Novel anti-inflammatory diketopiperazine alkaloids from the marine-derived fungus Penicillium brasilianum.

Diketopiperazine alkaloids have proven the most abundant heterocyclic alkaloids up to now, which usually process diverse scaffolds and rich biological activities. In our search for bioactive diketopiperazine alkaloids from marine-derived fungi, two novel diketopiperazine alkaloids, penipiperazine A (1) and its biogenetically related new metabolite (2), together with a known analogue neofipiperzine C (3), were obtained from the strain Penicillium brasilianum. Their planar structures and absolute configurations were elucidated by extensive spectroscopic analyses, 13C NMR calculation, Marfey's, ECD, and ORD methods. Compound 1 featured a unique 6/5/6/6/5 indole-pyrazino-pyrazino-pyrrolo system, and its plausible biogenetic pathway was also proposed. Additionally, compounds 1-3 have been tested for their inflammatory activities. 1 and 2 significantly inhibited the release of NO and the expression of related pro-inflammatory cytokines on LPS-stimulated RAW264.7 cells, suggesting they could be attracting candidate for further development as anti-inflammatory agent. KEY POINTS: • A novel diketopiperazine alkaloid featuring a unique 6/5/6/6/5 indole-pyrazino-pyrazino-pyrrolo system was isolated from the marine fungus Penicillium brasilianum. • The structure of 1 was elucidated by detailed analysis of 2D NMR data, 13C NMR calculation, Marfey's, ECD, and ORD methods. • Compounds 1 and 2 significantly inhibited the release of NO and the expression of related pro-inflammatory cytokines on LPS-stimulated RAW264.7 cells.

Diverse indole-diterpenoids with protein tyrosine phosphatase 1B inhibitory activities from the marine coral-derived fungus Aspergillus sp. ZF-104.

Eight previously undescribed indole-diterpenoids named penerpenes O-V (1-8), together with seven known analogues (9-14), were isolated from the marine soft coral-derived fungus Aspergillus sp. ZF-104. Their structures including the absolute configurations of these compounds were assigned on the basis of spectroscopic data and ECD analysis along with quantum ECD and NMR calculations. Compounds 4 and 5 bear rare indolin-2-one units in their structures and 6 bears a reconstructed novel skeleton in which the indole ring and the terpenoid substructure are cleaved before they are reconnected through the nitrogen atom. Compounds 1, 2, 7, and 10 showed protein tyrosine phosphatase 1B (PTP1B) inhibitory activities comparable to that of the positive control NaVO3.

Structural diversity and biological activities of indole-diterpenoids from Penicillium janthinellum by co-culture with Paecilomyces formosus.

Co-culturing the marine-derived fungi Penicillium janthinellium with Paecilomyces formosus led to the isolation of nine new indole-diterpenes, janthinellumines A-I (1-9), along with twelve known analogues (10-21). The chemical structures including their absolute configurations of them were assigned by the analysis of extensive spectroscopic data and calculated ECD and VCD methods. These indole-diterpenoids displayed extensive biological activities, including anti-influenza A virus, protein tyrosine phosphatase (PTP) inhibitory, and anti-Vibrio activities. Among them, the anti-influenza mechanism of compounds 1, 2, and 7 was further investigated using neuraminidase inhibitory assay, molecular docking, and reverse genetics methods, suggesting that 1, 2, and 7 could interact with Arg371 of the viral neuraminidase. The structure-activity relationship (SAR) of PTPs inhibitory activity for indole-diterpene derivatives (1, 2, 4, 5, 9-16, and 19-21) was also summarized.

Two new alkaloids from the endophytic fungus Schizophyllum sp. HM230 isolated from Vincetoxicum mongolicum Maxim.

Endophytic fungi is an important source for the discovery of bioactive natural compounds. A chemical investigation of the ethyl acetate extract of the endophytic fungus Schizophyllum sp. HM230 derived from stems of the herb Vincetoxicum mongolicum Maxim led to isolation of five alkaloids, including two new compounds, schizophyllins M (1) and N (2), along with three known ones (3-5). The planar structures of two new compounds were elucidated by extensive spectroscopic methods including MS, 1D and 2D NMR. Their absolute configurations were determined by Mosher's method and comparison of the ECD data. All the isolates were evaluated for their cytotoxicity and antioxidant activities. Compounds 1-4 showed middle cytotoxicity against MCF-7 cells with IC50 values range of 68.1 ∼ 87.32 µM. Compounds 1-5 displayed obvious antioxidant activity with the IC50 values range of 0.86 ∼ 5.78 mg/mL.

Application of Cellulase for Contributing Phenolic Release and Conversion in Oats (Avena sativa L.) During Microbial Fermentation.

In this work, Monascus fermentation and cellulase hydrolysis (MCF) of oats (Avena sativa L.) to release and convert phenolic fraction was investigated. Results showed the fungus Monascus grew well with a biomass of 27.03 mg/g glucosamine equivalent in MCF, following the destruction of oat cellular structures. SDS-PAGE revealed lots of enzymes were regulated with the α-amylase and FPase activity achieved 139.25 U/g and 1.84 U/g in MCF, respectively. Compared with unfermented oats, content of the total phenolic fractions was increased by 19.2 times in MCF, suggesting a phenolic release process occurred during fermentation. Moreover, the soluble-free chlorogenic acid upregulated to 510.00 mg/kg whereas the insoluble-bound ferulic acid downregulated to 193.36 mg/kg in MCF, indicating a transformation process of chlorogenic acid from ferulic acid in oats was enhanced. Based on this, a possible pathway of phenolic release and conversion in oats during fermentation with Monascus spp. was revealed. This study was helpful to enrich the theory of microbial metabolism and transformation in grain materials.

Carotane sesquiterpenoids A-G from the desert endophytic fungus Fusarium sp. HM 166.

Seven undescribed carotane sesquiterpenoids named fusanoids A-G (1-7), along with one known analog (8) and two known sesterterpenes (9 and 10), were isolated from the fermentation broth of the desert endophytic fungi Fusarium sp. HM166. The structures of the compounds, including their absolute configurations, were determined by spectroscopic data, single-crystal X-ray diffraction analysis, and ECD calculations. Compound 10 showed cytotoxic activities against human hepatoma carcinoma cell line (Huh-7) and human breast cell lines (MCF-7 and MDA-MB-231), and compound 2 showed cytotoxic activity against MCF-7, while compounds 4-9 were inactive against all the tested cell lines. Compounds 4 and 10 showed potent inhibitory activities against the IDH1R132h mutant.

Transcriptome Analysis of Liver Cancer Cell Huh-7 Treated With Metformin.

Metformin is a kind of widely used antidiabetic drug that regulates glucose homeostasis by inhibiting liver glucose production and increasing muscle glucose uptake. Recently, some studies showed that metformin exhibits anticancer properties in a variety of cancers. Although several antitumor mechanisms have been proposed for metformin action, its mode of action in human liver cancer remains not elucidated. In our study, we investigated the underlying molecular mechanisms of metformin's antitumor effect on Huh-7 cells of hepatocellular carcinoma (HCC) in vitro. RNA sequencing was performed to explore the effect of metformin on the transcriptome of Huh-7 cells. The results revealed that 4,518 genes (with log2 fold change > 1 or < -1, adjusted p-value < 0.05) were differentially expressed in Huh-7 cells with treatment of 25-mM metformin compared with 0-mM metformin, including 1,812 upregulated and 2,706 downregulated genes. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses identified 54 classical pathways that were significantly enriched, and 16 pathways are closely associated with cancer, such as cell cycle, DNA replication, extracellular matrix-receptor interaction, and so on. We selected 11 differentially expressed genes, which are closely associated with HCC, to validate their differential expressions through a quantitative real-time reverse transcription-polymerase chain reaction. The result exhibited that the genes of fatty acid synthase, mini-chromosome maintenance complex components 6 and 5, myristoylated alanine-rich C-kinase substrate, fatty acid desaturase 2, C-X-C motif chemokine ligand 1, bone morphogenetic protein 4, S-phase kinase-associated protein 2, kininogen 1, and proliferating cell nuclear antigen were downregulated, and Dual-specificity phosphatase-1 is significantly upregulated in Huh-7 cells with treatment of 25-mM metformin. These differentially expressed genes and pathways might play a crucial part in the antitumor effect of metformin and might be potential targets of metformin treating HCC. Further investigations are required to evaluate the metformin mechanisms of anticancer action in vivo.

Predator-Prey Interactions between Nonnative Juvenile Largemouth Bass (Micropterus salmoides) and Local Candidate Prey Species in the Pearl River Delta: Predation Capacity, Preference and Growth Performance.

An ontogenetic dietary shift is crucial for the survival and growth of piscivorous largemouth bass (LB). However, there is much to learn about the predator-prey interaction during the switching process. We carried out a series of indoor experiments to examine the predation capacity, predation preference, and growth performance of exotic juvenile LB feeding on candidate prey species in the Pearl River Delta. The widely distributed oriental river prawn (Macrobranchium nipponense), barcheek goby (Ctenogobius giurinus), western mosquitofish (Gambusia affinis), silver carp (Hypophthalmichthys molitrix), and mud carp (Cirrhinus molitorella), with relatively similar total lengths, were selected as potential prey based on their availability and habitat use. Our results show that predation capacity and preference varied quantitatively and qualitatively among prey species. The number of oriental river prawns killed was significantly less than that of fish species, comparing the 1st hour with the 24th hour (p < 0.01). The feeding rhythm of LB varied significantly from crayfish to fish. Numerically, Jacobs' selection index reinforced LB's special preference for predating G. affinis. Although there were obvious variations in predation capacity and feed selection, no statistically significant growth differences were detected among LB groups feeding on live M. nipponense, G. affinis, H. molitrix, and C. molitorella (p < 0.05). These findings suggest that the successful ontogenetic dietary shift of juvenile LB may depend on the availability and vulnerability of local fish species. Further study on the reproductive phenology of potential fish prey may help to predict LB's establishment.

Three New Quinazoline-Containing Indole Alkaloids From the Marine-Derived Fungus Aspergillus sp. HNMF114.

By feeding tryptophan to the marine-derived fungus Aspergillus sp. HNMF114 from the bivalve mollusk Sanguinolaria chinensis, 3 new quinazoline-containing indole alkaloids, named aspertoryadins H-J (1-3), along with 16 known ones (4-19), were obtained. The structures of the new compounds were elucidated by the analysis of spectroscopic data combined with quantum chemical calculations of nuclear magnetic resonance (NMR) chemical shifts and electron capture detector (ECD) spectra. Structurally, compound 3 represents the first example of this type of compound, bearing an amide group at C-3. Compounds 10 and 16 showed potent α-glucosidase inhibitory activity with IC50 values of 7.18 and 5.29 µM, and compounds 13 and 14 showed a clear activation effect on the ryanodine receptor from Spodoptera frugiperda (sfRyR), which reduced the [Ca2+] ER by 37.1 and 36.2%, respectively.

Construction and Comparison of ceRNA Regulatory Network for Different Age Female Breast Cancer.

Studies have shown the difference appearing among the prognosis of patients in different age groups. However, the molecular mechanism implicated in this disparity have not been elaborated. In this study, expression profiles of female breast cancer (BRCA) associated mRNAs, lncRNAs and miRNAs were downloaded from the TCGA database. The sample were manually classified into three groups according to their age at initial pathological diagnosis: young (age ≤ 39 years), elderly (age ≥ 65 years), and intermediate (age 40-64 years). lncRNA-miRNA-mRNA competitive endogenous RNA (ceRNA) network was respectively constructed for different age BRCA. Then, the biological functions of differentially expressed mRNAs (DEmRNAs) in ceRNA network were further investigated by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Finally, survival analysis was used to identify prognostic biomarkers for different age BRCA patients. We identified 13 RNAs, 38 RNAs and 40 RNAs specific to patients aged ≤ 39 years, aged 40-64 years, and aged ≥ 65 years, respectively. Furthermore, the unique pathways were mainly enriched in cytokine-cytokine receptor interaction in patients aged 40-64 years, and were mainly enriched in TGF-beta signaling pathway in patients aged ≥ 65 years. According to the survival analysis, AGAP11, has-mir-301b, and OSR1 were respectively functioned as prognostic biomarkers in young, intermediate, and elderly group. In summary, our study identified the differences in the ceRNA regulatory networks and provides an effective bioinformatics basis for further understanding of the pathogenesis and predicting outcomes for different age BRCA.

Regulation of phenolic release in corn seeds (Zea mays L.) for improving their antioxidant activity by mix-culture fermentation with Monascus anka, Saccharomyces cerevisiae and Bacillus subtilis.

In this study, mix-culture fermentation (MF) with M. anka, S. cerevisiae and B. subtilis to regulate phenolic release in corn seeds was investigated. Results showed the adding strategy of S. cerevisiae and B. subtilis significantly (p < 0.05) influenced the total phenolic content (TPC) of corn seeds, and the highest TPC in MF was 22.56-fold rise than that of unfermented control. Moreover, the phenolic compounds in corn seeds were shifted after MF. SDS-PAGE revealed some enzymes produced by microorganisms were regulated, and the α-amylase and FPase activities were changed to improvement in MF. SEM micrographs of corn seeds in MF varied markedly with the cell walls structure destroyed, so as to release of the phenolic fractions in corn seeds. Based on the phenolic mobilization, DPPH and ABTS+ radical scavenging abilities of phenolic fractions from corn seeds were improved in MF. This paper provides a prospect fermentation process for improving the phenolic release in corn cereals.

Characterization and phytotoxicity of ophiobolins produced by Bipolaris setariae.

The Bipolaris setariae NY1 strain, isolated from a diseased green foxtail plant in Henan Province, China, showed strong pathogenicity towards green foxtail. In order to clarify the role of phytotoxic substances in the fungal pathogenicity, bioassay-directed isolation and bioactivity assays of secondary metabolites produced by the fungal strain were carried out. Five ophiobolins were obtained: 3-anhydro-ophiobolin A, 6-epi-ophiobolin A, 6-epi-ophiobolin B, 3-anhydro-6-epi-ophiobolin B and ophiobolin I. Bioassays on punctured and intact detached leaves of green foxtail indicated that 3-anhydro-ophiobolin A was the most phytotoxic, followed by 6-epi-ophiobolin A. The other three ophiobolins appeared to be inactive against green foxtail. The effects of 3-anhydro-ophiobolin A and 6-epi-ophiobolin A were synergistic. The symptoms on green foxtail caused by 3-anhydro-ophiobolin A or its mixture with 6-epi-ophiobolin A resembled those caused by the fungus. 3-Anhydro-ophiobolin A and 6-epi-ophiobolin A are likely the main pathogenic determinants of B. setariae. 6-epi-Ophiobolin A caused cytotoxicity against five kinds of human cancer cells: human colon adenocarcinoma cells (HCT-8), human liver cancer cells (Bel-7402), human gastric cancer cells (BGC-823), human lung adenocarcinoma cells (A549), and human ovarian adenocarcinoma cells (A2780). The results provide information for the development of herbicides and antitumor potential of the ophiobolin sesterterpenes.

Dipleosporalones A and B, Dimeric Azaphilones from a Marine-Derived Pleosporales sp. Fungus.

Dipleosporalones A and B (1 and 2), two new [2 + 2] azaphilone dimers, were obtained from a marine-derived Pleosporales sp. fungus. The absolute configurations of 1 and 2 were elucidated by calculations of their ECD spectra. Dipleosporalone A (1) possessed an unprecedented skeleton with an uncommon 6/4/6 ring system. Compounds 1 and 2 showed cytotoxicity about 30-90-fold more potent than that of their monomer pinophilin B.

Chemical Deprenylation of N6 -Isopentenyladenosine (i6 A) RNA.

N6 -isopentenyladenosine (i6 A) is an RNA modification found in cytokinins, which regulate plant growth/differentiation, and a subset of tRNAs, where it improves the efficiency and accuracy of translation. The installation and removal of this modification is mediated by prenyltransferases and cytokinin oxidases, and a chemical approach to selective deprenylation of i6 A has not been developed. We show that a selected group of oxoammonium cations function as artificial deprenylases to promote highly selective deprenylation of i6 A in nucleosides, oligonucleotides, and live cells. Importantly, other epigenetic modifications, amino acid residues, and natural products were not affected. Moreover, a significant phenotype difference in the Arabidopsis thaliana shoot and root development was observed with incubation of the cation. These results establish these small organic molecules as direct chemical regulators/artificial deprenylases of i6 A.

Transcriptome Analysis of MDA-MB-231 Cells Treated with Fumosorinone Isolated from Insect Pathogenic Fungi.

BACKGROUND: In our previous study, we have isolated a new compound, named Fumosorinone (FU) from insect pathogenic fungi, and was found to inhibit proliferation, migration, and invasion of breast cancer MDA-MB-231 cells. OBJECTIVE: The aim of this study was to identify the underlying molecular mechanisms for FU effects on MDAMB- 231 cells. METHODS: After MDA-MB-231 cells were treated with FU for 48h, RNA sequencing was used to identify the effect of FU on the transcriptome of MDA-MB-231 cells. The validation of the relative expression of the selective genes was done using quantitative real-time PCR (qRT-PCR). RESULTS: The transcriptome results showed that 2733 genes were differentially expressed between the untreated and the FU-treated cells, including 1614 up-regulated and 1119 down-regulated genes. The multiple genes are associated with cancer cell growth, migration, and invasion. Functional analysis identified multitude of pathways related to cancer, such as cell cycle, ECM-receptor interaction, p53 signaling pathway. We selected 4 upregulated and 9 downregulated genes, which are associated with breast cancer to verify their expression using qRT-PCR. The validation showed that HSD3B1, ALOX5, AQP5, COL1A2, CCNB1, CCND1, VCAM-1, PTPN1 and PTPN11 were significantly downregulated while DUSP1, DUSP5, GADD45A, EGR1 were upregulated in FU-treated MDA-MB-231cells. CONCLUSION: These aberrantly expressed genes and pathways may play pivotal roles in the anti-cancer activity of FU, and maybe potential targets of FU treatments for TNBC. Further investigations are required to evaluate the FU mechanisms of anti-cancer action in vivo.

Indole-Diterpenoids with Protein Tyrosine Phosphatase Inhibitory Activities from the Marine-Derived Fungus Penicillium sp. KFD28.

Five new indole-terpenoids named penerpenes E-I (1-5), along with seven known ones (6-12), were isolated from the marine-derived fungus Penicillium sp. KFD28 from a bivalve mollusk, Meretrix lusoria. The structures of the new compounds were elucidated from spectroscopic data and ECD spectroscopic analyses. Compound 1 was assigned as an indole-diterpenoid with a unique 6/5/5/6/6/5/5 heptacyclic ring system. Compound 2 represents an indole-diterpenoid with a new carbon skeleton derived from paxilline by the loss of three carbons (C-23/24/25). Compound 3 contains an additional oxygen atom between C-21 and C-22 compared to paxilline to form an unusual 6/5/5/6/6/7 hexacyclic ring system bearing a 1,3-dioxepane ring, which is rarely encountered in natural products. Compounds 1, 2, 4, and 6 showed inhibitory activities against protein tyrosine phosphatase 1B (PTP1B) with IC50 values of 14, 27, 23, and 13 µM, respectively.

Penerpenes A-D, Four Indole Terpenoids with Potent Protein Tyrosine Phosphatase Inhibitory Activity from the Marine-Derived Fungus Penicillium sp. KFD28.

Four unusual indole-terpenoids, penerpenes A-D (1-4), along with two known ones paxilline (5) and emindole SB (6), were isolated from the marine-derived fungus Penicillium sp. KFD28. The absolute structures of 1-4 were elucidated on the basis of spectroscopic data and ECD spectra analysis along with quantum ECD calculations. Compounds 1 and 2 showed potent inhibitory activity toward protein tyrosine phosphatases (PTP1B and TCPTP). Plausible biosynthetic pathways of compounds 1-4 are proposed.

Divergolides T⁻W with Apoptosis-Inducing Activity from the Mangrove-Derived Actinomycete Streptomyces sp. KFD18.

Four new ansamycins, named divergolides T-W (1-4), along with two known analogs were isolated from the fermentation broth of the mangrove-derived actinomycete Streptomyces sp. KFD18. The structures of the compounds, including the absolute configurations of their stereogenic carbons, were determined by spectroscopic data and single-crystal X-ray diffraction analysis. Compounds 1-4 showed cytotoxic activity against the human gastric cancer cell line SGC-7901, the human leukemic cell line K562, the HeLa cell line, and the human lung carcinoma cell line A549, with 1 being the most active while compounds 5 and 6 were inactive against all the tested cell lines. Compounds 1 and 3 showed very potent and specific cytotoxic activities (IC50 2.8 and 4.7 µM, respectively) against the SGC-7901 cells. Further, the apoptosis-inducing effect of 1 and 3 against SGC-7901 cells was demonstrated by two kinds of staining methods for the first time.

The influence of warming on the biogeographic and phylogenetic dependence of herbivore-plant interactions.

Evolutionary experience and the phylogenetic relationships of plants have both been proposed to influence herbivore-plant interactions and plant invasion success. However, the direction and magnitude of these effects, and how such patterns are altered with increasing temperature, are rarely studied. Through laboratory functional response experiments, we tested whether the per capita feeding efficiency of an invasive generalist herbivore, the golden apple snail, Pomacea canaliculata, is dependent on the biogeographic origin and phylogenetic relatedness of host plants, and how increasing temperature alters these dependencies. The feeding efficiency of the herbivore was highest on plant species with which it had no shared evolutionary history, that is, novel plants. Further, among evolutionarily familiar plants, snail feeding efficiency was higher on those species more closely related to the novel plants. However, these biogeographic dependencies became less pronounced with increasing temperature, whereas the phylogenetic dependence was unaffected. Collectively, our findings indicate that the susceptibility of plants to this invasive herbivore is mediated by both biogeographic origin and phylogenetic relatedness. We hypothesize that warming erodes the influence of evolutionary exposure, thereby altering herbivore-plant interactions and perhaps the invasion success of plants.

Carotane-type sesquiterpenes from cultures of the insect pathogenic fungus Isaria fumosorosea.

Two new carotane-type sesquiterpenes named trichocaranes E (1) and F (2), along with two known ones CAF-603 (3) and trichocarane C (4), were isolated from cultures of the insect pathogenic fungus Isaria fumosorosea. Their structures and relative configurations were elucidated by extensive spectroscopic analysis and X-ray crystallography. Compounds 1-3 showed potent cytotoxic activities against six tumor cell lines MDA, MCF-7, SKOV-3, Hela, A549, HepG2 with IC50 values in a concentration range of 0.1-6.0 µg/ml.