Extended longevity has resulted in an increased number of patients with dementia. Therefore, new methods for treating and preventing dementia are highly desirable. Multiple health benefits have been reported for Grifola frondosa (maitake mushrooms). In this study, we investigated the safety and effects of continuous intake of maitake mushrooms on cognitive function in healthy older individuals. We conducted a randomized, double-blind, placebo-controlled trial over 18 wk targeting 47 healthy Japanese individuals aged 60 y or older. The individuals were randomly divided into three groups: a Y10M group consumed bread containing 50 g of the maitake mushroom strain Y10M, a C5304 group consumed bread containing 50 g of the maitake mushroom strain C5304, and a placebo group consumed bread without maitake mushrooms. No participants discontinued the intervention due to adverse effects in any group. The Y10M group showed a significant improvement in cognitive assessment based on the Japanese version of the Montreal Cognitive Assessment (MoCA-J) score compared to the placebo group after the intervention. However, no difference was observed in the MoCA-J scores between the C5304 and placebo groups. The activity of natural killer cells, a type of immune cell, was also significantly augmented in the Y10M group and positively correlated with MoCA-J scores. These results suggest that improved cognitive function is, at least in part, attributable to the activation of the immune system. This study suggests that consumption of maitake mushrooms may help inhibit the decline in cognitive function in healthy older individuals.

BACKGROUND:Cerebrotendinous xanthomatosis (CTX) is caused by biallelic mutations in the CYP27A1 gene, which encodes sterol 27-hydroxylase. Reduced activity of this enzyme decreases bile acid synthesis and increases cholestanol production. Cholestanol, a metabolic intermediate, can be used as a diagnostic marker; however, patients with CTX have been reported to have normal cholestanol levels. Cholesterol and bile acid metabolism is likely to be extensively disrupted even in these patients with CTX. OBJECTIVE:We aimed to clarify whether sterol markers, including oxysterols, can be used to screen for and diagnose CTX and assess treatment efficacy. METHODS:Fourteen patients with CTX (homozygotes, n = 7; compound heterozygotes, n = 5; unidentified, n = 2), 6 carriers, and 24 healthy controls were enrolled. We analyzed 9 sterol markers, including 4 oxysterols, using gas chromatography-mass spectrometry. We measured these markers in patients with CTX before and after treatment with chenodeoxycholic acid (CDCA) and/or lipid-lowering agents. RESULTS:Cholestanol and sterol markers of cholesterol synthesis and absorption were 1.3 to 8.3 times higher in patients with CTX than in carriers and controls. These marker levels decreased significantly in patients with CTX after treatment. Compared to carriers and controls, patients with CTX had extremely low levels of 27-hydroxycholesterol (27-OHC) and extremely high levels of 7α-hydroxycholesterol (7α-OHC). During treatment, 7α-OHC levels decreased significantly, while 27-OHC levels remained low. Receiver operating characteristic analysis demonstrated excellent diagnostic performance. The area under the curve was 1.000 for low 27-OHC and 0.991 for elevated 7α-OHC, outperforming cholestanol (0.903). CONCLUSION:Sterol markers, including oxysterols, can be used to screen for and diagnose CTX and to assess treatment efficacy with CDCA and/or lipid-lowering agents.

Postmenopausal women are at a higher risk of developing dyslipidemia and osteoporosis due to estrogen deficiency, necessitating regular vitamin D supplementation and the use of cholesterol inhibitors, respectively, to prevent these conditions. Despite current treatments, alternatives are needed to address both conditions simultaneously. Ergosterol, a precursor of vitamin D2, is a fungal sterol converted to brassicasterol by 7-dehydrocholesterol reductase, a cholesterol biosynthesis enzyme that converts 7-dehydrocholesterol (a precursor of vitamin D3) into cholesterol. Our previous research demonstrated that ergosterol decreases cholesterol levels and increases 7-dehydrocholesterol levels in human HepG2 hepatoma cells. Furthermore, we reported that ergosterol increases vitamin D2 levels, inhibits the cholesterol biosynthetic pathway, and potentially promotes vitamin D3 biosynthesis in male rats fed a high-fat and high-sucrose diet. In this study, we investigated the effects of daily high ergosterol intake on cholesterol, vitamin D2, and D3 biosynthetic pathways in ovariectomized (OVX) rats using GC-MS and LC with tandem mass spectrometry. OVX rats treated with ergosterol for 14 weeks exhibited significantly decreased plasma cholesterol levels and markers of cholesterol absorption, such as the plant sterol sitosterol. Furthermore, 7-dehydrocholesterol levels increased in these rats compared to untreated OVX rats. We observed that 1α, 25-dihydroxyvitamin D3 levels slightly increased in OVX rats treated with ergosterol. Additionally, ergosterol improved bone strength and increased OVX-induced bone resorption. These results indicate that daily ergosterol intake may aid in the simultaneous prevention of dyslipidemia and osteoporosis in postmenopausal women.

BackgroundDyslipidemia is a lifestyle-related disease; therefore, cholesterol biosynthesis inhibitors in foods can be easily ingested on a daily basis and are effective in aiding treatment and prevention. To assess the impact of this diet on health, it is of the essential thing that food intake can be properly measured, and it is important to find biomarkers of food intake. Previously, we reported that ergosterol, which is present in mushrooms, inhibits cholesterol biosynthesis. In this study, we measured serum ergosterol levels in healthy participants who consumed maitake mushroom bread to confirm actual ingestion of maitake mushrooms.MethodsSerum samples from healthy participants who consumed maitake mushroom bread (n = 24) or normal bread without maitake mushroom (placebo, n = 26) were analysed for ergosterol levels using liquid chromatography-tandem mass spectrometry with diene derivatization.ResultsIn the placebo group, there was no significant difference in ergosterol concentrations between baseline (before consumption) and 18 weeks. In contrast, the ergosterol concentration was 5-fold higher at 18 weeks than at baseline in the maitake mushroom bread-intake group.ConclusionMaitake mushroom bread intake for 18 weeks significantly increased serum ergosterol levels in healthy participants, suggesting that ergosterol is useful as a biomarker of mushroom intake.

BACKGROUND:Combination therapy with metformin and sodium-glucose cotransporter 2 (SGLT2) inhibitors is strongly recommended for adults with type 2 diabetes mellitus who have inadequate glycemic control. These inhibitors are particularly indicated for patients with a history or high risk of atherosclerotic cardiovascular disease, heart failure, or chronic kidney disease. Consequently, the number of patients receiving both metformin and SGLT2 inhibitors is expected to increase. However, these treatments pose a risk of life-threatening acidoses, specifically metformin-associated lactic acidosis (MALA) and euglycemic diabetic ketoacidosis (euDKA), which require prompt differentiation and distinct treatments. CASE REPORT:We present the case of a woman in her 50s with chronic heart failure who had been treated for type 2 diabetes for 15 years. After dehydration caused by working in hot conditions and fluid restriction, she presented with dehydration, fatigue, respiratory distress, hypotension, tachypnea, hypothermia, acute renal failure, hypoglycemia, and severe metabolic acidosis. Despite supportive care, lactic acidosis persisted, leading to a diagnosis of MALA. Hemodialysis improved lactate levels, but a persistent anion gap and elevated β-hydroxybutyrate level confirmed euDKA. Metabolic abnormalities resolved with continuous insulin infusion and the patient was discharged without sequelae. WHY SHOULD AN EMERGENCY PHYSICIAN BE AWARE OF THIS?: The combination of MALA and euDKA represents rare but potentially fatal conditions requiring distinct diagnostic and therapeutic approaches. With their growing use for type 2 diabetes management and cardiovascular/kidney benefits, emergency physicians are more likely to encounter complications associated with these medications.

BACKGROUND:Diarrhoea is one of the most serious complications in hereditary ATTR (ATTRv) amyloidosis. However, its precise pathomechanism remains unknown. The present study investigated the involvement of bile acid in diarrhoea along with the therapeutic effect of colestimide, a bile acid sequestrant, in ATTRv amyloidosis. METHODS:We prospectively enrolled 19 ATTRv amyloidosis patients (9 with refractory diarrhoea and 10 without diarrhoea) and 20 healthy individuals for measurements of serum 7a-hydroxy-4-cholesten-3-one (C4) levels. The patients with diarrhoea were then treated with oral colestimide (1.5 g twice daily) for 28 days. The frequency of diarrhoea and C4 level were evaluated before and after colestimide treatment. RESULTS:Mean serum C4 level was significantly higher in ATTRv patients with diarrhoea (62.3 ng/mL) than in ATTRv patients without diarrhoea (24.0 ng/mL, p = 0.03). Colestimide treatment significantly decreased mean diarrhoea frequency (pre-treatment period: 9.1 times/week, colestimide treatment period, 6.6 times/week, p = 0.04) and increased mean C4 level (before treatment: 66.2 ng/mL, after treatment: 187.1 ng/mL, p = 0.02). CONCLUSIONS:Bile acid status was significantly associated with diarrhoea in ATTRv amyloidosis. Colestimide and other bile acid sequestrants may reduce diarrhoea frequency in afflicted patients.

In adrenal fasciculata cells stimulated by ACTH, Ca2+ and cAMP play indispensable roles as second messengers in cortisol production. However, whether their second messengers cooperatively or independently participate in steroid production remains unclear. We focused on the roles of Ca2+ and cAMP in cortisol production in bovine adrenal fasciculata cells stimulated by ACTH for a relatively short period (1 h). Incubation of the cells with 100 pM ACTH in Ca2+-containing (normal) medium for 1 h increased cortisol production without affecting cAMP content. In contrast, treatment of the cells with the peptide at a higher concentration (1 nM) significantly augmented both cortisol production and cAMP content. However, ACTH did not increase either of them in the Ca2+-free medium. ACTH rapidly increased the intracellular free Ca2+ concentration ([Ca2+]i) in the normal medium, but did not influence [Ca2+]i in the Ca2+-free medium, indicating that ACTH caused Ca2+ influx into the cells. ACTH-induced Ca2+ influx and cortisol production were suppressed by a voltage-sensitive Ltype Ca2+ channel blocker but not by a T-type, N-type, or P-type Ca2+ channel blocker. In contrast, dibutyryl cAMP, a cellpermeable cAMP analog, greatly enhanced cortisol production in the normal or Ca2+-free medium and slowly caused Ca2+ influx into the cells. These results strongly suggest that Ca2+, as a second messenger, is more critical than cAMP for cortisol production. However, both second messengers jointly participate in the production in adrenal fasciculata cells stimulated by ACTH.

Background/Aim: The prognosis of patients with multiple myeloma (MM) has recently improved due to the emergence of new molecular targeting agents. However, MM remains incurable because MM stem cells are resistant to these agents. Therefore, it is essential to develop strategies to eradicate MM stem cells. We have previously demonstrated that MM cells cultured under prolonged hypoxic conditions (1% O2) (i.e., hypoxia-adapted MM cells; MM-HA cells) exhibited stem-cell-like characteristics. γδ T cells attack tumor cells by recognizing butyrophilin (BTN) 3A1 and BTN2A1, which are activated by the intracellular accumulation of isopentenyl pyrophosphate (IPP), an intermediate in the mevalonate pathway. In the present study, we investigated the cytotoxicity of γδ T cells against MM-HA stem-like cells. Materials and Methods: We used a combination of flow cytometry, liquid chromatography-tandem mass spectrometry, and western blotting methods to investigate the cytotoxicity of γδ T cells against MM-HA cells and measured the amounts of IPP in MM-HA cells and their supernatants. Results: The cytotoxicity of γδ T cells against MM-HA cells was significantly lower than that against MM cells cultured under normoxic conditions (20% O2; MM-Normo). Furthermore, the concentration of IPP in MM-HA cells was lower than that in MM-Normo cells. The expression of mevalonate decarboxylase and farnesyl diphosphate synthase proteins were decreased in MM-HA-cells. Conclusion: The cytotoxicity of γδ T cells against MM-HA cells was suppressed by the reduced IPP accumulation by modulating the mevalonate pathway in MM-HA cells.

Dyslipidemia is a lifestyle-related (physical inactivity or obesity) disease; therefore, dietary foods that can easily be consumed in daily life is important to prevent dyslipidemia. Ergosterol, a precursor of vitamin D2, is a fungal sterol present in the membranes of edible mushrooms and other fungi. Ergosterol is converted to brassicasterol by 7-dehydrocholesterol reductase (DHCR7), a cholesterol biosynthesis enzyme that converts 7-dehydrocholesterol (a precursor of vitamin D3) into cholesterol. Previously, we reported that ergosterol increases 7-dehydrocholesterol, decreases cholesterol levels by competitive effect of DHCR7, and reduces DHCR7 mRNA and protein levels in human HepG2 hepatoma cells. Here, we investigated the effects of long-term high ergosterol intake on the cholesterol, vitamin D2, and D3 biosynthetic pathways of rats fed a high-fat and high-sucrose (HFHS) diet using GC-MS and LC with tandem mass spectrometry. In HFHS rats, oral ergosterol administration for 14 weeks significantly decreased plasma low-density lipoprotein cholesterol, total bile acid, and cholesterol precursor (squalene and desmosterol) levels and increased 7-dehydrocholesterol levels compared to HFHS rats without ergosterol. Ergosterol, brassicasterol, and vitamin D2 were detected, cholesterol levels were slightly decreased, and levels of vitamin D3 and its metabolites were slightly increased in rats fed HFHS with ergosterol. These results showed that ergosterol increased vitamin D2 levels, inhibited the cholesterol biosynthetic pathway, and possibly promoted vitamin D3 biosynthesis in vivo. Therefore, daily ergosterol intake may aid in the prevention of dyslipidemia.

Simultaneous administration of enteral formula and phenytoin in the clinical setting is known to reduce the plasma concentration of phenytoin. In this study, we examined the binding of phenytoin with enteral formulas and its components by quantifying the free phenytoin concentration. Furthermore, we investigated the effect of enteral formulas on gastrointestinal absorption of phenytoin in rats. The free phenytoin rate was reduced in vitro when phenytoin and enteral formula or pectin, a dietary fiber in enteral formulas, were co-administered. In vivo, when phenytoin and the enteral formula Mei Balance R® were co-administered, the time to maximum plasma concentration (Tmax) after oral administration was significantly increased. Moreover, the area under the phenytoin concentration-time curve from time zero to 6 h (AUC0-6 h) was significantly increased by co-administration of phenytoin with the enteral formula PG Soft EJ®. These results showed the gastrointestinal absorption of phenytoin differs according to the type of enteral formula. In addition, we found the first time that plasma phenytoin levels increase when combined with enteral formula. Among the components of enteral formulas, in particular, milk protein delayed the absorption of phenytoin. Moreover, milk protein, casein and carrageenan tended to increase AUC0-6 h. These results suggest the change in phenytoin concentration is due not only to the binding of enteral formula but also to the disintegration of components such as protein. Therefore, when co-administrated of phenytoin and enteral formula, phenytoin must be monitored frequently according to the enteral formula interaction.

The purpose of this study was to clarify the effects of enteral formulas and their food protein and dietary fiber components on the pharmacokinetics of warfarin in rats. Blood samples were collected from 0 to 30 h after the oral administration of 0.5 (milk protein or casein), or dietary fiber (indigestible dextrin or carrageenan). Plasma warfarin concentration was determined by liquid chromatography-tandem mass spectrometry. Plasma warfarin concentration was lower at the initial phase after co-administration of F2 Light & REG;, milk protein, or casein compared with the control group, which might have been caused by the suppression of the gastrointestinal absorption of warfarin in accordance with the binding to food protein. Plasma warfarin concentration was significantly higher after co-administration of Mei Flow & REG;, indigestible dextrin, or carrageenan compared with the control group. This is the first study to find that the plasma concentration of warfarin increases when warfarin is co-administered with enteral formula Mei Flow & REG; or their dietary fiber components such as indigestible dextrin or carrageenan. Therefore, close attention should be paid to decreases or increases in the plasma warfarin concentration when warfarin is used with enteral formula.

Current treatment approaches for hyperlipidemia rely mainly on reducing the cholesterol level by inhibiting 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), which is involved in the presqualene pathway of cholesterol biosynthesis. Finding a compound that instead targets the postsqualene pathway could aid in the treatment of hyperlipidemia and synergistically reduce the cholesterol level when used in conjunction with HMGCR inhibitors. Ergosterol is a fungal sterol that is converted to brassicasterol by 7-dehydrocholesterol reductase (DHCR7). DHCR7 is also a cholesterol biosynthesis enzyme, and thus ergosterol may cause the accumulation of 7-dehydrocholesterol, a precursor of cholesterol and vitamin D3 , by a competitive effect. In this study, we examined the effect of ergosterol on the postsqualene pathway by quantifying cholesterol precursors and related sterols using gas chromatography-mass spectrometry and by conducting quantitative RT-PCR and western blot analysis for human HepG2 hepatoma cells. We found that ergosterol is converted into brassicasterol by the action of DHCR7 from HepG2 cells and that it induced the accumulation of cholesterol precursors (lathosterol, 7-dehydrocholesterol, and desmosterol) and decreased the cholesterol level by altering the mRNA and protein levels of cholesterol biosynthesis enzymes (increase of sterol 8,7-isomerase [EBP] and decrease of DHCR7 and 24-dehydrocholesterol reductase [DHCR24]). These results demonstrate that ergosterol inhibits the postsqualene pathway and may be useful for the prevention of hyperlipidemia.

Depression can be activated by dysfunction of the hypothalamus–pituitary–adrenal (HPA) axis and is sometimes accompanied by increased cortisol production. In this study, the effects of Kampo extract products on cortisol production in bovine adrenal fasciculata cells via adrenocorticotropic hormone (ACTH) stimulation were investigated with the aim of identifying substances that can improve the function of the HPA axis. Daiokanzoto extract product, which is prepared from Rhei Rhizoma and Glycyrrhizae Radix as the constituent crude drugs, was found to potently inhibit cortisol production in bovine adrenal fasciculata cells. Because only Rhei Rhizoma decoction showed potent inhibitory effect, the cortisol production inhibitory substances in Rhei Rhizoma was analyzed. As a result, six anthraquinones: chrysophanol, emodin, aloe-emodin, rhein, chrysophanol-8-O-β-D-glucopyranoside, and physicion-8-O-β-D-glucopyranoside were isolated and identified from Rhei Rhizoma decoction. Among these compounds, emodin showed the strongest inhibition of the ACTH-induced cortisol production in cells (IC50 = 13.1 ± 1.9 µM). These results suggest that emodin may help of stress-induced illnesses such as depression.

The androgens testosterone and dihydrotestosterone (DHT) are essential for a variety of systemic functions in mature males. Alteration of these hormones results in late-onset hypogonadism (LOH) and benign prostate hyperplasia (BPH). The fruit bodies of fungi of the genus Cordyceps have been regarded as folk medicine or health food with tonic and antifatigue effects. The extract from the fruit body of Cordyceps militaris parasitizing Samia cynthia ricini (CM) was evaluated as a novel-candidate natural product for ameliorating male andropause symptoms. To explore the effects of CM on LOH and BPH, CM was applied to rat models and cultured testicular cells and prostate cells. The concentrations of androgens in the serum and culture media were determined by ELISA. Expression of steroidogenic enzymes and androgen-related genes was evaluated by qPCR, and prostatic cell proliferation was assessed with the cell-viability assay. CM maintained the serum levels of testosterone and DHT, but inhibited testosterone-induced prostate hypertrophy. CM also increased the secretion of testosterone and DHT by primary testicular cells, with no changes in the mRNA expression of steroidogenic enzymes, but decreased the growth of prostatic cell lines. Our data suggest that CM could improve both LOH and BPH in males.

Obesity-induced endoplasmic reticulum (ER) stress contributes to low-grade chronic inflammation, in adipose tissue and results in metabolic disorders such as diabetes mellitus and dyslipidemia. Sepsis displays a major cause of death on intensive care units and also contributes substantially to mortality after surgery. The incidence of severe sepsis in obesity patients is higher than that in non-obesity patients. Sepsis is a complex acute inflammatory disease, and the inflammatory response is characterized by the systemic circulation of inflammatory cytokines. However, changes in adipocyte function under ER stress and the role of adipose tissues in sepsis remains understood. Our previous study revealed that alpha1-antitrypsin (SERPINA1: A1AT), which exhibits tissue protective action and the activity of serine protease inhibiter, is highly expressed in adipose tissues and adipocytes. We determined here the importance of adipocyte A1AT expression for adipocyte homeostasis under ER stress. To address this issue, the ER stress activator thapsigargin was treated to differentiated SW872 adipocytes after A1AT siRNA treatment. Thapsigargin markedly increased the basal levels of pro-inflammatory adipokines (IL-1beta, IL-6, and IL-8) expression. Knocking down A1AT enhanced the expression of thapsigargin-stimulated adipokines. Furthermore, the knockdown of A1AT reduced insulin-induced phosphorylation of Akt, whereas stimulated TNF-alpha-triggered pro-inflammatory cytokines. These findings indicate the possibility that endogenous A1AT may negatively regulate inflammatory mechanism to inhibit excessive pro-inflammatory adipokine expression and play a protective role in adipose tissue dysfunction mediated through ER stress activation.

Cystatins are thiol proteinase inhibitors present ubiquitously in mammalian system. They prevent unwanted proteolysis and are involved in several neurodegenerative diseases. Hydrogen peroxide, hydroxyl radical and superoxide radical were produced during the autoxidation of various cytotoxic agents namely 6-hydroxydopamine, 6-aminodopamine, 6,7-dihydroxytryptamine, and dialuric acid.Reactive oxygen species (ROS) such as singlet oxygen [O2−] and hydroxyl radicals result in nonspecific reactivity, thus involved in several pathological conditions such as inflammation and reperfusion injury leading to damage of different biological molecules due to the production of enzyme myeloperoxidase because of activation of neutrophils in these diseases.As Myeloperoxidase [MPO] is released extracellularly by activated monocytes and is highly basic protein that damages the localized sites. The onset and progression of several diseases is a complex process in which oxidation is believed to play significant role. Oxidized lipids and proteins that are detected in early, intermediate and advanced stage of human neurodegenerative diseases (plaques) have revealed considerably elevated levels of reactive oxygen species showing their involvement in brain diseases.In the present study, it has been demonstrated that H2O2 induced modifications in brain cystatin leads to its inactivation and degradation. It was also found that the damage from the oxidants occurred mainly because of the (ROS) hydroxyl radicals [OH] produced by H2O2 which in turn changes the conformation of the Cystatin, a major factor for the damage of this significant protein. It is one of the cause of protease-antiprotease imbalance in mammals leading to progression of several diseases.