Antidepressant-like effect of Ganoderma lucidum spore polysaccharide-peptide mediated by upregulation of prefrontal cortex brain-derived neurotrophic factor.

A 28-kDa polysaccharide-peptide (PGL) with antidepressant-like activities was isolated from spores of the mushroom Ganoderma lucidum. It was unadsorbed on DEAE-cellulose. Its internal amino acid sequences manifested pronounced similarity with proteins from the mushrooms Lentinula edodes and Agaricus bisporus. The monosaccharides present in 28-kDa PGL comprised predominantly of glucose (over 90%) and much fewer galactose, mannose residues, and other residues. PGL manifested antidepressant-like activities as follows. It enhanced viability and DNA content in corticosterone-injured PC12 cells(a cell line derived from a pheochromocytoma of the rat adrenal medulla with an embryonic origin from the neural crest containing a mixture of neuroblastic cells and eosinophilic cells) and reduced LDH release. A single acute PGL treatment shortened the duration of immobility of mice in both tail suspension and forced swimming tests. PGL treatment enhanced sucrose preference and shortened the duration of immobility in mice exposed to chronic unpredictable mild stress (CUMS). Chronic PGL treatment reversed the decline in mouse brain serotonin and norepinephrine levels but did not affect dopamine levels. PGL decreased serum corticosterone levels and increased BDNF mRNA and protein levels and increased synapsin I and PSD95 levels in the prefrontal cortex. This effect was completely blocked by pretreatment with the BDNF antagonist K252a, indicating that PGL increased synaptic proteins in a BDNF-dependent manner.Key points• An antidepressive polysaccharide-peptide PGL was isolated from G. lucidum spores.• PGL protected PC12 nerve cells from the toxicity of corticosterone.• PGL upregulated BDNF expression and influenced key factors in the prefrontal cortex.

Purification and properties of a laccase from the mushroom Agaricus sinodeliciosus.

A homogeneous monomeric laccase (ASL) from Agaricus sinodeliciosus, with a molecular mass of 65 kDa, was isolated using ion-exchange chromatography (CM-cellulose and Q-Sepharose) and gel-filtration chromatography (Superdex 75). This laccase exhibited maximum activity at 50 °C and pH 5.0. Hg2+ and Cd2+ significantly inhibited its activity. The laccase displayed a Km value of 0.9 mM toward 2,2'-azinobis-(3-ethylbenzthiazoline-6-sulfonate) (ABTS). In addition to ABTS, ASL exhibited higher affinity toward o-toluidine and benzidine than other substrates. ASL is able to decolorize malachite green and Eriochrome black T.

MAP30 protein from Momordica charantia is therapeutic and has synergic activity with cisplatin against ovarian cancer in vivo by altering metabolism and inducing ferroptosis.

Increasing evidence shows that Traditional Chinese Medicine (TCM) has an obvious appeal for cancer treatment, but there is still a lack of scientific investigation of its underlying molecular mechanisms. Bitter melon or bitter gourd (Momordica charantia) is an edible fruit that is commonly consumed, and it is used to cure different diseases in various ancient folk medical practices. We report that a bioactive protein, MAP30, isolated from bitter melon seeds exhibited potent anticancer and anti-chemoresistant effects on ovarian cancer cells. Functional studies revealed that MAP30 inhibited cancer cell migration, cell invasion, and cell proliferation in various ovarian cancer cells but not normal immortalized ovarian epithelial cells. When administered with cisplatin, MAP30 produced a synergistic effect on cisplatin-induced cell cytotoxicity in ovarian cancer cells. When low doses of cisplatin and MAP30 were co-injected intraperitoneally, a remarkable reduction of tumor dissemination and tumor growth was observed in an ovarian cancer ascites mouse model. Notably, blood tests confirmed that MAP30 did not cause any adverse effects on liver and kidney functions in the treated mice. MAP30 activated AMP-activated protein kinase (AMPK) signaling via CaMKKß and induced cell cycle arrest in the S-phase. MAP30 modulated cell metabolism of ovarian cancer cells via suppression of GLUT-1/-3-mediated glucose uptake, adipogenesis, and lipid droplet formation in tumor development and progression. MAP30 also induced an increase in intracellular Ca2+ ion concentration, which triggered ROS-mediated cancer cell death via apoptosis and ferroptosis. Collectively, these findings suggest that natural MAP30 is a non-toxic supplement that may enhance chemotherapeutic outcomes and benefit ovarian cancer patients with peritoneal metastases.

Traditional Chinese Medicine (TCM) and Allergic Diseases.

PURPOSE OF REVIEW: This paper purports to review recent relevant publications on the efficacy of traditional Chinese medicine in treating allergic diseases, to illustrate the pertinent mechanisms of action of TCM, and to explore the possible role of TCM in the management of allergic diseases in the foreseeable future. As TCM embodies multiple treatment modalities, only the most popular two, namely CHM (Chinese herbal medicine) and acupuncture, were discussed. Publications, especially reviews involving randomized controlled trials (RCTs) on the use of TCM on allergic diseases, published up to June 2019 were reviewed and analyzed. Papers reporting the mechanisms of action of TCM in allergic diseases were also included. Other publications in Chinese were also discussed. RECENT FINDINGS: A startling escalation in the incidence of allergic diseases in the last several decades has posed tremendous social and financial burdens on the community. Failing to locate a cure for these chronic diseases, patients have resorted to using alternative medications of which traditional Chinese medicine (TCM) is a popular one. Thus CHM has been extensively employed for treating allergic diseases. Some investigations have been conducted to ascertain the therapeutic efficacy of CHM for allergic diseases. Although CHM has been widely deployed for treating allergic diseases, it appears from the published data that there is a dearth of conclusive evidence to establish the effectiveness of CHM for allergic diseases. It is recommended that more large- scale RCTs with prolonged durations be carried out to corroborate the efficacy of CHM for allergic diseases. On the other hand, there is ample evidence indicating that acupuncture is useful when administered alone in allergic rhinitis and asthma or when applied as an adjunct to conventional western therapy. Evidence of its utility in atopic eczema and urticaria is not definitive. It is recommended that acupuncture be integrated into the therapy of allergic rhinitis and asthma, and that CHM be used as an adjunct in the treatment of allergic diseases on an individual basis.

Diversity of potentially exploitable pharmacological activities of the highly prized edible medicinal fungus Antrodia camphorata.

Antrodia camphorata, also known as A. cinnamomea, is a precious medicinal basidiomycete fungus endemic to Taiwan. This article summarizes the recent advances in research on the multifarious pharmacological effects of A. camphorata. The mushroom exhibits anticancer activity toward a large variety of cancers including breast, cervical, ovarian, prostate, bladder, colorectal, pancreatic, liver, and lung cancers; melanoma; leukemia; lymphoma; neuroblastoma; and glioblastoma. Other activities encompass antiinflammatory, antiatopic dermatitis, anticachexia, immunoregulatory, antiobesity, antidiabetic, antihyperlipidemic, antiatherosclerotic, antihypertensive, antiplatelet, antioxidative, antiphotodamaging, hepatoprotective, renoprotective, neuroprotective, testis protecting, antiasthmatic, osteogenic, osteoprotective, antiviral, antibacterial, and wound healing activities. This review aims to provide a reference for further development and utilization of this highly prized mushroom.

Purification of an Antifungal Peptide from Seeds of Brassica oleracea var. gongylodes and Investigation of Its Antifungal Activity and Mechanism of Action.

In this study, a 8.5-kDa antifungal peptide designated as BGAP was purified from the crude extract of the seeds of Brassica oleracea var. gongylodes by employing a protocol that comprised cation exchange chromatography on SP-Sepharose, cation exchange chromatography on Mono S and gel filtration chromatography on Superdex peptide. BGAP showed the highest amino acid sequence similarity to defensin peptides by mass spectrometric analysis. BGAP showed a broad spectrum of antifungal activity with a half maximal inhibitory concentration at 17.33 µg/mL, 12.37 µg/mL, 16.81 µg/mL, and 5.60 µg/mL toward Colletotrichum higginsianum, Exserohilum turcicum, Magnaporthe oryzae and Mycosphaerella arachidicola, respectively. The antifungal activity of BGAP remained stable (i) after heat treatment at 40-100 °C for 15 min; (ii) after exposure to solutions of pH 1-3 and 11-13 for 15 min; (iii) after incubation with solutions containing K⁺, Ca2+, Mg2+, Mn2+ or Fe3+ ions at the concentrations of 20-150 mmol/L for 2 h; and (iv) following treatment with 10% methyl alcohol, 10% ethanol, 10% isopropanol or 10% chloroform for 2 h. Fluorescence staining experiments showed that BGAP brought about an increase in cell membrane permeability, a rise in reactive oxygen species production, a decrease in mitochondrial membrane potential, and an accumulation of chitin at the hyphal tips of Mycosphaerella arachidicola.

CRISPR/Cas9 Technology Targeting Fas Gene Protects Mice From Concanavalin-A Induced Fulminant Hepatic Failure.

Fulminant hepatic failure is a life-threatening disease which occurs in patients without preexisting liver disease. Nowadays, there is no ideal therapeutic tool in the treatment of fulminant hepatic failure. Recent studies suggested that a novel technology termed CRISPR/Cas9 may be a promising approach for the treatment of fulminant hepatic failure. In this project, we have designed single chimeric guide RNAs specifically targeting the genomic regions of mouse Fas gene. The in vitro and in vivo effects of sgRNAs on the production of Fas protein were examined in cultured mouse cells and in a hydrodynamic injection-based mouse model, respectively. The in vivo delivery of CRISPR/Cas9 could maintain liver homeostasis and protect hepatocytes from Fas-mediated cell apoptosis in the fulminant hepatic failure model. Our study indicates the clinical potential of developing the CRISPR/Cas9 system as a novel therapeutic strategy to rescue Concanavalin-A-induced fulminant hepatic failure in the mouse model. J. Cell. Biochem. 118: 530-536, 2017. © 2016 Wiley Periodicals, Inc.

Chemical and ultrastructural studies of lignocellulose biodegradation during Agaricus bisporus cultivation.

During Agaricus bisporus cultivation, lignocellulose degradation is the result of the activity of both the mushroom and microbial communities developed during the composting. To investigate the lignocellulose degradation in detail from the beginning to the end of the process, the functional groups of cellulose, hemicellulose, and lignin have been studied with Fourier transform infrared spectroscopy and the morphological changes of lignocelluloses were elucidated with scanning electron microscopy. The aperture of lignin and cellulose increased to enable the mycelia of A. bisporus to penetrate into the medium and to degrade lignocelluloses in a more direct way. The chemical structure changes implied a preferential use of lignin that could make for better use of cellulose to boost growth of A. bisporus. Changes in chemical structure together with ultrastructural changes induced by the microbial flora during cultivation substrate production by the composting substrate are important in promoting the utilization of lignocelluloses by A. bisporus.

Purification and characterization of a ribonuclease with antiproliferative activity from the mystical wild mushroom Tuber indicum.

An RNase with a molecular mass of 28 kDa and with high ribonucleolytic activity toward poly(A) was purified from the ascocarps of Tuber indicum. The purification procedure involved ion exchange chromatography on diethylaminoethyl cellulose, Q-Sepharose and Mono Q, and gel filtration by fast protein liquid chromatography on Superdex 75. The pH and temperature optima of the RNase were 7.2 and 50 °C, respectively. The ranking of its activity toward various polyhomoribonucleotides was poly(A)>poly(C)>poly(G) ≈ poly(U). All of the metal ions used in this study, except for the K(+) ions, curtailed the activity of the RNase. The RNase activity was reduced by ethylene diamine tetraacetic acid (EDTA), dithiothreitol (DTT), and sodium dodecyl sulfate (SDS) by 42.2%, 75.5%, and 96.6%, respectively. The RNase inhibited the proliferation of hepatoma (HepG2) and human breast cancer cell lines (MCF7), with half-maximal inhibitory concentrations (IC50 ) of 12.6 and 16.6 µM, respectively.

Abnormal response of the proliferation and differentiation of growth plate chondrocytes to melatonin in adolescent idiopathic scoliosis.

Abnormalities in the melatonin signaling pathway and the involvement of melatonin receptor MT2 have been reported in patients with adolescent idiopathic scoliosis (AIS). Whether these abnormalities were involved in the systemic abnormal skeletal growth in AIS during the peripubertal period remain unknown. In this cross-sectional case-control study, growth plate chondrocytes (GPCs) were cultured from twenty AIS and ten normal control subjects. Although the MT2 receptor was identified in GPCs from both AIS and controls, its mRNA expression was significantly lower in AIS patients than the controls. GPCs were cultured in the presence of either the vehicle or various concentrations of melatonin, with or without the selective MT2 melatonin receptor antagonist 4-P-PDOT (10 µM). Then the cell viability and the mRNA expression of collagen type X (COLX) and alkaline phosphatase (ALP) were assessed by MTT and qPCR, respectively. In the control GPCs, melatonin at the concentrations of 1, 100 nM and 10 µM significantly reduced the population of viable cells, and the mRNA level of COLX and ALP compared to the vehicle. Similar changes were not observed in the presence of 4-P-PDOT. Further, neither proliferation nor differentiation of GPCs from AIS patients was affected by the melatonin treatment. These findings support the presence of a functional abnormality of the melatonin signaling pathway in AIS GPCs, which might be associated with the abnormal endochondral ossification in AIS patients.

Abnormal Skeletal Growth in Adolescent Idiopathic Scoliosis Is Associated with Abnormal Quantitative Expression of Melatonin Receptor, MT2.

The defect of the melatonin signaling pathway has been proposed to be one of the key etiopathogenic factors in adolescent idiopathic scoliosis (AIS). A previous report showed that melatonin receptor, MT2, was undetectable in some AIS girls. The present study aimed to investigate whether the abnormal MT2 expression in AIS is quantitative or qualitative. Cultured osteoblasts were obtained from 41 AIS girls and nine normal controls. Semi-quantification of protein expression by Western blot and mRNA expression by TaqMan real-time PCR for both MT1 and MT2 were performed. Anthropometric parameters were also compared and correlated with the protein expression and mRNA expression of the receptors. The results showed significantly lower protein and mRNA expression of MT2 in AIS girls compared with that in normal controls (p = 0.02 and p = 0.019, respectively). No differences were found in the expression of MT1. When dichotomizing the AIS girls according to their MT2 expression, the group with low expression was found to have a significantly longer arm span (p = 0.036). The results of this study showed for the first time a quantitative change of MT2 in AIS that was also correlated with abnormal arm span as part of abnormal systemic skeletal growth.

An autoimmunized mouse model recapitulates key features in the pathogenesis of Sjögren's syndrome.

The pathogenesis of Sjögren's syndrome (SS) is poorly understood. To evaluate an autoimmunization-induced experimental SS model, we firstly observed the phenotype of lymphocyte infiltration in the enlarged submandibular gland (SG). Furthermore, significant activation of caspase-3 and a high ratio of Bax-to-Bcl-2 were detected, indicating the inflammatory apoptosis associated with developmental foci. Meanwhile, the dysregulated cytokines, such as tumor necrosis factor α, IL-1ß and IL-6 mRNA expression, were found to be over-expressed. A progressive decrease of aquaporin 5 and its subcellular translocation from apical to basal membrane in SG was found to be associated with the abnormally expressed M3 muscarinic acetylcholine receptor. This pattern was found to be similar to that seen in human SS and possibly contributed to the saliva secretion deficiency. Thus, this autoimmunization-induced model recapitulates the key features of human SS and may have potential for studying the pathogenesis of human SS.

A laccase with HIV-1 reverse transcriptase inhibitory activity from the broth of mycelial culture of the mushroom Lentinus tigrinus.

A 59 kDa laccase with inhibitory activity against HIV-1 reverse transcriptase (IC(50) = 2.4 µM) was isolated from the broth of mycelial culture of the mushroom Lentinus tigrinus. The isolation procedure involved ion exchange chromatography on DEAE-cellulose and CM-cellulose, and gel filtration by fast protein liquid chromatography on Superdex 75. The laccase was adsorbed on both types of ion exchangers. About 95-fold purification was achieved with a 25.9% yield of the enzyme. The procedure resulted in a specific enzyme activity of 76.6 U/mg. Its N-terminal amino acid sequence was GIPDLHDLTV, which showed little similarity to other mushroom laccase and other Lentinus tigrinus strain laccase. Its characteristics were different from previously reported laccase of other Lentinus tigrinus strain. Maximal laccase activity was observed at a pH of 4 and at a temperature of 60°C, respectively. This study yielded the information about the potentially exploitable activities of Lentinus tigrinus laccase.

A laccase with antiproliferative and HIV-I reverse transcriptase inhibitory activities from the mycorrhizal fungus Agaricus placomyces.

A novel 68 kDa laccase was purified from the mycorrhizal fungus Agaricus placomyces by utilizing a procedure that comprised three successive steps of ion exchange chromatography and gel filtration as the final step. The monomeric enzyme exhibited the N-terminal amino acid sequence of DVIGPQAQVTLANQD, which showed only a low extent of homology to sequences of other fungal laccases. The optimal temperature for A. placomyces laccase was 30°C, and optimal pH values for laccase activity towards the substrates 2,7'-azinobis[3-ethylbenzothiazolone-6-sulfonic acid] diammonium salt (ABTS) and hydroquinone were 5.2 and 6.8, respectively. The laccase displayed, at 30°C and pH 5.2, K(m) values of 0.392 mM towards hydroquinone and 0.775 mM towards ABTS. It potently suppressed proliferation of MCF 7 human breast cancer cells and Hep G2 hepatoma cells and inhibited human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) activity with an IC(50) of 1.8 µM, 1.7 µM, and 1.25 µM, respectively, signifying that it is an antipathogenic protein.

Abnormal melatonin receptor 1B expression in osteoblasts from girls with adolescent idiopathic scoliosis.

Melatonin signaling dysfunction has been associated with the etiology of adolescent idiopathic scoliosis (AIS). Genetic analysis has also associated the occurrence of AIS with the MT2 gene. Thus, we determined whether there is abnormality in the protein expression of melatonin receptors (MT) in AIS osteoblasts. In this study, we recruited 11 girls with severe AIS and eight normal subjects for intraoperative bone biopsies. MT1 and MT2 receptor protein expressions in the isolated osteoblasts were detected. Also, cell proliferation assay using different melatonin concentrations (0, 10(-9), 10(-5), 10(-4) m) was carried out. The results showed that both MT1 and MT2 receptors are expressed in osteoblasts of the controls. While MT1 receptors were expressed in osteoblasts of all AIS subjects, osteoblasts of only 7 of 11 AIS showed expression of MT2 receptors. Melatonin stimulated control osteoblasts to proliferate. However, proliferation of AIS osteoblasts without expression of MT2 receptor, after treatment with melatonin, was minimal when compared with control and AIS osteoblasts with MT2 receptor expression. The proliferation of AIS osteoblasts with MT2 receptor was greater than those without. This is the first report demonstrating a difference between AIS and normal osteoblasts in the protein expression of MT2 receptor. The results suggest that there is a possible functional effect of MT2 receptor on osteoblast proliferation. AIS osteoblasts without expression of MT2 receptor showed the lowest percentage of viable cells after melatonin treatment. This possibly indicates the modulating role of melatonin through MT2 receptor on the proliferation of osteoblasts.

Abnormal proliferation and differentiation of osteoblasts from girls with adolescent idiopathic scoliosis to melatonin.

Melatonin deficiency has been postulated as an etiologic factors in adolescent idiopathic scoliosis (AIS). In previous studies, melatonin was shown to regulate skeletal growth and bone formation in both humans and rats. Although it remains controversial whether there are differences in serum melatonin level between AIS and control subjects, melatonin signaling pathway dysfunction in osteoblasts has been reported in patients with AIS. Recently, our group found that melatonin receptor 1B (MT2) gene polymorphism was associated with the occurrence of AIS. Hence, the present study investigated the effect of melatonin on AIS osteoblasts. In vitro assays were performed with osteoblasts isolated from 17 severe AIS girls and nine control subjects. The osteoblasts were exposed to different concentrations of melatonin for 3 days. The effects of melatonin on cell proliferation (as evidenced by MTT assay) and differentiation (demonstrated by alkaline phosphatase activity) were determined. In the control group, melatonin significantly stimulated osteoblasts to proliferate and differentiate. However, in the AIS group, the stimulatory effects of melatonin were not discernible. Importantly, this finding demonstrated that there is a significant difference between AIS and control osteoblasts in functional response toward melatonin. Melatonin-stimulated proliferation of control osteoblasts was inhibited by the MT2 antagonist, 4-phenyl-2-propionamidotetraline, as well as by luzindole, a nonselective melatonin receptor antagonist, suggesting that MT2 is associated with the proliferative action of melatonin. The lack of response in AIS osteoblasts might be because of dysfunction of the melatonin signaling pathway, which may contribute to the low bone mineral density and abnormal skeletal growth observed in patients with AIS.

Protective effect of bilberry (Vaccinium myrtillus L.) extracts on cultured human corneal limbal epithelial cells (HCLEC).

The use of bilberry (Vaccinium myrtillus L.) as a food and medicine for improving human vision has a long history all over the world. However, there is lack of convincing evidence from rigorous clinical trials or scientific research. This study investigated the effects of different concentrations of bilberry extracts on the cell viability, cell cycle and the expression of hyaluronic acid and glycosaminoglycans of cultured human corneal limbal epithelial cells. The data showed that bilberry extracts had no cytotoxicity to the corneal limbal epithelial cells at a wide range of concentrations (10(-9)-10(-4) M, equalized to the content of cyanidin-3-O-glucoside). Bilberry extract (10(-6), 10(-5) and 10(-4) M) increased cell viability after 48 h incubation. The number of cells decreased in G(0)/G(1) phase and increased prominently in S and G(2)/M phases after treatment with bilberry extracts at a high concentration (10(-4) M). The expression of glycosaminoglycans increased prominently after incubation with bilberry extracts (10(-7) and 10(-4) M) for 48 h while no significant changes were observed for the expression of hyaluronic acid. The results indicated that bilberry extract may be beneficial for the physiological renewal and homeostasis of corneal epithelial cells.

Purification and characterization of a laccase from the edible wild mushroom Tricholoma mongolicum.

A novel laccase from Tricholoma mongolicum was purified by using a procedure which entailed ion exchange chromatography on DEAE-cellulose, CM-cellulose, Q-Sepharose, and FPLC-gel filtration on Superdex 75. The purified enzyme was obtained with a specific activity of 1480 U/mg-protein and a final yield of 15%. It was found to be a monomeric protein with a molecular mass of 66 kDa as estimated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Its N-terminal amino acid sequence was GIGPVADLYVGNRIL, similar to some but different other mushroom laccase. The optimum pH and temperature for the purified enzyme were pH 2 to pH 3 and 30 degrees C, respectively. It displayed a low K(m) toward 2,7-azinobis (3-ethylbenzothiazolone-6-sulfonic acid) diammonium salt (ABTS) and high K(cat)/K(m) values. The purified laccase oxidized a wide range of lignin-related phenols, but exerted maximal activity on ABTS. It was significantly inhibited by Hg(2+) ions, and remarkably stimulated by Cu(2+) ions. It inhibited HIV-1 reverse transcriptase and proliferation of hepatoma HepG2 cells and breast cancer MCF7 cells with an IC50 of 0.65 microM, 1.4 microM, and 4.2 microM, respectively, indicating that it is also an antipathogenic protein.

Buckwheat Antifungal Protein with Biocontrol Potential To Inhibit Fungal ( Botrytis cinerea) Infection of Cherry Tomato.

A 11 kDa antifungal protein FEAP was purified from buckwheat ( Fagopyrum esculentum) seed extract with a procedure involving (NH4)2SO4 precipitation and chromatography on SP-Sepharose, Affi-gel blue gel, Mono S, and Superdex peptide. Its N-terminal sequence was AQXGAQGGGAT, resembling those of buckwheat peptides Fα-AMP1 and Fα-AMP2. FEAP exhibited thermostability (20-100 °C) and acid resistance (pH 1-5). Its antifungal activity was retained in the presence of 10-150 mmol/L of K+, Mn2+, or Fe3+ ions, 10-50 mmol/L of Ca2+ or Mg2+ ions, and 50% methanol, 50% ethanol, 50% isopropanol, or 50% chloroform. Its half-maximal inhibitory concentrations toward spore germination and mycelial growth in Botrytis cinerea were 79.9 and 236.7 µg/mL, respectively. Its antifungal activity was superior to the fungicide cymoxanil mancozeb (248.1 µg/mL). FEAP prevented B. cinerea from infecting excised leaves, intact leaves, and isolated fruits of cherry tomato. Its mechanism involved induction of an increase in cell membrane permeability and a decrease in mitochondrial membrane potential.