Comparative study on the anti-tumor effect of steroids derived from different organisms in H22 tumor-bearing mice and analysis of their mechanisms.

This study aimed to comparatively investigate the anti-tumor mechanisms of steroids including ergosterol, ß-sitosterol, cholesterol, and fucosterol. The model of H22 tumor-bearing mice was constructed based on histopathological data and biochemical parameters, while serums were subjected to metabolomics analysis to study the potential anti-tumor mechanisms. The results indicated that the four steroids exhibited different degrees of anti-tumor effects on H22 mice. The tumor inhibition rates were 63.25% for ergosterol, 56.41% for ß-sitosterol, 61.54% for cholesterol, and 72.65% for fucosterol. Metabolomic analyses revealed that 87, 71, and 129 differential metabolites were identified in ergosterol, cholesterol, and fucosterol treatment groups, respectively. The fucosterol treatment group had the highest number of differential metabolites. At the same time, it mainly inhibited purine and amino acid metabolism to exert anti-tumor effects. Ergosterol enhanced immunity and affected pyruvate metabolism, and cholesterol inhibited purine metabolism. The chemical structure difference among ergosterol, cholesterol, and fucosterol is mainly at the number and position of sterol double bonds and the number and length of side chain carbons. Therefore, there is a structure-activity relationship between the structure of steroid compounds and their efficacy. This study provides a key foundation for the exploitation of the anti-tumor effects of steroids derived from different organisms.

Genomic comparison between two Inonotus hispidus strains isolated from growing in different tree species.

Inonotus hispidus mainly growing in broad-leaved trees, including Morus alba, Fraxinus mandshurica, and Ulmus macrocarpa etc. The fruiting body of I. hispidus growing in M. alba (hereafter as MA) is used as a traditional Chinese medicine "Sanghuang". However, differences between the genetic material basis of I. hispidus growing in other tree species have not been reported. Therefore, in this paper, the genomic comparison between MA and I. hispidus growing in F. mandshurica (hereafter as FM) were studied. The whole genome of MA monokaryon was sequenced by Illumina combined with Pac Bio platform. Next, genome assembly, genome component prediction and genome functional annotation were performed. Comparative genomics analysis was performed between FM monokaryon and MA monokaryon, using MA as the reference. The results showed that, MA had 24 contigs with a N50 length of 2.6 Mb. Specifically, 5,342, 6,564, 1,595, 383 and 123 genes were annotated from GO, KEGG, KOG, CAZymes and CYP450, respectively. Moreover, comparative genomics showed that, the coding genes and total number of genes annotated in different databases of FM were higher than that of MA. This study provides a foundation for the medicinal application of FM as MA from the perspective of genetic composition.

Comparative transcriptome and proteome analysis explores the antitumor key regulators of ergosterone in H22 tumor-bearing mice.

Ergosterone has been proved to have potential antitumor effect on H22 tumor-bearing mice, but the antitumor mechanism and key regulators are still unclear. The current study was aimed to explore the key regulators responsible for antitumor of ergosterone using whole transcriptome and proteome analysis in H22 tumor-bearing mice model. The model of H22 tumor-bearing mice was constructed according to the histopathological data and biochemical parameters. The isolated tumor tissues of different treatment groups were subjected to transcriptomic and proteomic analysis. Our findings demonstrated that 472 differentially expressed genes and 658 proteins were identified in the tumor tissue of different treatment groups through RNA-Seq and liquid chromatography with tandem mass spectrometry-based proteomic analysis, respectively. The combined omics analysis revealed three critical genes/proteins, including Lars2, Sirpα and Hcls1 that could play a role in antitumor pathways. Furthermore, Lars2, Sirpα and Hcls1 genes/proteins, as key regulators of the antitumor effect of ergosterone, were verified by qRT-PCR and western blotting methods, respectively. In summary, our study provides new insights into analysing the antitumor mechanism of ergosterone from the point of view of gene and protein expression and will encourage further development of the antitumor pharmaceutical industry.

Systematic Characterization of Active Antitumor Constituents from the Shaggy Bracket Medicinal Mushroom Inonotus hispidus (Agaricomycetes) by UPLC-Q-TOF/MS and Network Pharmacology.

Inonotus hispidus is a well-known medicinal fungus and has been used in the treatment of cancer in China, but the material basis and potential mechanisms are still limited. The present study aimed to use in vitro experiments, UPLC-Q-TOF/MS and network pharmacology to predict active compounds and possible mechanisms of cultivated and wild I. hispidus. The cytotoxicity results in vitro showed that the extracts of cultivated and wild fruit bodies exhibited the highest inhibitory effects against MDA-MB-231 cells, and the 50% inhibition concentration, (IC50) values were 59.82 and 92.09 µg/mL, respectively. Of the two extracts, a total of 30 possible chemical components, including 21 polyphenols and nine fatty acids, were identified. Network pharmacology showed that five active polyphenols (osmundacetone, isohispidin, inotilone, hispolon, and inonotusin A) and 11 potential targets (HSP90AA1, AKT1, STAT3, EGFR, ESR1, PIK3CA, HIF1A, ERBB2, TERT, EP300 and HSP90AB1) were found to be closely associated with antitumor activity. Furthermore, 18 antitumor-related pathways were identified using the compound-target-pathway network. The molecular docking revealed that the active polyphenols had a good binding ability to the core targets, and the results were consistent with those of network pharmacology. Based on these findings, we speculate that I. hispidus can exert its antitumor activity through multicomponent, multitarget, and multichannel mechanisms of action.

Experimental and Numerical Investigations on Seismic Behavior of RC Beam to PVC-CFRP Confined Concrete Column Exterior Joint with Steel Tube Connector.

Recently, substantial investigations were developed on a polyvinyl chloride (PVC)-carbon fiber-reinforced polymer (CFRP) confined concrete (PFCC) structure owing to its superior mechanical behavior and durability. However, a convenient and effective joint configuration between the PFCC columns and reinforced concrete (RC) beams still requires in-depth study. In the present work, the seismic performance of an RC beam to PFCC column exterior joint with steel tube connector (STC) is systematically studied. Eleven joint specimens are fabricated and tested, with the steel ratio of STC, reinforcement ratio of the frame beam, axial compression ratio, stirrup ratio of the joint and CFRP strips spacing as the design parameters. The experimental results, that is, 8 the failure modes, hysteretic response, ductility, strength, stiffness and energy dissipation, are analyzed. All specimens exhibit joint shear failure, although the joints with STC exhibit significantly better performance those of ordinary joint. In addition to reducing the axial compression ratio, the reinforcement ratio of the frame beam or increasing the stirrup ratio of the joint can also produce a positive effect. Furthermore, the numerical analysis of the exterior joints is performed; the calculated skeleton curves agree well with the test results, and additional parametric studies (i.e., the diameter, height and concrete strength of the joint) are carried out based on the verified numerical model.

The regular pattern of metabolite changes in mushroom Inonotus hispidus in different growth periods and exploration of their indicator compounds.

Inonotus hispidus is a valuable and rare edible and medicinal mushroom with extremely high nutritional and medicinal value. However, there is no holistic insight to elucidate the molecular basis of the differentiated usage and accurate annotation of physiological maturity to fluctuating yields and quality. This study aimed to figure out the fruiting bodies' metabolites change regulation and potential maturating indicators to distinguish different quality I. hispidus. We applied non-targeted ultra-high performance liquid chromatography and high-resolution mass spectrometry combined and with multivariate analysis and analyzed cultivated and wild mushroom I. hispidus in different growth periods (budding, mature and aging). With the fruiting bodies maturating, 1358 metabolites were annotated, 822 and 833 metabolites abundances changed greater than or equal to 1 time from the budding period to the aging period in abundance in cultivated and wild, the total polysaccharides, crude fat, total flavonoids, and total terpenes increased at first and then decreased. Total amino acids, crude protein, and total polyphenols decreased, while the total steroids increased linearly. The change of metabolites showed certain regularity. Metabolic pathways enrichment analysis showed that these metabolites are involved in glycolysis, biosynthesis of amino acids, organic acid metabolism, glycine-serine-and-threonine metabolism, tricarboxylic acid cycle, purine metabolism, and pyrimidine metabolism. In addition, ergosterol peroxide and (22E)-ergosta-4,6,8(14),22-tetraen-3-one can be used as indicator compounds, and their contents increase linearly with the fruiting bodies of I. hispidus' physiological maturation. This comprehensive analysis will help to evaluate the edible values and facilitate exploitation in mushroom I. hispidus.

Phenolic Profile and Fingerprint Analysis of Akebia quinata Leaves Extract with Endothelial Protective Activity.

In contrast to the stem and fruit of Akebia quinata, A. quinata leaves as a source rich in phenolic compounds with potentially beneficial pharmacological activities have been largely overlooked. To develop and use A. quinata leaves as a resource, we evaluated its potential as a cardiovascular-protective agent. Herein, we investigated the effects and potential mechanisms of A. quinata leaves extract on lipopolysaccharide (LPS)-induced inflammatory responses in human umbilical vein endothelial cells. We found that A. quinata leaves extract pretreatment of 10 µg/mL significantly attenuated LPS-induced protein expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1. Furthermore, this extract also suppressed LPS-induced phosphorylation of nuclear factor-κB p65. In order to elucidate the chemical profiles of the samples, the HPLC fingerprint was established, and prominent peaks were identified via HPLC-electrospray ionization-mass spectrometry. Multivariate statistical analyses, including hierarchical cluster analysis, principal component analysis, and partial least-squares discriminant analysis, were performed to evaluate the clustering of the samples. It was found that isochlorogenic acid C was a key marker for the classification of A. quinata leaves from the Gongju and Muju city in Korea. Collectively, this study not only suggested the potential of A. quinata leaves as a novel therapeutic candidate for inflammatory cardiovascular disease but also developed a quality control method for A. quinata leaves, which could help to expand the application of A. quinata.

Deciphering key regulators of Inonotus hispidus petroleum ether extract involved in anti-tumor through whole transcriptome and proteome analysis in H22 tumor-bearing mice model.

ETHNOPHARMACOLOGICAL RELEVANCE: The mushroom Inonotus hispidus is traditional Chinese medicine, which has been used to treat tumor illness for a long history in China. Our previous research found that I. hispidus petroleum ether extract (IPE) has significant anti-tumor activity. However, the potential anti-tumor regulatory pathways and targets of I. hispidus remain unclear. OBJECTIVE: The present study was envisaged to explore the key regulators responsible for anti-tumor of IPE using whole transcriptome and proteome analysis in H22 tumor-bearing mice model. MATERIALS AND METHODS: The model of H22 tumor-bearing mice was constructed according to the histopathological data and biochemical parameters. The isolated tumor tissues of different treatment groups were subjected to transcriptomic and proteomic analysis. An integrated approach of RNA-Seq, proteomics, and system biology analysis was used to identify key regulators involved in antitumor pathways. The analyzed differential expression patterns were supported by gene and protein expression studies. RESULTS: These results indicated that 957 differentially expressed genes and 405 proteins were identified in the tumor tissue of different treatment groups through RNA-Seq and liquid chromatography with tandem mass spectrometry-based proteomic analysis, respectively. The combined omics analysis revealed five critical genes/proteins, including Lilrb4a, Nrp1, Gzma, Gstt1, and Pdk4 that could play a role in antitumor pathways. Furthermore, Lilrb4a, Nrp1, Gzma, Gstt1 and Pdk4 genes/proteins, as key regulators of the anti-tumor effect of IPE, were verified by qRT-PCR and western blotting methods, respectively. CONCLUSION: Our study provides new ideas for analyzing the antitumor mechanism of IPE from the point of view of gene and protein expression and will encourage further development of the I. hispidus pharmaceutical industry.

[Antitumor effect and mechanism of different extracts of cultivated Phellinus vaninii on H22 tumor bearing mice].

In order to explore the antitumor effect and mechanism of different extracts of cultivated Phellinus vaninii fruit body on H22 tumor bearing mice, 150 ICR mice were randomly divided into blank group, model group, CTX group, P. vaninii water extract group, ethanol extract group, petroleum ether extract group and crude polysaccharide group. H22 liver cancer cells were used to establish a solid tumor model and the mice were sacrificed on the 10th day after administration. The spleen and thymus organ index and tumor inhibition rate were calculated, the serum levels of TNF-α, INF-γ, VEGF, and hematoxylin-eosin were detected, and the immunohistochemical staining method was used to observe the pathological changes of tumor tissues, while Western blotting was used to detect the expression of tumor-related proteins. The high-dose petroleum ether extract group showed the best tumor inhibition rate (73.21%), increased serum levels of TNF-α, IFN-γ, and VEGF, as well as significantly promoted tumor necrosis and ablation. The immunohistochemistry of the water extract group showed negative regulation, indicating an insignificant tumor suppression. Western blotting showed the apoptosis genes Caspase-3, Caspase-9 and pathway genes NF-κB and JAK were all highly expressed in each administration group compared with the model group, and their expression levels gradually decreased with increasing doses. In summary, the petroleum ether extract of P. vaninii fruit body showed a significant anti-tumor effect which is presumably mediated through the mitochondrial pathway. The metabolism of drug in the body induces activation of Caspase-3 and Caspase-9 apoptotic proteins by Bax, Bcl-2, and TNF, which further caused nuclear chromatin or DNA to condense or degrade, and subsequently destroy the normal proliferation of tumor cells, thereby inducing their apoptosis and inhibiting tumor growth.

Comparative Analysis of Metabolic Compositions and Trace Elements of Inonotus hispidus Mushroom Grown on Five Different Tree Species.

Inonotus hispidus is a popular edible and medicinal mushroom widely used in China. I. hispidus mushroom mainly grows on five different tree species (Morus alba L., Ulmus macrocarpa Hance, Fraxinus mandshurica Rupr., Ziziphus jujuba Mill., and Malus pumila Mill.), and their fruiting bodies were all separately used in the market. However, there is no holistic insight to elucidate the molecular basis of the differentiated usage. This study aimed to investigate and compare the metabolite compositions and trace elements in I. hispidus grown on five different tree species. The metabolomic data, 8 kinds of principal components and 12 kinds of trace elements, were analyzed in this study. The results showed that the same 1353 metabolites were identified in I. hispidus grown on five different tree species, but the relative abundance was different. The principal components and trace elements contents are different, for example, polysaccharides, phenol metabolites, Ca, Na, Mg, Fe, and Mn were enriched in I. hispidus grown on M. alba, the flavonoids were enriched in Z. jujuba samples, and the steroids, terpenoids, and Zn were enriched in M. pumila samples. Further, the KEGG enrichment pathway and metabolic models were established. These findings provide a molecular basis for the unique use of the I. hispidus mushroom grown on different tree species.

Cation exchange as a single polishing step for conjugated peptides.

Purification of peptides typically includes expensive reverse phase (RP) processes, which utilize high pressure and large volumes of solvent. For two conjugated peptides, chromatography process development targeted a low-pressure aqueous process that could achieve target product purities of ≥95%, comparable to purities seen with traditional RP. A high throughput screening approach of different modalities was used to identify binding and elution conditions on a cation exchange resin and small-scale columns were used in order to assess impurity removal and process yield. The parameters for load and gradient elution were optimized to increase product purity and process productivity with a wide operating window identified where high purity and productivity are achieved. Computational modeling was then used to validate experimental chromatography results and to gain insight on the effect of the chemical modifications on the surface properties of the two peptides. Both modeling and experimental data showed that with optimization, cation exchange could be utilized as a single polishing step for conjugated peptides. Similar purities were achieved as those seen with RP with up to double the productivity.

Role of membrane structure on the filtrate flux during monoclonal antibody filtration through virus retentive membranes.

Virus removal filtration is a critical step in the manufacture of monoclonal antibody products, providing a robust size-based removal of both enveloped and non-enveloped viruses. Many monoclonal antibodies show very large reductions in filtrate flux during virus filtration, with the mechanisms governing this behavior and its dependence on the properties of the virus filter and antibody remaining largely unknown. Experiments were performed using the highly asymmetric Viresolve® Pro and the relatively homogeneous Pegasus™ SV4 virus filters using a highly purified monoclonal antibody. The filtrate flux for a 4 g/L antibody solution through the Viresolve® Pro decreased by about 10-fold when the filter was oriented with the skin side down but by more than 1000-fold when the asymmetric filter orientation was reversed and used with the skin side up. The very large flux decline observed with the skin side up could be eliminated by placing a large pore size prefilter directly on top of the virus filter; this improvement in filtrate flux was not seen when the prefilter was used inline or as a batch prefiltration step. The increase in flux due to the prefilter was not related to the removal of large protein aggregates or to an alteration in the extent of concentration polarization. Instead, the prefilter appears to transiently disrupt reversible associations of the antibodies caused by strong intermolecular attractions. These results provide important insights into the role of membrane morphology and antibody properties on the filtrate flux during virus filtration.

Anti-tumor effect of Inonotus hispidus petroleum ether extract in H22 tumor-bearing mice and analysis its mechanism by untargeted metabonomic.

ETHNOPHARMACOLOGICAL RELEVANCE: The mushroom Inonotus hispidus is traditional Chinese medicine, which has been used to treat tumor illness for many years in China. However, the potential anti-tumor mechanisms of I. hispidus remain unclear. OBJECTIVE: This study aimed to reveal the anti-tumor mechanism of I. hispidus petroleum ether extract (IPE) on H22 tumor-bearing mice from the point of view of metabonomics. MATERIALS AND METHODS: The model of H22 tumor-bearing mice was constructed according to the histopathological data and biochemical parameters, while the serum metabolomics was analyzed by non-targeted ultra-high performance liquid chromatography and high-resolution mass spectrometry (UPLC-MS/MS) to study the potential anti-tumor mechanisms of IPE. RESULTS: These results indicated that IPE has significant anti-tumor effect on H22 tumor-bearing mice and no obvious adverse reactions were observed. After the intervention of IPE, the biosynthesis of cortisol and corticosterone as the metabolics in the downstream of steroid biosynthesis pathway and the biosynthesis of succinate, fumarate and malate as the metabolics in the downstream of tricarboxylic acid cycle pathway were inhibited; but the metabolic pathways of the amino acids as tryptophan, lysine degradation, alanine, aspartate and glutamate and other amino acid were activated. CONCLUSION: IPE has significant anti-tumor effect in H22 tumor-bearing mice, and the anti-tumor activity of IPE is main through the regulation of energy, amino acids, and steroid hormone biosynthesis pathways expression.

Analyses of transcriptomes and the first complete genome of Leucocalocybe mongolica provide new insights into phylogenetic relationships and conservation.

In this study, we report a de novo assembly of the first high-quality genome for a wild mushroom species Leucocalocybe mongolica (LM). We performed high-throughput transcriptome sequencing to analyze the genetic basis for the life history of LM. Our results show that the genome size of LM is 46.0 Mb, including 26 contigs with a contig N50 size of 3.6 Mb. In total, we predicted 11,599 protein-coding genes, of which 65.7% (7630) could be aligned with high confidence to annotated homologous genes in other species. We performed phylogenetic analyses using genes form 3269 single-copy gene families and showed support for distinguishing LM from the genus Tricholoma (L.) P.Kumm., in which it is sometimes circumscribed. We believe that one reason for limited wild occurrences of LM may be the loss of key metabolic genes, especially carbohydrate-active enzymes (CAZymes), based on comparisons with other closely related species. The results of our transcriptome analyses between vegetative (mycelia) and reproductive (fruiting bodies) organs indicated that changes in gene expression among some key CAZyme genes may help to determine the switch from asexual to sexual reproduction. Taken together, our genomic and transcriptome data for LM comprise a valuable resource for both understanding the evolutionary and life history of this species.

Tumor suppressive microRNA-485-5p targets PRRX1 in human skin melanoma cells, regulating epithelial-mesenchymal transition and apoptosis.

Melanoma is one of the most aggressive skin cancers. Existing evidence has reported the aberrant expression of microRNAs (miRNAs) in melanoma, but their putative targets and underlying downstream effects remain to be further understood. Herein, we explored the suppressive role of miR-485-5p in melanoma progression. Initial bioinformatics analyses showed that the PRRX1 gene was differentially expressed in melanoma, while miR-485-5p was predicted to be a potential regulatory miRNA binding to PRRX1 mRNA. We confirmed that PRRX1 was upregulated, while miR-485-5p was downregulated in human melanoma samples compared with adjacent normal skin tissues. We then showed that PRRX1 was a target gene of miR-485-5p by dual-luciferase reporter gene assay. Moreover, a reduction in the expression of PRRX1 and downregulation of important proteins of the transforming growth factor-beta (TGFß) signaling pathway was observed after miR-485-5p overexpression. Furthermore, miR-485-5p overexpression or PRRX1 knockdown suppressed epithelial-mesenchymal transition, cell viability, migration, and invasion, and promoted cell apoptosis in melanoma cells. Our study demonstrates the tumor-suppressive functions of miR-485-5p in the development of human melanoma, providing a potential target for therapy.

Investigation of the possible mechanism of polysaccharides extracted from Leucocalocybe mongolica in exerting antitumor effects in H22 tumor-bearing mice.

A response surface method was used to optimize the extraction of polysaccharides from Leucocalocybe mongolica. Moreover, the preliminary structural characteristics and antitumor activity of L. mongolica polysaccharide (LMP) were investigated. The results showed that the optimized extraction technological parameters for LMP were 93°C extraction temperature, 5 hr extraction time, and 30 ml/g liquid-to-solid ratio. The LMP content extracted under the optimal conditions was 6.64%. LC-MS/MS results indicated that LMP is a neutral polysaccharide composed of d-fructose, d-mannose, dextrose anhydrate, d-xylose, trehalose, and galactose. The tumor inhibition rate was significantly improved by LMP treatment. LMP had minimal toxicity based on the significant decrease in AST and BUN levels; VEGF protein levels were also significantly decreased. In contrast, the levels of IFN-γ, IL-2, IL-6, and TNF-α were improved. The results of ELISA, H&E staining, TUNEL assay, immunohistochemistry, and western blotting indicated that the LMP exhibited antitumor activity in vivo by promoting apoptosis, mediating inflammatory responses, and inhibiting angiogenesis. PRACTICAL APPLICATIONS: As one of the main bioactive components, fungal polysaccharide has always been a hot research topic. Fungal polysaccharides are carbohydrate polymers composed of monosaccharide units bound together by glycosidic linkages, which have been found to be involved in many biological processes. In this research, the LMP structure was analyzed, and the immunohistochemical and western blot analysis confirmed that, LMP could effectively reduce the generation of tumor angiogenesis, promote apoptosis of tumor cell sand inhibit tumor growth. The results of this study can effectively provide a basis for clinical research and development of antitumor drugs, and lay a foundation for the study of the antitumor effects of wild edible and medicinal fungi.

Quality Control of Radix Astragali (The Root of Astragalus membranaceus var. mongholicus) Along Its Value Chains.

Radix Astragali (RA), the root of Astragalus membranaceus var. mongholicus (Bunge) P.K. Hsiao, known as "Huangqi" in Chinese, has been used as a traditional herbal medicine or food in China for more than 2,000 years and is now consumed globally. Unfortunately, the increasing demand for RA has led to the overexploitation of its wild stock, as well as quality problems, including adulteration and contamination. Therefore, the sustainable cultivation of RA is urgently needed. In the present research, semi-structured interviews and key informant interviews were conducted, over a 2-year period, to collect data from stakeholders in the main production areas; moreover, a targeted chemical analysis-based quality assessment strategy was applied to understand the quality of RA. Accordingly, 10 different types of value chains (VCs) were identified in RA production; meanwhile, the contents of the main active ingredients (astragaloside and calycosin-7-O-ß-D-glucoside) were analyzed by HPLC-ELSD-UV and the yield of medicinal material was demined and further analyzed using k-means clustering analysis. The results show that the tight relationship between quality of the RA and stakeholders' revenues among the VCs, which reflects a more general trend in the production system. Over the past few decades, vertical coordination has emerged increasingly in VCs of RA, which leads to a more coherent traceability system and rigorous regulations in the supply chains. Daodi herbs can be considered to be a standard that is distinctive with good quality characteristics that emphasize the origins of the medicinal plants. We find that the suitability of geographical areas and vertical integration can improve the VCs of RA, which further contributes to its quality control, as well as its sustainable production.

Investigation of the possible mechanism of two kinds of sterols extracted from Leucocalocybe mongolica in inducing HepG2 cell apoptosis and exerting anti-tumor effects in H22 tumor-bearing mice.

Sterols are one of the main components of medicinal fungi with an anti-tumor effect. In this study, ergosta-4, 6, 8(14), 22-tetraen-3-one (ET) and (22E, 24R)-ergosta-7, 22-dien-3ß, 5α, 6ß-triol (ED) were obtained from Leucocalocybe mongolica and were used for the first time to study their ability to induce apoptosis in HepG2 cells and their anti-tumor effects and related mechanism in H22 tumor-bearing mice. METHOD: The chemical structures were defined by IR and NMR. In vitro, the CCK8 assay was used as a cytotoxicity assay. Flow cytometry was used for the HepG-2 cell apoptosis analysis, which was examined via annexin V-FITC/PI double staining, and the related expression levels of the apoptosis-associated proteins were determined by western blot analysis. In vivo, ICR male mice were randomly assigned to eight groups: the model group, CTX (25 mg/kg/d) group, and ET and ED groups, which were treated with three different concentrations of each compound (0.025, 0.05, and 0.1 mmol/kg/d). Relevant biochemical indicators were detected by ELISA assay, H & E staining, TUNEL assay, immunohistochemical staining and western blot. RESULTS: In vitro, ET and ED showed significant cytotoxic effects against HepG2, MCF-7, and HeLa cells, especially HepG-2 cells, and both ED and ET demonstrated a good effect in inhibiting the proliferation of HepG-2 cells. In vivo, ET and ED significantly decreased the tumor volume and VEGF levels but increased the serum cytokine levels of IFN-γ, IL-2, IL-6 and TNF-α. H & E staining, TUNEL assay, immunohistochemical analysis, and western blotting indicated that the both ET and ED exhibited anti-tumor activity in vivo by promoting apoptosis and inhibiting angiogenesis. CONCLUSION: These results indicated that both ET and ED have a strong inhibitory effect on the proliferation of HepG-2 cells in vitro and an anti-H22 tumor effect in vivo.

Preventive Effect of Small Molecular Fraction of Irpex lacteus (Agaricomycetes) Fruiting Body against Chronic Nephritis in Mice and Identification of Active Compounds.

Previous study found that the fruit body of Irpex lacteus has an effect on the prevention and treatment of chronic nephritis. In this study, we systematically investigated the preventive effect of small molecular fraction (SMF) of the fungal fruit body against chronic nephritis. In addition, we analyzed, isolated, and identified the chemical constituents of SMF, and screened the activity of three small peptides isolated in vitro. The results showed SMF significantly reduced amounts of urine protein (UP), the content of urea (BUN), creatinine (Cr), tumor necrosis factor-α (TNF-α), and maleic dialdehyde (MDA) in serum, and significantly increased superoxide dismutase (SOD) level in renal tissue homogenate (P < 0.05). Moreover, the results of hematoxylin and eosin (H&E) and Masson staining of renal tissues indicated that SMF has protective effects on renal tissues and prevents renal interstitial from fibrosis. The peptide sequences isolated from SMF were identified as WSMGPAPDSVH (SP1), QCTGNASCSPPC (SP2), and HYCCTAKYA (SP3), which were active compounds for the prevention of nephritis, and these new peptides were isolated for the first time. The cell proliferation assay showed that 10 µg/L transforming growth factor-ß1 (TGF-ß1) significantly induced the proliferation of human renal tubular epithelial cells (HK-2), compared with the control group, and the difference was statistically significant (P < 0.01). However, when combined with three small peptides, respectively, the cell proliferation was significantly inhibited (P < 0.05). These results suggest that isolated peptides can maintain the morphological stability of HK-2 cells, inhibit cell proliferation induced by TGF-ß1 to some extent, and prevent cell fibrosis.

GC-MS Analysis and Cytotoxicity Detection of Volatile Oil from the Fragrant Bracket Mushroom, Trametes suaveolens (Agaricomycetes).

Trametes suaveolens is a medicinal mushroom known as Baizhi in traditional Chinese medicine. Our previous research has found that it has some pharmacological activity in vivo. The aim of the study was to investigate the chemical compounds and cytotoxic effects of volatile oil from T. suaveolens. In this study, internal transcribed spacer (ITS) sequence analysis was used to determine wild T. suaveolens collected. To fully analyze the composition of volatile oil extracted from T. suaveolens, hydrodistillation (HD) and solid phase microextraction (SPME) were adopted simultaneously. In both cases, the analysis was carried out using gas chromatography-mass spectrometry (GC-MS) and the cytotoxic effects of T. suaveolens volatile oil on human NCI-H460 lung non-small cell carcinoma cells and MCF-7 breast adenocarcinoma cells were investigated. The results indicated that all these wild samples were identified as T. suaveolens. Thirty-one components in HD and 62 components in SPME were identified, respectively. Furthermore, the volatile compounds obtained from T. suaveolens by HD indicated that esters compounds were a major class (68.47%), followed by acids (25.06%), aldehydes (4.20%), and alcohols (1.48%). SPME found that the largest content were aldehydes (45.47%), followed by alcohols (31.42%), ketones (6.89%), and esters (6.72%). In the cytotoxic assays, the volatile oil was found to have toxic effect on NCI-H460 and MCF-7 tumor cells but not BEAS-2B and MCF-10A normal cells, and the IC50 values of NCI-H460 and MCF-7 tumor cells were 24.1 and 19.2 µg/ml, respectively. The present study shows that the composition of essential oil from T. suaveolens has potential value for the prevention and treatment of lung cancer.