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Research background: Tiger milk mushroom (Lignosus rhinocerus) is a medicinal mushroom that is geographically distributed in the region of South China, Thailand, Malaysia, Indonesia, Philippines and Papua New Guinea. Consumption of its sclerotium has been reported to treat various ailments. However, its anticancer potential towards oral cancer cell lines is yet to be determined considering the traditional method of its consumption by biting/chewing of the sclerotium. Experimental approach: Mushroom sclerotial powder of cultivar TM02® was extracted and fractionated in a chromatographic column prior to cytotoxicity testing against a panel of human oral cancer cell lines. The capability of the identified bioactive fraction in regulating several molecules associated with its tumour necrosis factor (TNF) pathway was investigated. Results and conclusions: 2,5-Diphenyl-2H-tetrazolium bromide (MTT) proliferation assay indicated that cell lines ORL-48 (derived from gingiva), ORL-188 (derived from the tongue) and ORL-204 (derived from buccal mucosa) were inhibited by cold water extract of L. rhinocerus sclerotia and its high-molecular-mass fraction (HMM) in varying degrees with ORL-204 being most affected. Hence, the treatment of ORL-204 with HMM mushroom extract was further investigated. HMM mushroom extract induced apoptosis and G0/G1 phase cell cycle arrest through caspase-3/7 cleavage. Activities of MIP2 and COX-2 were downregulated by 0.2- and 4.6-fold respectively in the HMM mushroom extract-treated ORL-204 cells. Novelty and scientific contribution: Using ORL-204, we showed that HMM mushroom extract may act via the TNF pathway at various network sites as a potential dietary compound for cancer prevention and natural adjunct therapeutic to conventional cancer treatment.
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RESEARCH BACKGROUND: Ophiocordyceps sinensis, a highly valued medicinal fungus, is close to extinction due to overexploitation. Successful cultivation of O. sinensis fruiting body (OCS02®) shows that the cultivar has a promising nutritional value and numerous bioactive compounds. Antioxidant and antiproliferative properties and biologically active proteins of the OCS02® are investigated for possible development into nutraceuticals. EXPERIMENTAL APPROACH: The chemical composition of the OCS02® cold water extract was determined, and the antioxidant activities were examined using ferric reducing, DPPH⢠and O2 â¢- scavenging assays. Tetrazolium dye (MTT) cytotoxic assay was performed to assess the antiproliferative activity of the extract. Bioactive proteins in the active fraction of the extract were identified using liquid chromatography (LC) and tandem-mass spectrometry (MS/MS). RESULTS AND CONCLUSIONS: The OCS02® extract exhibited strong O2 â¢- scavenging (expressed as Trolox equivalents (18.4±1.1) mol/g) and potent cytotoxic activities against adenocarcinomic human alveolar basal epithelial (A549) cells (IC50=(58.2±6.8) µg/mL). High molecular mass polysaccharides, proteins and protein-polysaccharide complexes could have contributed to the antioxidant and cytotoxic selectivity of the OCS02®. LC-MS/MS analysis identified several potential cytotoxic proteases and an oxalate decarboxylase protein which may exhibit protection effects on kidneys. NOVELTY AND SCIENTIFIC CONTRIBUTIONS: The findings demonstrate the potential of OCS02® to be developed into functional food due to its promising superoxide anion radical scavenging capacity, cytotoxic effect and presence of biopharmaceutically active proteins.
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Sclerotial powder of a cultivated species of the Tiger Milk Mushroom, Lignosus cameronensis was analysed for its nutritional components and compared against species of the same genus, Lignosus rhinocerus and Lignosus tigris. All three species have been used by indigenous tribes in Peninsular Malaysia as medicinal mushrooms. Content of carbohydrate, fibre, mineral, amino acid, palatable index, fat, ash and moisture were determined. L. cameronensis sclerotial material consists of carbohydrate (79.7%), protein (12.4%) and dietary fibre (5.4%) with low fat (1.7%) and no free sugar. It has the highest content of total carbohydrate (791 g kg-1 ), energy value (3,700 kcal kg-1 ) and calcium (0.85 g kg-1 ). The crude protein content (123 g kg-1 ) is comparable to that of L. rhinocerus with its main amino acids consisting of glutamic acid, aspartic acid and leucine. The umami index is determined to be 0.27. The total essential amino acid (45 g kg-1 ) is comparable to that of L. tigris. The main mineral is potassium (1.51 g kg-1 ) and the Na/K ratio was <0.6. Heavy metals such as mercury, cadmium, lead and arsenic were absent. L. cameronensis has the highest amount of food energy, total carbohydrate and calcium compared to those of both L. rhinocerus and L. tigris. The essential amino acids comprised almost 40% of the total amino acid content, slightly more than that reported from sclerotial powder of the L. tigris. © 2019 IUBMB Life, 9999(9999):1-6, 2019.
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Aminoácidos/análisis , Carbohidratos/análisis , Metales Pesados/análisis , Minerales/análisis , Nutrientes/análisis , Plantas Medicinales/metabolismo , Polyporaceae/metabolismo , Humanos , Valor Nutritivo , Plantas Medicinales/crecimiento & desarrollo , Polyporaceae/crecimiento & desarrolloRESUMEN
BACKGROUND: Genome mining facilitated by heterologous systems is an emerging approach to access the chemical diversity encoded in basidiomycete genomes. In this study, three sesquiterpene synthase genes, GME3634, GME3638, and GME9210, which were highly expressed in the sclerotium of the medicinal mushroom Lignosus rhinocerotis, were cloned and heterologously expressed in a yeast system. RESULTS: Metabolite profile analysis of the yeast culture extracts by GC-MS showed the production of several sesquiterpene alcohols (C15H26O), including cadinols and germacrene D-4-ol as major products. Other detected sesquiterpenes include selina-6-en-4-ol, ß-elemene, ß-cubebene, and cedrene. Two purified major compounds namely (+)-torreyol and α-cadinol synthesised by GME3638 and GME3634 respectively, are stereoisomers and their chemical structures were confirmed by 1H and 13C NMR. Phylogenetic analysis revealed that GME3638 and GME3634 are a pair of orthologues, and are grouped together with terpene synthases that synthesise cadinenes and related sesquiterpenes. (+)-Torreyol and α-cadinol were tested against a panel of human cancer cell lines and the latter was found to exhibit selective potent cytotoxicity in breast adenocarcinoma cells (MCF7) with IC50 value of 3.5 ± 0.58 µg/ml while α-cadinol is less active (IC50 = 18.0 ± 3.27 µg/ml). CONCLUSIONS: This demonstrates that yeast-based genome mining, guided by transcriptomics, is a promising approach for uncovering bioactive compounds from medicinal mushrooms.
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Agaricales/química , Antineoplásicos/metabolismo , Saccharomyces cerevisiae/metabolismo , Sesquiterpenos/metabolismo , Agaricales/genética , Transferasas Alquil y Aril/genética , Transferasas Alquil y Aril/metabolismo , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Estructura Molecular , Saccharomyces cerevisiae/genética , Sesquiterpenos/química , Sesquiterpenos/farmacología , Relación Estructura-ActividadRESUMEN
Lignosus rhinocerotis (Cooke) Ryvarden (Polyporales, Basidiomycota), also known as the tiger milk mushroom, has received much interest in recent years owing to its wide-range ethnobotanical uses and the recent success in its domestication. The sclerotium is the part with medicinal value. Using two-dimensional gel electrophoresis coupled with mass spectrometry analysis, a total of 16 non-redundant, major proteins were identified with high confidence level in L. rhinocerotis sclerotium based on its genome as custom mapping database. Some of these proteins, such as the putative lectins, immunomodulatory proteins, superoxide dismutase, and aegerolysin may have pharmaceutical potential; while others are involved in nutrient mobilization and the protective antioxidant mechanism in the sclerotium. The findings from this study provide a molecular basis for future research on potential pharmacologically active proteins of L. rhinocerotis.
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Proteínas Fúngicas/aislamiento & purificación , Proteínas Fúngicas/farmacología , Polyporaceae/química , Proteómica/métodos , Antioxidantes/aislamiento & purificación , Antioxidantes/farmacología , Electroforesis en Gel Bidimensional , Proteínas Hemolisinas/farmacología , Espectrometría de Masas/métodos , Sistemas de Lectura Abierta , Plantas Medicinales/química , Polyporaceae/genética , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
BACKGROUND: The sclerotium of Lignosus rhinocerotis (Cooke) Ryvarden or Tiger milk mushroom (Polyporales, Basidiomycota) is a valuable folk medicine for indigenous peoples in Southeast Asia. Despite the increasing interest in this ethnobotanical mushroom, very little is known about the molecular and genetic basis of its medicinal and nutraceutical properties. RESULTS: The de novo assembled 34.3 Mb L. rhinocerotis genome encodes 10,742 putative genes with 84.30% of them having detectable sequence similarities to others available in public databases. Phylogenetic analysis revealed a close evolutionary relationship of L. rhinocerotis to Ganoderma lucidum, Dichomitus squalens, and Trametes versicolor in the core polyporoid clade. The L. rhinocerotis genome encodes a repertoire of enzymes engaged in carbohydrate and glycoconjugate metabolism, along with cytochrome P450s, putative bioactive proteins (lectins and fungal immunomodulatory proteins) and laccases. Other genes annotated include those encoding key enzymes for secondary metabolite biosynthesis, including those from polyketide, nonribosomal peptide, and triterpenoid pathways. Among them, the L. rhinocerotis genome is particularly enriched with sesquiterpenoid biosynthesis genes. CONCLUSIONS: The genome content of L. rhinocerotis provides insights into the genetic basis of its reported medicinal properties as well as serving as a platform to further characterize putative bioactive proteins and secondary metabolite pathway enzymes and as a reference for comparative genomics of polyporoid fungi.
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Genómica , Medicina Tradicional de Asia Oriental , Polyporales/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Genes Fúngicos/genética , Filogenia , Polyporales/metabolismo , Polisacáridos/genética , Polisacáridos/metabolismoRESUMEN
The Lignosus is a genus of fungi that have useful medicinal properties. In Southeast Asia, three species of Lignosus (locally known collectively as Tiger milk mushrooms) have been reported including L. tigris, L. rhinocerotis, and L. cameronensis. All three have been used as important medicinal mushrooms by the natives of Peninsular Malaysia. In this work, the nutritional composition and antioxidant activities of the wild type and a cultivated strain of L. tigris sclerotial extracts were investigated. The sclerotia are rich in carbohydrates with moderate amount of protein and low fat content. Free radical scavenging activities of L. tigris sclerotial extracts correlate with their phenolic content, which ranges from 6.25 to 45.42 mg GAE/g extract. The FRAP values ranged from 0.002 to 0.041 mmol/min/g extract, while the DPPH(â¢), ABTS(â¢+), and superoxide anion (SOA) scavenging activities ranged from 0.18 to 2.53, 0.01 to 0.36, and -4.53 to 10.05 mmol Trolox equivalents/g extract, respectively. L. tigris cultivar shows good prospect to be developed into functional food due to its good nutritional value and potent SOA scavenging activity.
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Antioxidantes/administración & dosificación , Fenoles/metabolismo , Polyporaceae/química , Superóxidos/metabolismo , Agaricales/química , Antioxidantes/química , Depuradores de Radicales Libres/metabolismo , Fenoles/químicaRESUMEN
BACKGROUND: The sclerotium of Lignosus rhinocerotis (Cooke) Ryvarden (Tiger Milk mushroom) is used as a traditional medicine to relieve cough, asthma and chronic hepatitis. The traditional uses of the sclerotium are presumably related to its anti-inflammatory effect. The present study was carried out to evaluate the anti-inflammatory activity of the sclerotial powder of L. rhinocerotis (Cooke) Ryvarden (Tiger Milk mushroom) cultivar TM02. METHODS: The anti-acute inflammatory activity of the sclerotial powder of L. rhinocerotis cultivar TM02 was investigated using carrageenan-induced paw edema test while the inhibition of transudative and proliferative phases of chronic inflammation were studied by cotton pellet induced granuloma model. Sprague Dawley rats were used in both studies. The anti-inflammatory activity was also measured by inhibition of lipopolysaccharide induced TNF-alpha production in RAW 264.7 macrophage cells. RESULTS: Cold water extract (CWE), hot water extract (HWE) and methanol extract (ME) of the sclerotial powder of L. rhinocerotis cultivar TM02 possessed anti-acute inflammatory activity as was measured by carrageenan-induced paw edema test, with CWE being the most potent. The acute anti-inflammatory activity of the cold water extract (CWE) was mainly contributed by its high molecular weight (HMW) fraction isolated by Sephadex G50 gel filtration chromatography. Its protein component was very potent in the inhibition of TNF-alpha production with an IC50 of 0.76 µg/ml. CWE at 200 mg/kg did not inhibit transudative and proliferative phase of chronic inflammation as shown by using the cotton pellet induced granuloma model. CONCLUSIONS: Our results suggested that most of the bioactive substance(s) contributed to the acute-inflammatory activity of the sclerotial powder of L. rhinocerotis cultivar TM02 appear to be in the CWE, particularly its HMW fraction. The anti-inflammatory activity of CWE was mainly contributed by the protein component of the high molecular weight (HMW) fraction and it exhibited strong inhibitory effect on TNF-alpha production but the possibility of synergistic effect between HMW, MMW and LMW fractions cannot be excluded. Future studies will provide new insights into the anti-inflammatory activity of the sclerotial powder.
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Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Micelio/química , Polyporaceae/química , Animales , Antiinflamatorios/química , Carragenina/toxicidad , Línea Celular , Edema/inducido químicamente , Edema/tratamiento farmacológico , Granuloma , Masculino , Ratones , Ratas , Ratas Sprague-Dawley , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Lignosus cameronensis holds promise for exploration given its morphological likeness to the renowned Lignosus rhinocerus, or tiger milk mushroom. Investigating its potential medicinal and industrial applications addresses a significant knowledge gap in this field. A comparative analysis with other Lignosus species and cultivars provides insights into biopharmacological potential. L. cameronensis cold water extract (LC-CWE) displayed moderate antioxidant activity, demonstrating promising Trolox equivalent antioxidant -capacity. Variable cytotoxicity was observed in different cell lines, with an IC50 of 215 µg/ml against breast cancer cells (MCF-7) cells. LC-CWE exhibited anti-inflammatory potential with an ED50 of 60 mg/kg in a λ-carrageenan-induced rat paw oedema model. Comparison with other Lignosus species and cultivars emphasised LC-CWE's distinct attributes, including high phenolic content and moderate antioxidant capacity. LC-CWE displayed potential in selectively inhibiting MCF-7 cells and reducing inflammation, highlighting its medicinal promise. This research expands our understanding of L. cameronensis and underscores the need for further mechanistic exploration.
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ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal mushrooms belonging to the Lignosus spp., colloquially known as Tiger Milk mushrooms (TMMs), are used as traditional medicine by communities across various regions of China and Southeast Asia to enhance immunity and to treat various diseases. At present, three Lignosus species have been identified in Malaysia: L. rhinocerus, L. tigris, and L. cameronensis. Similarities in their macroscopic morphologies and the nearly indistinguishable appearance of their sclerotia often lead to interchangeability between them. Hence, substantiation of their traditional applications via identification of their individual bioactive properties is imperative in ensuring that they are safe for consumption. L. tigris was first identified in 2013. Thus far, studies on L. tigris cultivar sclerotia (Ligno TG-K) have shown that it possesses significant antioxidant activities and has greater antiproliferative action against selected cancer cells in vitro compared to its sister species, L. rhinocerus TM02®. Our previous genomics study also revealed significant genetic dissimilarities between them. Further omics investigations on Ligno TG-K hold immense potential in facilitating the identification of its bioactive compounds and their associated bioactivities. AIM OF STUDY: The overall aim of this study was to investigate the gene expression profile of Ligno TG-K via de novo RNA-seq and pathway analysis. We also aimed to identify highly expressed genes encoding compounds that contribute to its cytotoxic and antioxidant properties, as well as perform a comparative transcriptomics analysis between Ligno TG-K and its sister species, L. rhinocerus TM02®. MATERIALS AND METHODS: Total RNA from fresh 3-month-old cultivated L. tigris sclerotia (Ligno TG-K) was extracted and analyzed via de novo RNA sequencing. Expressed genes were analyzed using InterPro and NCBI-Nr databases for domain identification and homology search. Functional categorization based on gene functions and pathways was performed using Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and Clusters of Orthologous Genes (COG) databases. Selected genes were subsequently subjected to phylogenetic analysis. RESULTS: Our transcriptomics analysis of Ligno TG-K revealed that 68.06% of its genes are expressed in the sclerotium; 80.38% of these were coding transcripts. Our analysis identified highly expressed transcripts encoding proteins with prospective medicinal properties. These included serine proteases (FPKM = 7356.68), deoxyribonucleases (FPKM = 3777.98), lectins (FPKM = 3690.87), and fungal immunomodulatory proteins (FPKM = 2337.84), all of which have known associations with anticancer activities. Transcripts linked to proteins with antioxidant activities, such as superoxide dismutase (FPKM = 1161.69) and catalase (FPKM = 1905.83), were also highly expressed. Results of our sequence alignments revealed that these genes and their orthologs can be found in other mushrooms. They exhibit significant sequence similarities, suggesting possible parallels in their anticancer and antioxidant bioactivities. CONCLUSION: This study is the first to provide a reference transcriptome profile of genes expressed in the sclerotia of L. tigris. The current study also presents distinct COG profiles of highly expressed genes in Ligno TG-K and L. rhinocerus TM02®, highlighting that any distinctions uncovered may be attributed to their interspecies variations and inherent characteristics that are unique to each species. Our findings suggest that Ligno TG-K contains bioactive compounds with prospective medicinal properties that warrant further investigations. CLASSIFICATION: Systems biology and omics.
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Agaricales , Polyporaceae , Antioxidantes/metabolismo , Transcriptoma , RNA-Seq , Agaricales/genética , Filogenia , Estudios Prospectivos , Polyporaceae/genéticaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Lignosus rhinocerus (Cooke) Ryvarden (also known as Tiger Milk mushroom, TMM), is a basidiomycete belonging to the Polyporaceae family. It has been documented to be used by traditional Chinese physicians and indigenous people in Southeast Asia to treat a variety of illnesses, such as gastritis, arthritis, and respiratory conditions, as well as to restore patients' physical well-being. TMM has also been used in folk medicine to treat cancer. For example, people from the indigenous Kensiu tribe of northeast Kedah (Malaysia) apply shredded TMM sclerotium mixed with water directly onto breast skin to treat breast cancer, while Chinese practitioners from Hong Kong, China prescribe TMM sclerotium as a treatment for liver cancer. L. rhinocerus has previously been demonstrated to possess selective anti-proliferative properties in vitro, however pre-clinical in vivo research has not yet been conducted. AIM OF STUDY: This study aimed to examine the anti-tumor activities of L. rhinocerus TM02®, using two different sample preparations [cold water extract (CWE) and fraction] via various routes of administration (oral and intraperitoneal) on an MCF7-xenograft nude mouse model. This study also investigated the inhibitory effect of TM02® CWE and its fractions against COX-2 in vitro using LPS-induced RAW264.7 macrophages, on the basis of the relationship between COX-2 and metastasis, apoptosis resistance, as well as the proliferation of cancer cells. MATERIALS AND METHODS: The first preparation, L. rhinocerus TM02® sclerotium powder (TSP) was dissolved in cold water to obtain the cold water extract (CWE). It was further fractionated based on its molecular weight to obtain the high (HMW), medium (MMW) and low (LMW) molecular weight fractions. The second preparation, known as the TM02® rhinoprolycan fraction (TRF), was obtained by combining the HMW and MMW fractions. TSP was given orally to mimic the daily consumption of a supplement; TRF was administered intraperitoneally to mimic typical tumorous cancer treatment with a rapid and more thorough absorption through the peritoneal cavity. Another experiment was conducted to examine changes in COX-2 activity in LPS-induced RAW264.7 macrophages after a 1-h pre-treatment with CWE, HMW, and MMW. RESULTS: Our results revealed that intraperitoneal TRF-injection (90 µg/g BW) for 20 days reduced initial tumor volume by â¼64.3% (n = 5). The percentage of apoptotic cells was marginally higher in TRF-treated mice vs. control, suggesting that induction of apoptosis as one of the factors that led to tumor shrinkage. TSP (500 µg/g BW) oral treatment (n = 5) for 63 days (inclusive of pre-treatment prior to tumor inoculation) effectively inhibited tumor growth. Four of the five tumors totally regressed, demonstrating the effectiveness of TSP ingestion in suppressing tumor growth. Although no significant changes were found in mouse serum cytokines (TNF-α, IL-5, IL-6 and CCL2), some increasing and decreasing trends were observed. This may suggest the immunomodulatory potential of these treatments that can directly or indirectly affect tumor growth. Pre-treatment with CWE, HMW and MMW significantly reduced COX-2 activity in RAW264.7 macrophages upon 24 h LPS-stimulation, suggesting the potential of L. rhinocerus TM02® extract and fractions in regulating M1/M2 polarization. CONCLUSION: Based on the findings of our investigation, both the rhinoprolycan fraction and crude sclerotial powder from L. rhinocerus TM02® demonstrated tumor suppressive effects, indicating that they contain substances with strong anticancer potential. The antitumor effects of L. rhinocerus TM02® in our study highlights the potential for further explorations into its mechanism of action and future development as a prophylactic or adjunct therapeutic against tumorous cancer.
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Lipopolisacáridos , Polyporaceae , Humanos , Ratones , Animales , Ratones Desnudos , Polvos , Ciclooxigenasa 2 , XenoinjertosRESUMEN
Lignosus rhinocerus has bioactive properties, such as anti-inflammatory, antioxidant, antiglycation, and immunomodulatory effects, that are beneficial to the skin. Gas chromatography-mass spectrometry analysis revealed the presence of cosmeceutical compounds, including linoleates, ascorbic acid dipalmitate, and all-trans-squalene, in the mushroom sclerotium. We further investigated the depigmentation and antiaging activities of L. rhinocerus cultivar TM02 sclerotia cold water extract (TM-CWE). TM-CWE at 10% (v/v) inhibited melanin production at the intra- and extracellular levels of B16-F1 melanoma cells (50.6-53.3%) and downregulated microphthalmia-associated transcription factor, the key protein for melanogenic enzyme expression, at the intracellular level (44.5 ± 2.6%). The extract also promoted human procollagen type 1 production in HaCat cells by 51.30 ± 2.87% at 12.5% (v/v). Irritation tests using three-dimensional eye and skin models further indicated that TM-CWE is a nonirritant with > 75% viability. Therefore, L. rhinocerus TM02 demonstrated the capacity to be a beneficial ingredient for skin care application.
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Agaricales , Ascomicetos , Colágeno , Humanos , Melaninas , Polyporaceae , AguaRESUMEN
COVID-19 infection has been a key threat to the public health system globally, with an estimated 248 million cases worldwide. COVID-19 patients are subject to a higher risk of developing chronic respiratory disorders that are closely associated with long-term disability, multi-morbidity, and premature mortality. Although there have been recent advancements in respiratory treatment regimens, there has also been increased interest in the use of medicinal mushrooms in bridging the unaddressed pathways of action within the treatment algorithms. In this review, we provide a collection of medicinal mushrooms that are beneficial in promoting respiratory health and potentially reducing COVID-19 symptoms in patients who are newly diagnosed and those who have recovered. While reviewing the use of immunomodulatory pathways, which have shown promising results in tackling side effects and post-COVID syndromes, we also provide insights into how the antioxidant elements present in medicinal mushrooms help to achieve the same results, especially in the prophylactic and therapeutic management of COVID-19 infection. To date, medicinal mushrooms are regarded as a functional food, which, however, need further quality, safety, and efficacy assessments. These requirements are also highlighted in the present review to promote the future development and application of medicinal mushrooms for better respiratory health.
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Agaricales , Tratamiento Farmacológico de COVID-19 , COVID-19 , Fitoterapia , Humanos , COVID-19/epidemiología , PandemiasRESUMEN
Ophiocordyceps sinensis is a popular medicinal mushroom used for various health conditions, including alleviation of frequent urination, which is a major symptom of overactive bladder (OAB) syndrome. This study aimed to investigate the effect of O. sinensis (OCS02 cultivar) cold-water extract (CWE) against bladder contractility using the organ bath technique. The bladder was removed from male Sprague-Dawley rats and cut into longitudinal strips of 2 mm × 8 mm. In some experiments, the urothelium was removed to study its role in CWE-induced responses. CWE elicited a biphasic response consisting of an immediate, transient contraction that was followed by a sustained relaxation in bladder strips precontracted with carbachol, a muscarinic agonist. Removal of urothelium did not alter the magnitude of the contractile response but significantly attenuated the relaxation response. In the presence of L-NAME (nitric oxide synthase inhibitor) and sodium nitroprusside (nitric oxide donor), CWE-induced transient contraction was enhanced, whereas the relaxation response was significantly reduced. Following preincubation with CWE, the amplitude and the frequency of the spontaneous myogenic contractions induced by carbachol, as well as the contractile response toward calcium, were significantly suppressed. Findings from this study show that the urothelium plays a role in the relaxant effect of CWE. Its mechanisms of action include the regulation of nitric oxide and inhibition of calcium influx.
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Cordyceps , Vejiga Urinaria , Animales , Calcio/farmacología , Carbacol/farmacología , China , Masculino , Músculo Liso/fisiología , Ratas , Ratas Sprague-Dawley , Vejiga Urinaria/fisiología , AguaRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Lignosus rhinocerotis (Cooke) Ryvarden cultivar TM02, also known as tiger's milk mushroom, is regarded as important folk medicine in Malaysia, while is used for the treatment of liver cancer, chronic hepatitis, gastric ulcer in traditional Chinese medicine. However, there is no compilation of scientific evidence that its protection for gastric, and no attempts have been made to understand how polysaccharides in Lignosus rhinocerotis might promote intestinal mucosal wound healing. AIM OF THE STUDY: This study aimed to investigate the effect and mechanism of ß-glucan prepared from L. rhinocerotis using an enzymatic method on epithelial restitution during intestinal mucosal damage. MATERIALS AND METHODS: Based on FT-IR, MALDI-TOF-MS, HPSEC-MALLS-RID, and AFM, the structure of polysaccharides from L. rhinocerotis was analysed. In addition, polysaccharides were used to test for wound healing activity in IEC-6 cells by measuring cell migration, proliferation, and expression of cell division control protein 42, Rac-1, RhoA, and Par-3. RESULTS: ß-glucan was extracted using enzyme-assisted extraction, and a yield of approximately 8.5 ± 0.8% was obtained from the dried biomass. The ß-glucan extracted by enzyme-assisted extraction (EAE) of polysaccharides was composed entirely of D-glucose with a total carbohydrate content of 95.5 ± 3.2%. The results of HPLC, FTIR, and MALDI-TOF-MS analyses revealed EAEP to be confirmed as ß-glucan. The molecular weight of prepared ß-glucan was found to be 5.315 × 104 g/mol by HPSEC-MALLS-RID. Furthermore, mucosal wound healing studies showed that the treatment of IEC-6 with a ß-glucan concentration of 200 µg/mL promoted cell migration and proliferation, and it enhanced the protein expression of cell division control protein 42, Rac-1, RhoA, and Par-3. CONCLUSIONS: The present study reveals that the prepared ß-glucan accelerates intestinal epithelial cell proliferation and migration via activation of Rho-dependent pathway. Hence, ß-glucan can be employed as a prospective therapeutic agent for the treatment of diseases associated with gastrointestinal mucosal damage, such as peptic ulcers and inflammatory bowel disease.
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Células Epiteliales/efectos de los fármacos , Mucosa Intestinal/efectos de los fármacos , Polyporaceae/química , Cicatrización de Heridas/efectos de los fármacos , beta-Glucanos/farmacología , Animales , Línea Celular , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Malasia , Medicina Tradicional de Asia Oriental , Ratas , beta-Glucanos/análisis , beta-Glucanos/química , Proteínas de Unión al GTP rho/metabolismoRESUMEN
BACKGROUND: Lignosus tigris, a recently discovered species of the unique Lignosus family, has been traditionally used by the indigenous communities in Peninsular Malaysia to treat various ailments and as an alternative medicine for cancer treatment. The L. tigris cultivar sclerotia (Ligno TG-K) was found to contain numerous bioactive compounds with beneficial biomedicinal properties and the sclerotial extract exhibited potent antioxidant activity. However, the anticancer property of the Ligno TG-K including in vitro and in vivo antitumor effects as well as its anticancer active compounds and the mechanisms has yet to be investigated. METHODS: The cytotoxicity of the Ligno TG-K against human breast (MCF7), prostate (PC3) and lung (A549) adenocarcinoma cell lines was evaluated using MTT cytotoxicity assay. The cytotoxic mechanisms of the active high molecular weight proteins (HMWp) fraction were investigated through detection of caspases activity and apoptotic-related proteins expression by Western blotting. The in vivo antitumor activity of the isolated HMWp was examined using MCF7 mouse xenograft model. Shotgun LC-MS/MS analysis was performed to identify the proteins in the HMWp. RESULTS AND DISCUSSION: Cold water extract of the sclerotia inhibited proliferation of MCF7, A549 and PC3 cells with IC50 ranged from 28.9 to 95.0 µg/mL. Bioassay guided fractionation of the extract revealed that HMWp exhibited selective cytotoxicity against MCF7 cells via induction of cellular apoptosis by the activation of extrinsic and intrinsic signaling pathways. HMWp activated expression of caspase-8 and -9 enzymes, and pro-apoptotic Bax protein whilst inhibiting expression of tumor survivor protein, Bcl-2. HMWp induced tumor-cell apoptosis and suppressed growth of tumor in MCF-7 xenograft mice. Lectins, serine proteases, RNase Gf29 and a 230NA deoxyribonuclease are the major cytotoxic proteins that accounted for 55.93% of the HMWp. CONCLUSION: The findings from this study provided scientific evidences to support the traditional use of the L. tigris sclerotia for treatment of breast cancer. Several cytotoxic proteins with high abundance have been identified in the HMWp of the sclerotial extract and these proteins have potential to be developed into new anticancer agents or as adjunct cancer therapy.
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Ophiocordyceps sinensis (=Cordyceps sinensis) has been known for its various medicinal properties, in particular immunomodulatory activities associated with its polysaccharides. In this study, the fruiting body of O. sinensis cultivar OCS02® was investigated for its chemical composition and monosaccharide profile. Cold water extract (CWE) obtained from this fruiting body was fractionated by molecular weight (MW) into high (HMW), medium (MMW), and low (LMW) fractions. Polysaccharides in the extract and fractions were identified as heteroglycans containing mostly glucose and mannose with small amounts of galactose, fucose, arabinose, and xylose. The immunomodulatory potential of these heteroglycans was evaluated by induction of cytokine/chemokine secretion using murine macrophage RAW 264.7. All treatments showed significant modulation of IL-6, IL-9, MIP-2, and TIMP-1, especially for CWE, HMW, and MMW, which might be due to their high ratios of glucose and the presence of protein. Further investigation on the structure-function relationship of these fruiting body polysaccharide fractions is needed to delineate the underlying mechanism of their immunomodulatory effect both in vitro and in vivo.
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Agaricales/química , Cuerpos Fructíferos de los Hongos/química , Hypocreales/química , Factores Inmunológicos/farmacología , Extractos Vegetales/farmacología , Polisacáridos/farmacología , Animales , China , Citocinas/inmunología , Factores Inmunológicos/química , Factores Inmunológicos/aislamiento & purificación , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Ratones , Peso Molecular , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Polisacáridos/química , Polisacáridos/aislamiento & purificación , Células RAW 264.7RESUMEN
A cultivar of fruiting bodies of Ophiocordyceps sinensis (FBOS; OCS02) was analyzed for nutrients, bioactive compounds, and heavy metal content to showcase its potential as a competitive, sustainable, and safe alternative to wild types and other cultivars. A previous 28-day subacute toxicity study showed that doses up to 1 g · kg-1 did not cause any adverse effects in Sprague-Dawley rats. The OCS02 cultivar contained large amounts of cordycepin, polysaccharides, and essential and semi-essential amino acids (0.66, 482.80, 99.02, and 101.04 g · kg-1, respectively) compared with levels reported in wild types and in cultivated mycelia. ß-1,3/1,6-glucan content was considerably high at 342.50 g · kg-1. The potassium level (5.14 g kg-1) tied in well with the low sodium content (0.121 g · kg-1)-6 times lower than amounts in wild types. We found no detectable levels of heavy metals such as lead, arsenic, cadmium, and mercury. The major amino acids found in FBOS (0CS02 cultivar) were arginine, lysine, serine, and threonine at 45.20, 20.30, 18.60, and 18.20 g · kg-1, respectively. The cultivated FBOS (OCS02 cultivar) is a comparable alternative to wild-type and other cultivated strains of O. sinensis. It has potential as a nutraceutical to meet market demand.
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Ascomicetos/química , Alimentos , Cuerpos Fructíferos de los Hongos/química , Aminoácidos/análisis , Cromatografía de Gases , Desoxiadenosinas/análisis , Metales Pesados/análisis , Micelio/química , Micelio/crecimiento & desarrollo , Polisacáridos/análisisRESUMEN
BACKGROUND: The highly valued medicinal tiger milk mushroom (also known as Lignosus rhinocerus) has the ability to cure numerous ailments. Its anticancer activities are well explored, and recently a partially purified cytotoxic protein fraction termed F5 from the mushroom's sclerotial cold water extract consisting mainly of fungal serine proteases was found to exhibit potent selective cytotoxicity against a human breast adenocarcinoma cell line (MCF7) with IC50 value of 3.00 µg/ml. However, characterization of its cell death-inducing activity has yet to be established. METHODS: The mechanism involved in the cytotoxic activities of F5 against MCF7 cells was elucidated by flow cytometry-based apoptosis detection, caspases activity measurement, and expression profiling of apoptosis markers by western blotting. Molecular attributes of F5 were further mined from L. rhinocerus's published genome and transcriptome for future exploration. RESULTS AND DISCUSSION: Apoptosis induction in MCF7 cells by F5 may involve a cross-talk between the extrinsic and intrinsic apoptotic pathways with upregulation of caspase-8 and -9 activities and a marked decrease of Bcl-2. On the other hand, the levels of pro-apoptotic Bax, BID, and cleaved BID were increased accompanied by observable actin cleavage. At gene level, F5 composed of three predicted non-synonymous single nucleotide polymorphisms (T > C) and an alternative 5' splice site. CONCLUSIONS: Findings from this study provide an advanced framework for further investigations on cancer therapeutics development from L. rhinocerus.
RESUMEN
Naturally occurring anti-glycation compounds have drawn much interest in recent years as they show potential in reducing or preventing the risk of chronic complications for diabetic patients. In this study, annotation of the genome-transcriptome data from tiger milk mushroom (Lignosus rhinocerus, syn. Lignosus rhinocerotis) to PlantCyc enzymes database identified transcripts that are related to anti-diabetic properties, and these include genes that are involved in carotenoid and abscisic acid biosynthesis as well as genes that code for glyoxalase I, catalase-peroxidases, and superoxide dismutases. The existence of these genes suggests that L. rhinocerus may contain bioactive compound(s) with anti-glycation properties that can be exploited for management of diabetic complications. A medium-molecular-weight (MMW) fraction which was obtained from a combination of cold water extraction and Sephadex® G-50 (fine) gel filtration chromatography of L. rhinocerus sclerotia powder was demonstrated to exhibit potent anti-glycation activity. The fraction specifically inhibited the formation of Nð-(carboxymethyl)lysine, pentosidine, and other advanced glycation end-product (AGE) structures in a human serum albumin-glucose system, with an IC50 value of 0.001 mg/ml, almost 520 times lower than that of the positive control, aminoguanidine hydrochloride (IC50 = 0.52 mg/ml). Its ability to suppress protein glycation may be partly associated with its strong superoxide anion radical scavenging activity (10.16 ± 0.12 mmol TE/g). Our results suggest that the MMW fraction of L. rhinocerus shows potential to be developed into a potent glycation inhibitor for preventing AGE-mediated diabetic complications.