The unique Akt inhibitor SC66 suppressed AMPK activity and abolished autophagy through the EGFR-p62 pathway.

Akt is usually considered to be a negative regulator of both autophagy and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) signaling. In the present study, we found that SC66, a pyridine-based allosteric Akt inhibitor, suppressed basal and H2 O2 -induced autophagy concurrent with decreased phosphorylation and activity of AMPK. SC66 treatment led to the formation of a high molecular weight (HMW) form of SQSTM1/p62 (p62), which is an autophagic substrate and is essential for selective autophagy. Moreover, we observed that SC66 inhibited the binding of p62 and microtubule-associated protein light chain 3 (LC3). The immunoprecipitation results revealed the interaction between p62 and epidermal growth factor receptor (EGFR), and knockdown of EGFR reversed SC66-mediated autophagy inhibition without affecting the phosphorylation of acetyl-CoA carboxylase (ACC), a well-known substrate of AMPK. SC66 increased the interaction between EGFR and Beclin 1 and markedly decreased the association of EGFR with VPS34, a critical protein for autophagy induction. Collectively, the data presented here indicate that EGFR-p62 pathway plays a critical role in Akt-mediated positive regulation of autophagy.

Different Role of Raptor and Rictor in Regulating Rasfonin-Induced Autophagy and Apoptosis in Renal Carcinoma Cells.

Both Raptor and Rictor are the key components in the complexes of mammalian target of rapamycin (mTOR), which play a vital role in mediating autophagy. Unlike mTOR, the regulatory role of either Raptor or Rictor in the regulation of autophagic process is relatively less explored. In present study, we found that rasfonin, which isolated from Talaromyces sp. 3656-A1 and was a fungal natural product, activated both caspase-dependent apoptosis and autophagy in ACHN, a renal carcinoma cell line. Knockdown of Raptor decreased both rasfonin-induced autophagic flux and PARP-1 cleavage, and in contrast, Rictor silencing increased apoptosis concomitantly enhancing rasfonin-induced autophagy. Unexpectedly, API-2, which was widely used as an inhibitor of Akt, promoted rasfonin-dependent autophagy in Raptor-depleted but not Rictor-deprived cells. Collectively, these results demonstrated that Raptor and Rictor could play a distinctly regulatory role in rasfonin-enhanced autophagy and apoptosis.

Pestalotiones A-D: Four New Secondary Metabolites from the Plant Endophytic Fungus Pestalotiopsis Theae.

Two new xanthone derivatives, pestalotiones A (1) and B (2), one new diphenyl ketone riboside, pestalotione C (7), and one new diphenyl ether, pestalotione D (8), along with five known compounds isosulochrin dehydrate (3), 3,8-dihydroxy-6-methyl-9-oxo-9H-xanthene-1-carboxylate (4), isosulochrin (5), chloroisosulochrin (6), and pestalotether D (9), were isolated from the crude extract of the plant endophytic fungus Pestalotiopsis theae (N635). The structures of the new compounds were unambiguously deduced by HRESIMS and 1D/2D-NMR spectroscopic data. Compound 6 showed modest cytotoxicity against the HeLa cell line with an IC50 value of 35.2 µM. Compound 9 also showed cytotoxic to the HeLa and MCF-7 cell lines, with IC50 values of 60.8 and 22.6 µM, respectively. Additionally, compounds 1 and 2 exhibited antioxidant activity in scavenging DPPH radical with IC50 values of 54.2 and 59.2 µg/mL, respectively.

A novel high-throughput nematicidal assay using embryo cells and larvae of Caenorhabditis elegans.

Human health safety and environmental concerns have resulted in the widespread deregistration of several agronomic important nematicides. New and safer nematicides are urgently needed. However, a high-throughput bioassay for screening potential nematicides has not been established. We developed a two-step high-throughput nematicidal screening method to combine a cell-based MTS colorimetric assay with Caenorhabditis elegans embryo cells for preliminary cytotoxicity screening (step 1) followed by in vitro larval assay for nematicidal activity (step 2). Based on three conventional nematicides' test, high correlations were obtained between cell viability and larval viability and "r" values were 0.78 for Avermectin, 0.95 for Fosthiazate, and 0.65 for Formaldehyde solution. Further assays with 60 fungal secondary metabolites (extracts, fractions and pure compounds) also demonstrated the high correlation between cell viability and larval viability (r=0.60) and between the C. elegans cell viability and the juvenile viability of soybean cyst nematode Heterodera glycines (r=0.48) and pine wood nematode Bursaphelenchus xylophilus (r=0.56). Six metabolites with high cytotoxicity have performed high larval mortality with a LC50 range of 6.8-500µg/ml. These results indicate that the proposed two-step screening assay represents an efficient and labor-saving method for screening natural nematicidal products.

2H-pyran-2-one and 2H-furan-2-one derivatives from the plant endophytic fungus Pestalotiopsis fici.

Two new α-pyrones (=2H-pyran-2-ones), ficipyrones A and B (1 and 2, resp.), and two new α-furanones (=2H-furan-2-ones), ficifuranones A and B (3 and 4, resp.), together with three known metabolites, antibiotic F 0368 (5), hydroxyseiridin (6), and hydroxyisoseiridin (7), were isolated from solid cultures of the plant endophytic fungus Pestalotiopsis fici. Their structures were elucidated primarily by NMR spectroscopy, and the absolute configuration of 1 was deduced from the circular-dichroism (CD) data. Compound 1 showed antifungal activity against the plant pathogen Gibberella zeae (CGMCC 3.2873) with an IC50 value of 15.9 µM.

The modulatory effects of gut microbes and metabolites on blood-brain barrier integrity and brain function in sepsis-associated encephalopathy.

Background: Intestinal microbiota homeostasis and the gut-brain axis are key players associated with host health and alterations in metabolic, inflammatory, and neurodegenerative disorders. Sepsis-associated encephalopathy (SAE), which is closely associated with bacterial translocation, is a common secondary organ dysfunction and an urgent, unsolved problem affecting patient quality of life. Our study examined the neuroprotective effects of the gut microbiome and short-chain fatty acid (SCFA) metabolites on SAE. Methods: Male C57BL/6 mice were administered SCFAs in drinking water, then subjected to cecal ligation and puncture (CLP) surgery to induce SAE. 16S rRNA sequencing was used to investigate gut microbiome changes. The open field test (OFT) and Y-maze were performed to evaluate brain function. The permeability of the blood-brain barrier (BBB) was assessed by Evans blue (EB) staining. Hematoxylin and eosin (HE) staining was used to examine intestinal tissue morphology. The expression levels of tight junction (TJ) proteins and inflammatory cytokines was assessed by western blots and immunohistochemistry. In vitro, bEND.3 cells were incubated with SCFAs and then with lipopolysaccharide (LPS). Immunofluorescence was used to examine the expression of TJ proteins. Results: The composition of the gut microbiota was altered in SAE mice; this change may be related to SCFA metabolism. SCFA treatment significantly alleviated behavioral dysfunction and neuroinflammation in SAE mice. SCFAs upregulated occludin and ZO-1 expression in the intestine and brain in SAE mice and LPS-treated cerebromicrovascular cells. Conclusions: These findings suggested that disturbances in the gut microbiota and SCFA metabolites play key roles in SAE. SCFA supplementation could exert neuroprotective effects against SAE by preserving BBB integrity.

New furanones from the plant endophytic fungus Pestalotiopsis besseyi.

Pestalafuranones A-E (compounds 1-5), five new 2(5H)-furanones, have been isolated from cultures of an isolate of Pestalotiopsis besseyi. The structures of these compounds were elucidated mainly by analysis of their NMR spectroscopic data and HRESIMS experiments. Pestalafuranones A-C (compounds 1-3) displayed weak inhibitory effects against HIV-1 replication in C8166 cells, whereas pestalafuranones D (4) and E (5) showed moderate antifungal activity against the plant pathogens Verticillium dahiae and Alternaria longipes.

Attenuation of in vitro and in vivo melanin synthesis using a Chinese herbal medicine through the inhibition of tyrosinase activity.

BACKGROUND: In traditional Chinese medicine, the skin reflects the health of body organs. A skin whitening agent, named seven whitening creams (also called Chi-Bai-San), has been used since ancient times in China. Chi-Bai-San reduces melanin and helps to reduce wrinkles. PURPOSE: We aimed to determine the skin-whitening ability and safe dose of the seven compounds in Chi-Bai-San. STUDY DESIGN: A common use for Chinese medicine is decocted in water. To mimic the function of Chi-Bai-San apply in clinical, we boiled all seven compound in water, respectively. These single recipe extractions and a mixture of these seven items were used in zebrafish embryo and B16F10 melanoma cell to identify the anti-melanogenesis function. METHODS: Chi-Bai-San comprises Bai-Lian (Ampelopsis japonica), Bai-Ji (Bletilla striata), Bai-Zhi (Angelica dahurica), Bai-Zhu (Atractylodes macrocephala), Bai-Shau (Paeonia lactiflora), Fu-Ling (Wolfiporia cocos), and Jen-Ju-Fen (Pearl powder). All components were extracted by heating in distilled water. The supernatant was collected after centrifugation. The extracted components were introduced into zebrafish embryos at different doses to determine the safe dose. B16F10 melanoma cells were treated with the final dose of each component and the component mixture. Melanin content and tyrosinase activity were assessed in zebrafish and B16F10 cells. Chi-Bai-San and its components were exposed to α MSH-induced B16F10 cells, and detected for mechanism of anti-melanogenesis pathway. RESULTS: Most compounds were not toxic at a low dose (0.1 mg/ml), except A. macrocephala, which resulted in a survival rate of only 30% at 72 hpf. The final dose of A. dahurica, P. lactiflora, W. cocos, and pearl was 1 mg/ml; that of A. japonica was 0.5 mg/ml; and that of A. macrocephala and B. striata was 0.1 mg/ml. Chi-Bai-San markedly decreased melanin content 37.47% in zebrafish embryos. Further, Chi-Bai-San abolished tyrosinase activity and MITF-mediated tyrosinase expression by down regulating the upstream transcription factors ZEB2, ß-catenin, and CREB2 in α MSH-induced B16F10 cells. Additionally, Chi-Bai-San might reduce melanosome secretion from melanocytes. CONCLUSION: Our findings indicate that safety and efficacy of heat-extracted Chi-Bai-San, which can reduce αMSH-induced melanin production by inhibiting the key role of melogenic-related transcription factor and promote the synergic effect of seven types of traditional Chinese herbal medicines.

The Cycas genome and the early evolution of seed plants.

Cycads represent one of the most ancient lineages of living seed plants. Identifying genomic features uniquely shared by cycads and other extant seed plants, but not non-seed-producing plants, may shed light on the origin of key innovations, as well as the early diversification of seed plants. Here, we report the 10.5-Gb reference genome of Cycas panzhihuaensis, complemented by the transcriptomes of 339 cycad species. Nuclear and plastid phylogenomic analyses strongly suggest that cycads and Ginkgo form a clade sister to all other living gymnosperms, in contrast to mitochondrial data, which place cycads alone in this position. We found evidence for an ancient whole-genome duplication in the common ancestor of extant gymnosperms. The Cycas genome contains four homologues of the fitD gene family that were likely acquired via horizontal gene transfer from fungi, and these genes confer herbivore resistance in cycads. The male-specific region of the Y chromosome of C. panzhihuaensis contains a MADS-box transcription factor expressed exclusively in male cones that is similar to a system reported in Ginkgo, suggesting that a sex determination mechanism controlled by MADS-box genes may have originated in the common ancestor of cycads and Ginkgo. The C. panzhihuaensis genome provides an important new resource of broad utility for biologists.

Two new imidazolone-containing alkaloids and further metabolites from the ascomycete fungus Tricladium sp.

Tricladins A and B (1 and 2, resp.), new imidazolone-containing alkaloids, together with five known metabolites, bacillamides A, B (3 and 4, resp.), 20-hydroxyaflavinine (5), N-(2-phenylethyl)acetamide (6), and N(b)-acetyltryptamine (7), have been isolated from the crude extract of the ascomycete fungus Tricladium sp. The structures of 1 and 2 were elucidated primarily by NMR and MS methods. Compound 2 showed marginal cytotoxicity against MDA-MB-231 human breast cancer cells, whereas the known metabolite 4 displayed an inhibitory effect on HIV-1 replication in C8166 cells.

Erratum: MicroRNA-26b suppresses autophagy in breast cancer cells by targeting DRAM1 mRNA, and is downregulated by irradiation.

[This corrects the article DOI: 10.3892/ol.2017.7452.].

Oxepinochromenones, furochromenone, and their putative precursors from the endolichenic fungus Coniochaeta sp.

Six new polyketides including four oxepinochromenones, conioxepinols A-D (1-4), one furochromenone, coniofurol A (5), and one xanthone, conioxanthone A (6), have been isolated from the crude extract of the endolichenic fungus Coniochaeta sp. The absolute configurations of C-7 in 1 and the 7,8-diol moiety in 3 were assigned using the modified Mosher's and Snatzke's method, respectively, whereas that of C-8 in 5 was deduced via the circular dichroism data of the [Rh(2)(OCOCF(3))(4)] complex. Compounds 2-4 showed modest cytotoxicity against a small panel of human tumor cell lines.

Polyketides from the ascomycete fungus Leptosphaeria sp.

The new polyketides leptosphaerins A-G (1-7) and the known compounds monodictysin B (8), 2-(2,6-dihydroxybenzoyl)-3-hydroxy-5-methylbenzoic acid (9), and 2-(2,6-dihydroxy-4-methylbenzoyl)-6-hydroxybenzoic acid (10) have been isolated from solid cultures of the ascomycete fungus Leptosphaeria sp. The absolute configurations of compounds 1, 2, and 6 were assigned using the modified Mosher method, whereas that of C-2 in 3 was determined via the circular dichroism data of the [Rh(2)(OCOCF(3))(4)] complex. Compound 7 showed antifungal activity against Aspergillus flavus.

The relationships between body mass index and television viewing, internet use and cellular phone use: the moderating effects of socio-demographic characteristics and exercise.

OBJECTIVE: This study aimed to examine the relationships between body mass index (BMI) and television viewing, Internet use and cellular phone use and the moderators for these relationships in adolescents. METHOD: The relationship between BMI and the time spent on three kinds of sedentary activities and the moderators for these relationships were analyzed among 9,278 Taiwanese adolescents. The different relationships between BMI and various Internet and cellular phone-related activities were analyzed. RESULTS: High television viewing and high Internet use were associated with increased BMI in adolescents. Exercising had a moderating effect on the relationship between BMI and television viewing. Several Internet and cellular phone-related activities were associated with increased BMI. DISCUSSION: The results support the relationships between adolescent BMI and television viewing and Internet use. The moderating effect of exercise and various Internet and cellular phone-related activities should be considered when developing intervention strategies for overweight adolescents.

Ultrasound, X-ray, computed tomography and clinical tests for diagnosis of abdominal purpura in children: A retrospective study.

Imaging modalities are not included in The European League Against Rheumatism, The Pediatric Rheumatology International Trials Organization and The Pediatric Rheumatology European Society 2010 criteria for the evaluation of abdominal purpura. The objective of the present study was to compare diagnostic parameters of ultrasound, X-ray and computed tomography (CT) for diagnosis of abdominal purpura considering the American College of Rheumatology (ACR) criteria as 'reference standard' in children with acute abdominal pain. Medical records of 215 children with acute abdominal pain were reviewed. Data regarding demographics and clinical characteristics, laboratory tests, X-ray, ultrasound findings, and computed tomographic images were collected and analyzed. Decision curve analysis was used for evaluation of the beneficial score for each diagnostic modality. Among diagnostic modalities, CT had the highest sensitivity (0.939); however, ultrasound findings had the highest accuracy (0.861) for diagnosis of abdominal purpura. Unlike X-ray and laboratory tests, ultrasound and CT were successful at detecting abdominal purpura when children had only colicky pain and were aged <20 years; however, occult blood stool test and granulocytes in the walls of small venules and arterioles (biopsy results) were negative. With respect to the ACR criteria, there were seven and three inconclusive results for ultrasound and CT, respectively. Abdominal ultrasound is an easy, non-invasive and safe method for the detection of abdominal purpura in children.

[Design and application of pre-hospital "Owl" first aid kit].

Pre-hospital first aid can reduce the mortality and morbidity of critically ill patients, and first aid kit plays an important role in pre-hospital first aid. The first aid kit used in clinic is easy to cause the injection to be broken, and it is inconvenient to use at night. Thus, we have designed an "Owl" first aid kit and obtained a national utility model patent (ZL 2018 2 0727656.7) which includes an injection storage cover and a medicine storage box. A number of injection storage plates are arranged on the injection storage cover, which are used for clamping injections of different sizes, etc., and can effectively prevent injection drugs from being extruded, shedding and broken. There is a supporting rod on one side of the injection storage cover, which can improve the efficiency to take medicine from the first aid kit. There are several partitions in the drug storage box, which can be adjusted according to the size and type of articles placed. A number of symmetrical elastic buckles are arranged on the bottom plate for picking up medicine in bottles, which is convenient for emergency personnel to operate with one hand. There is an illuminating lamp at the edge of the injection storage cover, and a number of fluorescent strips are arranged on the inner wall of the medicine storage box. Based on the above device, it is convenient for medical personnel to pick up medicines at night, which can avoid potential errors effectively. The "Owl" first aid kit has a simple structure and low cost. It can prevent the injection from being broken and convenient to operate. At the same time, it is convenient to be used at night, so it is worthy of clinical application.

Pestalofones A-E, bioactive cyclohexanone derivatives from the plant endophytic fungus Pestalotiopsis fici.

Pestalofones A-E (1-5), five new cyclohexanone derivatives, have been isolated from cultures of the plant endophytic fungus Pestalotiopsis fici, along with the known compounds, isosulochrin (6), isosulochrin dehydrate (7), and iso-A82775C (8). The structures of 1-5 were determined by NMR spectroscopy, and the absolute configuration of 1 was assigned using the modified Mosher method. Compounds 1, 2, and 5 displayed inhibitory effects on HIV-1 replication in C8166 cells, whereas 3 and 5 showed significant antifungal activity against Aspergillus fumigatus.

Pyrrolidinones from the ascomycete fungus Albonectria rigidiuscula.

The new pyrrolidinones, rigidiusculamides A-D (1-4), have been isolated from the crude extract of the ascomycete fungus Albonectria rigidiuscula. The structures of these compounds were elucidated primarily by NMR experiments. The absolute configuration of the 3,4-diol moieties in 1 and 4 was assigned using Snatzke's method. Compounds 1 and 2 showed modest cytotoxicity against the human tumor cell lines HeLa and MCF-7.

Allenyl and alkynyl phenyl ethers from the endolichenic fungus Neurospora terricola.

Terricollenes A-C (1-3), new allenyl phenyl ethers, and terricolynes (4-6), new alkynyl phenyl ethers, have been isolated from the crude extract of the endolichenic fungus Neurospora terricola. The structures of these compounds were elucidated primarily by NMR experiments. The absolute configurations of 1 and 2 were determined by reversed-phase HPLC analysis on the o-tolylthiocarbamates of the liberated sugars upon acid hydrolysis, whereas that of 4 was assigned using the modified Mosher method. Compound 1 showed modest cytotoxicity against the human tumor cell lines HeLa and MCF-7.

Ambuic acid and torreyanic acid derivatives from the endolichenic fungus Pestalotiopsis sp.

Six new ambuic acid (1) derivatives (2-7) and a new torreyanic acid analogue (8) have been isolated from the crude extract of endophytic fungus Pestalotiopsis sp. inhabiting the lichen Multiclavula [corrected] sp. The structures of these compounds were elucidated primarily by NMR and MS methods, and their absolute configurations were assigned by application of the CD excitation chirality method. Compounds 1 and 2 displayed antimicrobial activity against the Gram-positive bacterium Staphylococcus aureus.