology is demonstrated by the rapid characterization of regio- and stereoisomers using three types of CSs with differing tolerances to the solvents used in SFC and guest-soaking.

Bitter taste receptor activation by hop-derived bitter components induces gastrointestinal hormone production in enteroendocrine cells.

Matured hop bitter acids (MHBA) are bitter acid oxides derived from hops, widely consumed as food ingredients to add bitterness and flavor in beers. Previous studies have suggested a potential gut-brain mechanism in which MHBA simulates enteroendocrine cells to produce cholecystokinin (CCK), a gastrointestinal hormone which activates autonomic nerves, resulting in body fat reduction and cognitive improvement; however, the MHBA recognition site on enteroendocrine cells has not been fully elucidated. In this study, we report that MHBA is recognized by specific human and mouse bitter taste receptors (human TAS2R1, 8, 10 and mouse Tas2r119, 130, 105) using a heterologous receptor expression system in human embryonic kidney 293T cells. In addition, knockdown of each of these receptors using siRNA transfection partially but significantly suppressed an MHBA-induced calcium response and CCK production in enteroendocrine cells. Furthermore, blocking one of the essential taste signaling components, transient receptor potential cation channel subfamily M member 5, remarkably inhibited the MHBA-induced calcium response and CCK production in enteroendocrine cells. Our results demonstrate that specific bitter taste receptor activation by MHBA drives downstream calcium response and CCK production in enteroendocrine cells. These findings reveal a mechanism by which food ingredients derived from hops in beer activate the gut-brain axis for the first time.

Hop bitter acids containing a ß-carbonyl moiety prevent inflammation-induced cognitive decline via the vagus nerve and noradrenergic system.

The prevention of age-related cognitive decline and dementia is becoming a high priority because of the rapid growth of aging populations. We have previously shown that hop bitter acids such as iso-α-acids (IAAs) and matured hop bitter acids (MHBAs) activate the vagus nerve and improve memory impairment. Moreover, supplements with MHBAs were shown to improve memory retrieval in older adults. However, the underlying mechanisms have not been entirely elucidated. We aimed to investigate the effects of MHBAs and the common ß-tricarbonyl moiety on memory impairment induced by the activation of microglia and the loss of the noradrenergic system. MHBAs and a model compound with ß-tricarbonyl moiety were administered to LPS-inoculated mice and 5 × FAD Alzheimer's disease (AD) model mice, following the evaluation in behavioral tests and microglial activation. To evaluate the association of noradrenaline with MHBAs effects, mice treated with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4), a noradrenergic neurotoxin that selectively damages noradrenergic projections from the locus coeruleus, were subjected to the behavioral evaluation. MHBAs reduced brain inflammation and improved LPS-induced memory impairment. A model compound possessing the ß-tricarbonyl moiety improved the LPS-induced memory impairment and neuronal loss via the vagus nerve. Additionally, the protective effects of MHBAs on memory impairment were attenuated by noradrenaline depletion using DSP-4. MHBAs suppressed the activation of microglia and improved the memory impairment in 5 × FAD mice, which was also attenuated by noradrenaline depletion. Treatment with MHBAs increased cholecystokinin production from the intestinal cells. Generally, cholecystokinin activates the vagal nerve, which stimulate the noradrenergic neuron in the locus ceruleus. Taken together, our results reveal that food ingredients such as hop bitter acids with a ß-tricarbonyl moiety suppress microglial activation and improve memory impairment induced by inflammation or AD pathology via the activation of the gut-brain axis and noradrenergic system. Supplements with hop bitter acids, including MHBAs, might be a novel approach for the prevention of cognitive decline and dementia.

Exploiting the Potential of Meroterpenoid Cyclases to Expand the Chemical Space of Fungal Meroterpenoids.

Fungal meroterpenoids are a diverse group of hybrid natural products with impressive structural complexity and high potential as drug candidates. In this work, we evaluate the promiscuity of the early structure diversity-generating step in fungal meroterpenoid biosynthetic pathways: the multibond-forming polyene cyclizations catalyzed by the yet poorly understood family of fungal meroterpenoid cyclases. In total, 12 unnatural meroterpenoids were accessed chemoenzymatically using synthetic substrates. Their complex structures were determined by 2D NMR studies as well as crystalline-sponge-based X-ray diffraction analyses. The results obtained revealed a high degree of enzyme promiscuity and experimental results which together with quantum chemical calculations provided a deeper insight into the catalytic activity of this new family of non-canonical, terpene cyclases. The knowledge obtained paves the way to design and engineer artificial pathways towards second generation meroterpenoids with valuable bioactivities based on combinatorial biosynthetic strategies.

An Expansion of Crystalline Sponge X-ray Analysis to Elucidate the Molecular Structure of Reactive Compounds via Ion Pair Formation.

The crystalline sponge (CS) method allows structural elucidation of a target compound (guest) in solution by single crystal X-ray diffraction through trapping the guest into the CS framework. In principle, the CS method is inapplicable to reactive compounds that break the CS framework, such as acidic, basic, or nucleophilic ones. Here, a solution to this problem is disclosed wherein an ion pair of the guest compound is formed during the guest-soaking step by adding a suitable reagent. The ion pair can be observed and does not damage the CS framework. Using the developed method, amino, guanidino, and amidino compounds have been successfully analyzed as ion pairs with sulfonic acids. Practical utility has been shown because the absolute configurations of optically resolved amine derivatives were revealed with only a few micrograms. This demonstrates that the ion-pair-soaking method is simple and expands the range of compounds applicable to the CS method.

Colletofurans A-E, 1-Octyl-1,3-dihydroisobenzofuran Derivatives from Colletotrichum boninense AM-12-2.

3-(3-Hydroxy-2-(hydroxymethyl)phenyl)propanoic acid, 2-hydroxymethyl-3-hydroxy-(E)-cinnamic acid, and colletofurans A-E were isolated from Colletotrichum boninense AM-12-2. Colletofurans A-E are the first natural compounds featuring a 1-octyl-1,3-dihydroisobenzofuran core. Their structures were initially established by 1D/2D-NMR and HRESITOFMS. Mosher's ester method was used to determine the absolute configurations of secondary alcohols in colletofurans A-C. The structures of colletofurans A and B-E were further confirmed by DFT GIAO calculations and the X-ray crystalline sponge method, respectively.

Exploiting a C-N Bond Forming Cytochrome†P450 Monooxygenase for C-S Bond Formation.

C-S bond formation reactions are widely distributed in the biosynthesis of biologically active molecules, and thus have received much attention over the past decades. Herein, we report intramolecular C-S bond formation by a P450 monooxygenase, TleB, which normally catalyzes a C-N bond formation in teleocidin biosynthesis. Based on the proposed reaction mechanism of TleB, a thiol-substituted substrate analogue was synthesized and tested in the enzyme reaction, which afforded the unprecedented sulfur-containing thio-indolactam V, in addition to an unusual indole-fused 6/5/8-tricyclic product whose structure was determined by the crystalline sponge method. Interestingly, conformational analysis revealed that the SOFA conformation is stable in thio-indolactam V, in sharp contrast to the major TWIST form in indolactam V, resulting in differences in their biological activities.

Biomimetic Synthesis of Meroterpenoids by Dearomatization-Driven Polycyclization.

A biomimetic route to farnesyl pyrophosphate and dimethyl orsellinic acid (DMOA)-derived meroterpenoid scaffolds has yet to be reported despite great interest from the chemistry and biomedical research communities. A concise synthetic route with the potential to access DMOA-derived meroterpenoids is highly desirable to create a library of related compounds. Herein, we report novel dearomatization methodology followed by polyene cyclization to access DMOA-derived meroterpenoid frameworks in six steps from commercially available starting materials. Furthermore, several farnesyl alkene substrates were used to generate structurally novel, DMOA-derived meroterpenoid derivatives. DFT calculations combined with experimentation provided a rationale for the observed thermodynamic distribution of polycyclization products.

Roasted Barley Extract (Mugicha) Containing Cyclo(D-Phe-L-Pro) Prevents a Decrease in Skin Temperature in Cold Conditions: A Randomized, Double-Blind, Placebo-Controlled, Crossover Study.

Roasted barley extract (RBE) is a traditional Japanese beverage. Previously, we reported the effects of RBE containing cyclo(d-Phe-l-Pro) on blood flow in animals and humans and investigated rapid skin temperature recovery from cold-water immersion in women. The present randomized, double-blind study investigated the effects of RBE containing cyclo(d-Phe-l-Pro) on men's and women's skin temperature in excessively air-cooled conditions. Participants felt cold in the test room (25.5±0.5ºC). They ingested an RBE or placebo beverage and remained in the air-conditioned room for 100 min. Skin temperature of the left foot was measured every 5 min using infrared thermography. We evaluated effect of RBE administration by paired t-test. The skin temperature of the RBE group remained higher than that of the placebo group. The skin temperature changes 100 min after RBE or placebo ingestion were -3.67±1.14ºC and -4.59±0.89ºC, respectively in all participants. We also did subclass analysis focusing on men or women. In a previous study, RBE efficacy for skin temperature in men was not clearly demonstrated. RBE consumption was also effective not only in female participants but also in male participants. The skin temperature changes 100 min after RBE or placebo ingestion were -3.65±0.64ºC and -4.55±0.32ºC, respectively in male participants. Therefore, RBE containing cyclo(d-Phe-l-Pro) prevented skin temperature decreases in excessively air-cooled conditions in both men and women.

Theaflavins Improve Memory Impairment and Depression-Like Behavior by Regulating Microglial Activation.

Inflammation in the brain is associated with various disorders including Alzheimer's disease and depression. Thus, inflammation has received increasing attention regarding preventive approaches to such disorders. Epidemiological investigations have reported that drinking tea reduces the risk of dementia and depression. Theaflavins, a polyphenol found in black tea, are known to have anti-oxidative and anti-inflammation effects, but the effects of theaflavins on cognitive decline and depression induced by inflammation have not been investigated. To address this research gap, the present study assessed whether theaflavins could protect synapses and dendrites damaged by inflammation and prevent concomitant memory impairment and depression-like behavior in mice. Intracerebroventricular injection with lipopolysaccharide (LPS) induces neural inflammation associated with reduced spontaneous alternations in the Y-maze test and increased immobility in the tail suspension test, indicating impaired spatial memory and depression-like behavior, respectively. Oral administration with theaflavins prevented these behavioral changes induced by LPS. Theaflavins also suppressed productions of inflammatory cytokines and prevented dendritic atrophy and spine loss in the brain. Notably, theaflavins have a stronger anti-inflammatory effect than other polyphenols such as catechin, chlorogenic acid, and caffeic acid. These results suggest that theaflavins can suppress neural inflammation and prevent the symptoms of inflammation-related brain disorders.

Secretion of a gastrointestinal hormone, cholecystokinin, by hop-derived bitter components activates sympathetic nerves in brown adipose tissue.

Matured hop bitter acids (MHBA) are oxidation products from bitter components in hops, which are used widely as food materials to add flavor and bitterness in beer production. Our previous study has shown that MHBA induces thermogenesis in brown adipose tissue (BAT) via sympathetic nerves in rodents and reduces body fat in healthy adults. However, it is unclear how MHBA affects the sympathetic nervous system. In this study, we demonstrate that MHBA treatment of enteroendocrine cells increases Ca2+ levels and induces the secretion of the gastrointestinal hormone, cholecystokinin (CCK), in a dose-dependent manner. These effects were eliminated by Ca2+ depletion from the medium or blockers of L-type voltage-sensitive Ca2+ channels during pretreatment. Induction of CCK secretion by MHBA was also confirmed using isolated rat small intestines. Elevation of the sympathetic nerve activity innervating BAT (BAT-SNA) and BAT temperature by MHBA administration in rats was blocked by pretreatment with a CCK receptor 1 (CCK1R) antagonist. Moreover, the intraperitoneal injection of CCK fragment elevated BAT-SNA, and this increase was blocked by subdiaphragmatic vagotomy. These results demonstrate that MHBA induces CCK secretion in the gastrointestinal tracts and elevates BAT-SNA via CCK1R and vagal afferent nerves. In addition, MHBA increases BAT temperature via CCK1R. Our findings reveal a novel mechanism of the beneficial metabolic effects of food ingredients.

Matured Hop-Derived Bitter Components in Beer Improve Hippocampus-Dependent Memory Through Activation of the Vagus Nerve.

Improving and maintaining memory function is effective in preventing cognitive decline and dementia. Previously, we demonstrated that iso-α-acids, the hop-derived bitter components in beer, prevent cognitive impairment in an Alzheimer's disease mouse model. In this report, we investigated the effects of matured hop bitter acids (MHBA) containing components of oxides derived from α- and ß-acids, and structurally similar to iso-α-acids, on cognitive function using behavioral pharmacological procedures. MHBA and the representative components of MHBA, 4'-hydroxyallohumulinone (HAH) and 4'-hydroxy-cis-alloisohumulone (HAIH) improved spatial working memory in scopolamine-induced amnesia mice. MHBA also enhanced episodic memory in the novel object recognition test (NORT). The administration of MHBA increased the amount of norepinephrine (NE) and NE release into cerebrospinal fluid (CSF) in hippocampus. The MHBA activity in improving memory function was attenuated by treatment with a ß-adrenergic receptor inhibitor. In addition, vagotomized mice did not display the memory improvement induced by MHBA. Together, our results suggest that MHBA improves memory function via stimulation of the vagus nerve and enhancement of NE release in the hippocampus. Vagus nerve activation by the intake of food materials including MHBA may be a safe and effective approach for improving cognitive function.

Genetic, acute and subchronic toxicity studies of matured hop extract produced by extraction from heat-treated hops.

It has been demonstrated that successive ingestion of matured hop extract (MHE), produced by extraction from heat-treated hops, results in body fat reduction in animals and humans; however, preclinical safety studies have not been reported. In this study, we conducted in vitro and in vivo safety studies for MHE. Genotoxicity was evaluated using the Ames test, in vitro chromosomal aberration test, and in vivo micronucleus test. To assess acute safety, a single, oral administration of MHE to rats was monitored. Subchronic safety was assessed by repeated feeding with MHE for 90 days. The in vitro chromosomal aberration test was positive at 3,330 µg/mL and 5,000 µg/mL without metabolic activation. However, MHE did not induce any reverse mutation with or without metabolic activation in the Ames test, and no abnormalities were observed at a dose of 2,000 mg/kg body weight in the rat micronucleus test. In the acute and subchronic safety studies, no deaths or toxicological signs were recorded during the observation period. In addition, no changes in body weights, feed/water consumption, clinical signs, ophthalmoscopy, urinalysis, hematology, blood biochemistry, organ weights, or histopathology were observed after repeated administration of MHE. Therefore, the no-observed-adverse-effect-level (NOAEL) of MHE was considered to be over 3,484 and 4,022 mg/kg body weight/day in males and females, respectively. These results indicate that there is no safety concern for MHE in the present preclinical safety study.

Roasted Barley Extract (Mugi-cha) Containing Cyclo(d-Phe-l-Pro) Prevents Lowering of the Cutaneous Blood Flow and Skin Temperature under Air Conditioning: A Randomized, Double-Blind, Placebo-Controlled, Crossover Study.

Roasted barley extract (RBE), also known as mugi-cha, is a well-known healthy non-caffeinated beverage, and its health functionality has been widely reported. Our previous clinical study showed that RBE affects the cutaneous blood flow and skin temperature after cold-water immersion and that cyclo(d-Phe-l-Pro) is responsible for its effect. In this study, we investigated whether cyclo(d-Phe-l-Pro)-containing RBE prevents the decrease in the cutaneous blood flow and skin temperature. Subjects remained in the air-conditioned room while ingesting RBE or a placebo. We measured the cutaneous blood flow and skin temperature. We evaluated the effect of RBE administration by two-way repeated measures analysis of variance. A total of 15 subjects were enrolled. The change in cutaneous blood flow in the RBE and placebo groups was -0.79 ± 0.38 and -2.03 ± 0.35 mL min-1 100 g-1, respectively ( p value of 0.041). The change in the skin temperature in the RBE and placebo groups was -1.85 ± 0.35 and -3.02 ± 0.30 °C, respectively ( p value of <0.001). We also did subclass analysis with cold-feeling subjects. For the seven subjects who had cold sensation, the change in the cutaneous blood flow in the RBE and placebo groups was -0.48 ± 0.58 and -2.56 ± 0.48 mL min-1 100 g-1, respectively ( p value of 0.008). The change in the skin temperature in the RBE and placebo groups was -1.46 ± 0.74 and -2.89 ± 0.39 °C, respectively ( p value of 0.009). Thus, RBE containing cyclo(d-Phe-l-Pro) prevents the decrease in the cutaneous blood flow and skin temperature under air conditioning.

Roasted Barley Extract Affects Blood Flow in the Rat Tail and Increases Cutaneous Blood Flow and Skin Temperature in Humans.

Roasted barley extract (RBE, "Mugicha") is a traditional Japanese beverage reported to improve blood viscosity and affect food functionality. RBE is suggested to contain 2,5-diketopiperazines, which are the functional component with neuroprotective and immunostimulatory effects that are produced in food through roasting. In this study, we investigated the effects of RBE on blood circulation, both clinically and in rats. At first, we confirmed five 2,5-diketopiperazine derivatives in RBE by LC-MS analysis. Secondarily, we revealed that RBE affects blood flow in the rat tail and compared the efficacy on rat tail blood flow among five 2,5-diketopiperazines in RBE. Especially, cyclo(d-Phe-l-Pro) was the most effective in increasing blood flow in the rat tail. We also researched the mechanism of cyclo(d-Phe-l-Pro) with rat aorta study. As a result, we confirmed that cyclo(d-Phe-l-Pro) has an effect on vasodilatation through the release of nitric oxide in the vascular endothelium. Finally, we also confirmed that RBE affects cutaneous blood flow and increases skin temperature in humans.

Iso-α-acids, Bitter Components of Beer, Prevent Inflammation and Cognitive Decline Induced in a Mouse Model of Alzheimer's Disease.

Alongside the rapid growth in aging populations worldwide, prevention and therapy for age-related memory decline and dementia are in great demand to maintain a long, healthy life. Here we found that iso-α-acids, hop-derived bitter compounds in beer, enhance microglial phagocytosis and suppress inflammation via activation of the peroxisome proliferator-activated receptor γ. In normal mice, oral administration of iso-α-acids led to a significant increase both in CD11b and CD206 double-positive anti-inflammatory type microglia (p < 0.05) and in microglial phagocytosis in the brain. In Alzheimer's model 5xFAD mice, oral administration of iso-α-acids resulted in a 21% reduction in amyloid ß in the cerebral cortex as observed by immunohistochemical analysis, a significant reduction in inflammatory cytokines such as IL-1ß and chemokines including macrophage inflammatory protein-1α in the cerebral cortex (p < 0.05) and a significant improvement in a novel object recognition test (p < 0.05), as compared with control-fed 5xFAD mice. The differences in iso-α-acid-fed mice were due to the induction of microglia to an anti-inflammatory phenotype. The present study is the first to report that amyloid ß deposition and inflammation are suppressed in a mouse model of Alzheimer's disease by a single component, iso-α-acids, via the regulation of microglial activation. The suppression of neuroinflammation and improvement in cognitive function suggests that iso-α-acids contained in beer may be useful for the prevention of dementia.