Protective effect of Ganoderma lucidum spore extract in trimethylamine-N-oxide-induced cardiac dysfunction in rats.

Previous research has shown that the extracts from the Ganoderma lucidum spore (GS) have potentially cardioprotective effects, but there is still abundant room for development in determining its mechanism. In this study, the rat model of cardiac dysfunction was established by intraperitoneal injection of trimethylamine-N-oxide (TMAO), and the extracts of GS (oil, lipophilic components, and polysaccharides) were given intragastrically at a dose of 50 mg/kg/day to screen the pharmacological active components of GS. After 50 days of treatments, we found that the extraction from GS reduced the levels of total cholesterol, triglyceride, and low-density lipoprotein; increased the levels of high-density lipoprotein; and reduced the levels of serum TMAO when compared to the model group (P < 0.05); especially the GS polysaccharides (DT) and GS lipophilic components (XF) exhibited decreases in serum TMAO compared to TMAO-induced control. The results of 16S rRNA sequencing showed that GS could change the gut microbiota, increasing the abundance of Firmicutes and Proteobacteria in the DT-treated group and XF-treated group, while reducing the abundance of Actinobacteria and Tenericutes. Quantitative proteomics analysis showed that GS extracts (DT and XF) could regulate the expression of some related proteins, such as Ucp1 (XF-TMAO/M-TMAO ratio is 2.76), Mpz (8.52), Fasn (2.39), Nefl (1.85), Mtnd5 (0.83), Mtnd2 (0.36), S100a8 (0.69), S100a9 (0.70), and Bdh1 (0.72). The results showed that XF can maintain the metabolic balance and function of the heart by regulating the expression of some proteins related to cardiovascular disease, and DT can reduce the risk of cardiovascular diseases by targeting gut microbiota.

Corrigendum: Anti-breast Cancer Enhancement of a Polysaccharide From Spore of Ganoderma lucidum With Paclitaxel: Suppression on Tumor Metabolism With Gut Microbiota Reshaping.

[This corrects the article DOI: 10.3389/fmicb.2018.03099.].

Alcohol Extracts From Ganoderma lucidum Delay the Progress of Alzheimer's Disease by Regulating DNA Methylation in Rodents.

Age-related changes in methylation are involved in the occurrence and development of tumors, autoimmune disease, and nervous system disorders, including Alzheimer's disease (AD), in elderly individuals; hence, modulation of these methylation changes may be an effective strategy to delay the progression of AD pathology. In this study, the AD model rats were used to screen the main active extracts from the mushroom, Ganoderma lucidum, for anti-aging properties, and their effects on DNA methylation were evaluated. The results of evaluation of rats treated with 100 mg/kg/day of D-galactose to induce accelerated aging showed that alcohol extracts of G. lucidum contained the main active anti-aging extract. The effects on DNA methylation of these G. lucidum extracts were then evaluated using SAMP8 and APP/PS1 AD model mice by whole genome bisulfite sequencing, and some methylation regulators including Histone H3, DNMT3A, and DNMT3B in brain tissues were up-regulated after treatment with alcohol extracts from G. lucidum. Molecular docking analysis was carried out to screen for molecules regulated by specific components, including ganoderic acid Mk, ganoderic acid C6, and lucidone A, which may be active ingredients of G. lucidum, including the methylation regulators of Histone H3, MYT, DNMT3A, and DNMT3B. Auxiliary tests also demonstrated that G. lucidum alcohol extracts could improve learning and memory function, ameliorate neuronal apoptosis and brain atrophy, and down-regulate the expression of the AD intracellular marker, Aß1-42. We concluded that alcohol extracts from G. lucidum, including ganoderic acid and lucidone A, are the main extracts involved in delaying AD progression.

Antitumor Activity of Extract From the Sporoderm-Breaking Spore of Ganoderma lucidum: Restoration on Exhausted Cytotoxic T Cell With Gut Microbiota Remodeling.

As breast cancer is the leading cause of cancer-related deaths in women population worldwide, ongoing endeavor has been made for alternative regimens with improved efficacy but fewer adverse effects. Recently, active components from the spores of Ganoderma lucidum have attracted much attention for their versatile biological activities owing to the advance in sporoderm-breaking technology. Here, anticancer potential of an extract derived from the sporoderm-breaking spores of G. lucidum (ESG) was explored in a 4T1-breast cancer xenograft mice model. Results showed that ESG was able to suppress 4T1 tumor growth in vivo rather than in vitro. Flowcytometry analysis revealed that ESG could significantly increase both cytotoxic T cell (Tc) population and the ratio of Tc to helper T cell (Th) in peripheral blood of the tumor-bearing mouse; similar promotion on Tc was also found in tumor-infiltrating lymphocyte. Moreover, ESG evidently downregulated the two immune checkpoints, programmed cell death protein-1 (PD-1, in the spleen) and cytotoxic T lymphocyte antigen-4 (CTLA-4, in the tumor), suggesting that ESG could effectively restore the T cell paradigm by recovering the exhaustion status via suppressing the co-inhibitory checkpoints. By 16S rRNA gene sequence analysis on the fecal microbiota, it was found that ESG would remodeling the overall structure of the samples from tumor-bearing mice toward that of the normal counterparts, including 18 genera in 5 phyla, together with regulations on several genes that are responsible for signaling pathways involved in metabolism, cellular processes, and environmental information processing. Collectively, this study demonstrated that ESG would serve as a natural anticancer adjuvant via a restoration on the exhausted Tc, highlighting important clinical implications for the treatment of breast cancer.

Hypouricemic Effects of Extracts From Agrocybe aegerita on Hyperuricemia Mice and Virtual Prediction of Bioactives by Molecular Docking.

Agrocybe aegerita has long been utilized for promoting diuresis in traditional Chinese medicine (TCM) with a close correlation to hypouricemia. Ethanol (AAE) and water (AAW) extracts of the compound led to a remarkable decrease in serum uric acid levels (SUA) in hyperuricemia mice, approaching that of the normal control. Both AAE and AAW exhibited suppression effects on hepatic xanthine oxidase (XOD) activities and elevation effects on renal OAT1 (organic anion transporter 1). However, only little negative impact was observed on the inner organ functions. The molecular docking was used to screen our in-home compound database for A. aegerita, and four compounds including 2-formyl-3,5-dihydroxybenzyl acetate, 2,4-dihydroxy-6-methylbenzaldehyde, 2-(6-hydroxy-1H-indol-3-yl)acetamide, and 6-hydroxy-1H-indole-3-carbaldehyde (HHC) were identified as potential active compounds. Their inhibitory mechanism on XOD might be attributed to their localization in the tunnel for the entrance of substrates to XOD active site, preventing the entrance of the substrates. To confirm the activity of the screened compounds experimentally, HHC was selected due to its high ranking and availability. The assaying result suggested the significant inhibitory activity of HHC on XOD. Also, these compounds were predicted to carry good ADME (absorption, distribution, metabolism, and excretion) properties, thereby necessitating further investigation. The current results provided an insight into the hypouricemic effects of macrofungi and their bioactives, which might provide the significant theoretical foundation for identifying and designing novel hypouricemia compounds.

Hypouricemic Effects of Armillaria mellea on Hyperuricemic Mice Regulated through OAT1 and CNT2.

Ethanol and water extracts of Armillaria mellea were prepared by directly soaking A. mellea in ethanol (AME) at 65[Formula: see text]C, followed by decocting the remains in water (AMW) at 85[Formula: see text]C. Significantly, AME and AMW at 30, 60 and 120[Formula: see text]mg/kg exhibited excellent hypouricemic actions, causing remarkable declines from hyperuricemic control (351[Formula: see text][Formula: see text]mol/L, [Formula: see text]) to 136, 130 and 115[Formula: see text][Formula: see text]mol/L and 250, 188 and 152[Formula: see text][Formula: see text]mol/L in serum uric acid, correspondingly. In contrast to the evident renal toxicity of allopurinol, these preparations showed little impacts. Moreover, they showed some inhibitory effect on XOD (xanthine oxidase) activity. Compared with hyperuricemic control, protein expressions of OAT1 (organic anion transporter 1) were significantly elevated in AME- and AMW-treated mice. The levels of GLUT9 (glucose transporter 9) expression were significantly decreased by AMW. CNT2 (concentrative nucleoside transporter 2), a key target for purine absorption in gastrointestinal tract was involved in this study, and was verified for its innovative role. Both AME and AMW down-regulated CNT2 proteins in the gastrointestinal tract in hyperuricemic mice. As they exhibited considerable inhibitory effects on XOD, we selected XOD as the target for virtual screening by using molecular docking, and four compounds were hit with high ranks. From the analysis, we concluded that hydrogen bond, Pi-Pi and Pi-sigma interactions might play important roles for their orientations and locations in XOD inhibition.

Cordycepin, a Characteristic Bioactive Constituent in Cordyceps militaris, Ameliorates Hyperuricemia through URAT1 in Hyperuricemic Mice.

Recently, we've reported the anti-hyperuricemic effects of Cordyceps militaris. As a characteristic compound of C. militaris, we hypothesized that cordycepin may play a role in preventing hyperurecimia. Remarkably, cordycepin produced important anti-hyperuricemic actions, decreasing SUA (serum uric acid) to 216, 210, and 203 µmol/L (P < 0.01) at 15, 30, and 60 mg/kg in comparison of hyperuricemic control (337 µmol/L), closing to normal control (202 µmol/L). Elisa, RT-PCR and western blot analysis demonstrated that the actions may be attributed to its downregulation of uric acid transporter 1 (URAT1) in kidney. Serum creatinine levels and blood urine nitrogen and liver, kidney, and spleen coefficients demonstrated that cordycepin may not impact liver, renal, and spleen functions. In addition, we used computational molecular simulation to investigate the binding mechanism of cordycepin. Of which, van der Waals interaction dominated the binding. Residues TRP290, ARG17, ALA408, GLY411, and MET147 contributed mainly on nonpolar energy. This provided the theoretical guidance to rationally design and synthesis novel URAT1 inhibitors.

Anti-breast Cancer Enhancement of a Polysaccharide From Spore of Ganoderma lucidum With Paclitaxel: Suppression on Tumor Metabolism With Gut Microbiota Reshaping.

Increasing evidence highlights the cardinal role of gut microbiota in tumorigenesis and chemotherapy outcomes. Paclitaxel (PTX), although as a first-line chemotherapy reagent for breast cancer, still requires for improvement on its efficacy and safety due to drug resistance and adverse effects. The present work explored the enhancement of a polysaccharide derived from spore of Ganoderma lucidum (SGP) with PTX in a murine 4T1-breast cancer model. Results showed that the combination of PTX and SGP displayed an improved tumor control, in which mRNA expression of several Warburg effect-related proteins, i.e., glucose transporter 3 (Glut3), lactate dehydrogenase A (Ldha), and pyruvate dehydrogenase kinase (Pdk), and the metabolite profile of tumor was evidently altered. Flowcytometry analysis revealed that the combination treatment recovered the exhausted tumor infiltration lymphocytes (TILs) via inhibiting the expressions of immune checkpoints (PD-1 and Tim-3), while PTX alone evidently increased that of CTLA-4. 16S rRNA sequencing revealed a restoration by the combination treatment on gut microbiota dysbiosis induced by PTX, especially that Bacteroides, Ruminococcus, and other 5 genera were significantly enriched while the cancer-risk genera, Desulfovibrio and Odoribacter, were decreased. Moreover, spearman correlation analysis showed that abundance of Ruminococcus was significantly negative-associated with the amount of frucotose-6-phosphate within the tumor. Collectively, the present study suggests the clinical implication of SGP as an adjuvant candidate for PTX against breast cancer, which possibly relies on the regulation of tumor metabolism and gut microbiota.

Prebiotic Effect of Fructooligosaccharides from Morinda officinalis on Alzheimer's Disease in Rodent Models by Targeting the Microbiota-Gut-Brain Axis.

Gut microbiota influences the central nervous system disorders such as Alzheimer's disease (AD). The prebiotics and probiotics can improve the host cognition. A previous study demonstrated that fructooligosaccharides from Morinda officinalis (OMO) exert effective memory improvements in AD-like animals, thereby considered as potential prebiotics; however, the underlying mechanism still remains enigma. Thus, the present study investigated whether OMO is effective in alleviating AD by targeting the microbiota-gut-brain axis. OMO was administered in rats with AD-like symptoms (D-galactose- and Aß1-42-induced deficient rats). Significant and systematic deterioration in AD-like animals were identified, including learning and memory abilities, histological changes, production of cytokines, and microbial community shifts. Behavioral experiments demonstrated that OMO administration can ameliorate the learning and memory abilities in both AD-like animals significantly. AD parameters showed that OMO administration cannot only improve oxidative stress and inflammation disorder, but also regulate the synthesis and secretion of neurotransmitter. Histological changes indicated that OMO administration ameliorates the swelling of brain tissues, neuronal apoptosis, and down-regulation of the expression of AD intracellular markers (Tau and Aß1-42). 16S rRNA sequencing of gut microbiota indicated that OMO administration maintains the diversity and stability of the microbial community. In addition, OMO regulated the composition and metabolism of gut microbiota in inflammatory bowel disease (IBD) mice model treated by overdosed antibiotics and thus showed the prebiotic potential. Moreover, gut microbiota plays a major role in neurodevelopment, leading to alterations in gene expression in critical brain and intestinal regions, thereby resulting in perturbation to the programming of normal cognitive behaviors. Taken together, our findings suggest that the therapeutic effect of the traditional medicine, M. officinalis, on various neurological diseases such as AD, is at least partially contributed by its naturally occurring chemical constituent, OMO, via modulating the interaction between gut ecology and brain physiology.

Hypouricemic Effects of Ganoderma applanatum in Hyperuricemia Mice through OAT1 and GLUT9.

Ganoderma applanatum (G. applanatum) dispels wind to eliminate dampness and exhibited nephron- and liver-protective effects as noted in Chinese herbal classic literature; it might also affect hyperuricemia. Therefore, we examined the hypouricemia effects and mechanisms underlying G. applanatum on chemical-induced hyperuricemia in mice. Ethanol (GAE) and water (GAW) extracts were prepared by extracting G. applanatum in ethanol (GAE), followed by bathing the remains in water to yield GAW. GAE and GAW were administered orally at different doses to hyperuricemia mice, while allopurinol and benzbromarone served as positive controls. Both GAE and GAW showed remarkable hypouricemia activities, rendering a substantial decline in the SUA (serum uric acid) level in hyperuricemia control (P < 0.01). Moreover, the urine uric acid (UUA) levels were enhanced by GAE and GAW. In contrast to the evident renal toxicity of allopurinol, GAE and GAW did not show a distinct renal toxicity. Almost no suppressing effect was observed on the XOD activities. However, compared to the hyperuricemia control, OAT1 was elevated remarkably in mice drugged with GAE and GAW, while GLUT9 was significantly decreased. Similar to benzbromarone, GAE decreased the URAT1 protein levels significantly (P < 0.01), while GAW did not display a similar effect. GAE and GAW downregulated the level of CNT2 proteins in the gastrointestinal tract of hyperuricemia mice. Thus, G. applanatum produced outstanding hypouricemic effects, mediated by renal OAT1, GLUT9, and URAT1 and gastrointestinal CNT2 that might elevate urine uric secretions and decline in the absorption of purine in the gastrointestinal tracts. G. applanatum showed little negative influence on inner organs. By docking screening, four top-ranked compounds were identified that necessitated further investigation. Compounds: potassium oxonate, hypoxanthine, allopurinol, benzbromarone.

Actions of water extract from Cordyceps militaris in hyperuricemic mice induced by potassium oxonate combined with hypoxanthine.

ETHNOPHARMACOLOGICAL RELEVANCE: Cordyceps militaris was recorded in the classic traditional Chinese medicine book with the main functions of "protecting liver and enhancing kidney functions", influencing serum uric acid levels. AIM OF STUDY: The aim is to investigate the hypouricemic effects and possible mechanism of C. militaris in hyperuricemic mice. MATERIALS AND METHODS: A water extract (WECM) was prepared by decocting C. militaris directly at 80 °C in water bath, followed by lyophilization. WECM at 50, 100 and 200mg/kg was orally administered to hyperuricemic mice induced by potassium oxonate and hypoxanthine combinedly and allopurinol (5mg/kg) was served as a positive control. RESULTS: WECM exhibited excellent hypouricemic activity, which could decrease the serum uric acid levels of the hyperuricemic mice (306µmol/L) to 189, 184 and 162µmol/L at different doses respectively (P<0.01), approaching the levels of normal mice (184µmol/L). The urate transporter 1 (URAT1) protein levels of kidney at different doses of WECM were 28.15, 17.43, 9.03pg/mL respectively, much lower than that in the hyperuricemia group (93.45pg/mL, P<0.01); and suggested WECM may interact with URAT1. Docking simulations using modeled structure of URAT1 suggested that LYS145, ARG325, ARG477 and ASP168 of URAT1 are key functional residues of URAT1. Four active compounds in C. militaris were identified and their interaction energies with target were estimated between -200 and -400kcal/mol. CONCLUSIONS: These findings suggested that C. militaris produced significant hypouricemic actions and the hypouricemic effects of WECM may be attributed to the inhibitive effect of WECM on URAT1 protein levels. The results of blood urine nitrogen and serum creatinine levels and liver, kidney and spleen coefficients showed that WECM have no negative impacts on liver, renal and spleen functions. The screened four active compounds using molecular docking method deserve further investigation in other work.

Effect of bajijiasu isolated from Morinda officinalis F. C. how on sexual function in male mice and its antioxidant protection of human sperm.

ETHNOPHARMACOLOGICAL RELEVANCE: In recent years, the physiological aspects of human fertility have been seriously influenced by the interactions of genetic and environmental factors. Almost one in 20 males has been affected by male infertility, providing a great challenge and an opportunity to use natural compounds as alternatives to chemical drugs with comprehensive adverse effects. However, ample evidences are scanty to support the physiological mechanisms of natural compounds used to treat male infertility. In traditional Chinese medicine, Morinda officinalis F. C. How is widely used as a herb that invigorates the kidneys and supports yang, the original energy in the human body, to resist diseases and in treating male infertility. In this study, we evaluated whether bajijiasu isolated from the roots of M. officinalis F.C. How is a potential agent for the treatment of male infertility. MATERIALS AND METHOD: In this study, both normal and kidney-yang-deficient mice were administered bajijiasu orally at different concentrations. To determine the pharmacological mechanism of bajijiasu, we observed the sexual behavior and genital organ coefficients, determined their serum hormone levels, analyzed their sperm quality parameters, and examined histopathological sections from them. We also used enzymatic assays to determine the effects of bajijiasu on superoxide dismutase, glutathione peroxidase, and malondialdehyde. Confocal micro-Raman spectroscopy was used to investigate the changes in the DNA of H2O2-damaged human sperm after treatment with bajijiasu in vitro. RESULTS: Our results showed that bajijiasu enhanced the sexual behavior of both normal and kidney-yang-deficient mice. It also markedly increased the testosterone concentrations, reduced the levels of cortisol, improved the quality of the sperm, and counteracted the histopathological impairment induced by hydroxyurea in the kidney-yang-deficient mice. The enzymatic assay and Raman spectra showed that bajijiasu protects the DNA of sperm from damage by H2O2. CONCLUSION: Bajijiasu is a potential androgen-like drug that modulates hormone levels to some extent without producing reproductive-organ lesions, enhances the sexual function of male mice, and protects the DNA of human sperm from H2O2 damage. Thus, bajijiasu is an active ingredient of M. officinalis F.C. How that improves the human reproductive capacity.

Protective effect of Bajijiasu against ß-amyloid-induced neurotoxicity in PC12 cells.

Beta-amyloid peptide (Aß), a major protein component of senile plaques associated with Alzheimer's disease (AD), is also directly neurotoxic. Mitigation of Aß-induced neurotoxicity is thus a possible therapeutic approach to delay or prevent onset and progression of AD. This study evaluated the protective effect of Bajijiasu (ß- D-fructofuranosyl (2-2) ß- D-fructofuranosyl), a dimeric fructose isolated from the Chinese herb Radix Morinda officinalis, on Aß-induced neurotoxicity in pheochromocytoma (PC12) cells. Bajijiasu alone had no endogenous neurotoxicity up to 200 µM. Brief pretreatment with 10-40 µM Bajijiasu (2 h) significantly reversed the reduction in cell viability induced by subsequent 24 h exposure to Aß25-35 (21 µM) as measured by MTT and LDH assays, and reduced Aß25-35-induced apoptosis as indicated by reduced annexin V-EGFP staining. Bajijiasu also decreased the accumulation of intracellular reactive oxygen species and the lipid peroxidation product malondialdehyde in PC12 cells, upregulated expression of glutathione reductase and superoxide dismutase, prevented depolarization of the mitochondrial membrane potential (Ψm), and blocked Aß25-35-induced increases in [Ca(2+)] i . Furthermore, Bajijiasu reversed Aß25-35-induced changes in the expression levels of p21, CDK4, E2F1, Bax, NF-κB p65, and caspase-3. Bajijiasu is neuroprotective against Aß25-35-induced neurotoxicity in PC12 cells, likely by protecting against oxidative stress and ensuing apoptosis.