Effects of cadmium and pyrene on earthworm-associated bacterial communities: Unveiling new perspectives for soil pollution management.

Earthworms are considered to be excellent bioindicators of soil pollution. In recent years, there has been increasing interest in examining the effects of soil pollution on earthworm-associated microbiomes, with a particular focus on the gut microbiomes. However, relatively little effort has been invested in comprehensively investigating other microbiomes associated with earthworms and their responses to soil pollution. To fill this gap, we systematically studied the effects of Cd, pyrene, and combined pollution on the bacterial community in different vermicompartments, i.e., burrow wall, gut, and cast, in both epigeic Eisenia fetida and anecic Metaphire guillelmi, using a 2D-terraria incubator and high-throughput sequencing techniques. The results showed that bacterial alpha diversity followed the order of burrow wall > cast > gut, and this did not vary with soil pollution or earthworm ecotypes. Moreover, the dominant phyla in the vermicompartments were similar across different pollution treatments. Principal coordinate analysis (PCoA) revealed that the bacterial communities in different vermicompartments and ecotypes of earthworm were separated from each other, whereas they were grouped together in polluted treatments and unpolluted conditions. These results imply that even in polluted soil, vermicompartment and earthworm ecotypes remain the most significant factors affecting earthworm-associated microbiomes. However, the impacts of soil pollution on the bacterial composition in each vermicompartment were still evident. A comprehensive analysis revealed that the gut bacterial communities are more sensitive to soil contamination than casts and burrow wall in different ecotypes. Additionally, linear discriminant analysis of effect size (LefSe) identified several bacteria in Gemmatimonadota, the Firmicutes phylum in the burrow walls, and Patescibacteria (phyla) in the gut as potential biomarkers for pyrene contamination in soil. This research provides a comprehensive understanding of the effects of soil pollution on earthworm-associated microbiomes, thereby enhancing our understanding of earthworm ecotoxicology and soil pollution management.

Cellular signaling crosstalk between Wnt signaling and gap junctions inbenzo[a]pyrene toxicity.

Gap junctional intercellular communication (GJIC) is considered a key biological mechanism to maintain homeostasis in cell differentiation and growth. In addition, as another major signaling pathway associated with cell proliferation and differentiation, Wnt/ß-catenin signaling appears to trigger several cellular responses against injury. The purpose of the present study was to investigate the effects of a known toxic agent, benzo[a]pyrene (BaP), on the regulation and interaction between GJIC and Wnt/ß-catenin signaling. BaP treatment resulted in GJIC inhibition and decreases the major GJIC protein connexin 43 (Cx43) in WB-F344 rat liver epithelial cells. We also found BaP-mediated downregulation of Wnt/ß-catenin signaling related to the PI3K-Akt pathway. To identify the relationship between GJIC and Wnt/ß-catenin signaling, we treated WB-F344 cells with the Wnt agonist CHIR99021 and found that it inhibited GJIC while causing a significant reduction in Cx43 expression at both the mRNA and protein levels, through the repression of promoter activity. This Wnt agonist-mediated GJIC inhibition was confirmed using a small interfering RNA directed against the Wnt antagonist Dact2, indicating that Wnt/ß-catenin signaling negatively regulates GJIC. Despite the inverse correlation between Wnt/ß-catenin signaling and Cx43 promoter activation as indicated by downregulation of ß-catenin nuclear translocation and upregulation of Cx43 promoter activation involving HNF3ß, BaP treatment decreased the Cx43 protein expression, which was associated with protein degradation, possibly through protein kinase C activation. In conclusion, our results revealed the mechanism of BaP-induced inhibition of GJIC and Wnt/ß-catenin signaling. More importantly, linking Wnt/ß-catenin signaling to Cx protein expression will have profound implications in understanding the relationships among different major signaling pathways associated with cell proliferation and differentiation in toxicity.

Wedelolactone, a Component from Eclipta prostrata (L.) L., Inhibits the Proliferation and Migration of Head and Neck Squamous Cancer Cells through the AhR Pathway.

BACKGROUND: Ecliptae prostrata (L.) L. has been widely used in East Asia with reported biological activities, including anti-cancer properties. OBJECTIVES: We aimed to investigate the effect of ethyl acetate extract of Ecliptae prostrata (L.) L. (EAE) and its component wedelolactone on the proliferation and migration of head and neck squamous cancer cells. METHODS: The proliferation of human SCC-4 and mouse CU110-1 tongue squamous carcinoma cells was assessed using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) method. Scratch wound assays were performed to assess cell migration rates. The levels of Ecadherin and vimentin were used as markers of the epithelial-to-mesenchymal transition (EMT). AhR, CYP1A1, and CYP1B1 levels were examined to uncover the mechanism of inhibition of cell migration by wedelolactone. RESULTS: We found that EAE and wedelolactone decreased the proliferation of human SCC-4 cells and mouse CU110-1 cells at doses of EAE at > 25 µg/ml and wedelolactone at > 6.25 µg/ml. Similarly, both EAE and wedelolactone produced inhibitory effects against migration, but the effective doses that significantly inhibited migration were lower than those affecting proliferation. Wedelolactone below 12.5 µg/ml inhibited the epithelial-to-mesenchymal transition (EMT) with increased expression of E-cadherin and decreased expression of vimentin in SCC-4 and CU110-1 cells. Further analysis showed wedelolactone inhibited the expression of AhR and its downstream target molecules CYP1A1 and CYP1B1 in both squamous carcinoma cells at the same doses inhibiting cell migration. The addition of benzo (a)pyrene [B(a)P], an agonist of AhR, stimulated migration, especially in the CU110-1 cells with reported cancer stem cell-like characteristics. Instructively, B(a)P reversed the inhibitory effects of wedelolactone on AhR expression and cell migration, suggesting that wedelolactone antagonizes cell migration through the AhR pathway. Moreover, the higher activity of EAE and wedelolactone against the migration of cancer stem-like CU110-1 cells relative to SCC-4 cells suggests selective activity against cancer stem cells. CONCLUSION: Our study identifies wedelolactone as a major active component of Ecliptae prostrata (L.) L. with promising anti-cancer properties against head and neck squamous cancer cells.

Effect of pyrene and phenanthrene in shaping bacterial communities in seagrass meadows sediments.

Polycyclic aromatic hydrocarbons (PAHs), originating from anthropogenic and natural sources, are highly concerned environmental pollutants. This study investigated the impact of two model PAHs (pyrene and phenanthrene) on bacterial community succession in the seagrass meadows sediment in a lab-scale microcosm. Halophila ovalis sediment slurry microcosms were established, one group was placed as a control, and the other two were treated with pyrene and phenanthrene. Bacterial community succession in response to respective PAHs was investigated by 16S rRNA amplicon sequencing. The results demonstrated that bacterial diversity decrease in each microcosm during the incubation process; however, the composition of bacterial communities in each microcosm was significantly different. Proteobacteria (37-89%), Firmicutes (9-41%), and Bacteroides (7-21%) were the predominant group at the phylum levels. Their abundance varies during the incubation process. Several previously reported hydrocarbon-degrading genera, such as Pseudomonas, Spinghobium, Sphingobacterium, Mycobacterium, Pseudoxanthomonas, Idiomarina, Stenotrophomonas, were detected in higher abundance in pyrene- and phenanthrene-treated microcosms. However, these genera were distinctly distributed in the pyrene and phenanthrene treatments, suggesting that certain bacterial groups favorably degrade different PAHs. Statistical analyses, such as ANOSIM and PERMANOVA, also revealed that significant differences existed among the treatments' bacterial consortia (P < 0.05). This work showed that polycyclic aromatic hydrocarbon significantly affects bacterial community succession, and different PAHs might influence the bacterial community succession differently.

Toxic mechanism of pyrene to catalase and protective effects of vitamin C: Studies at the molecular and cell levels.

Polycyclic aromatic hydrocarbons, distributing extensively in the soil, would potentially threaten the soil organisms (Eisenia fetida) by triggering oxidative stress. As a ubiquitous antioxidant enzyme, catalase can protect organisms from oxidative damage. To reveal the potential impact of polycyclic aromatic hydrocarbon pyrene (Pyr) on catalase (CAT) and the possible protective effect of Ascorbic acid (vitamin C), multi-spectral and molecular docking techniques were used to investigate the influence of structure and function of catalase by pyrene. Fluorescence and circular dichroism analysis showed that pyrene would induce the microenvironmental changes of CAT amino acid residues and increase the α-helix in the secondary structure. Molecular simulation results indicated that the main binding force of pyrene around the active center of CAT is hydrogen bonding force. Furthermore, pyrene inhibited catalase activity to 69.9% compared with the blank group, but the degree of inhibition was significantly weakened after vitamin C added into the research group. Cell level experiments showed that pyrene can increase the level of ROS in the body cavity cell of earthworms, and put the cells under the threat of potential oxidative damage. Antioxidants-vitamin C has a protective effect on catalase and maintains the stability of intracellular ROS levels to a certain extent.

Application of self-assembly peptides targeting the mitochondria as a novel treatment for sorafenib-resistant hepatocellular carcinoma cells.

Currently, there is no appropriate treatment option for patients with sorafenib-resistant hepatocellular carcinoma (HCC). Meanwhile, pronounced anticancer activities of newly-developed mitochondria-accumulating self-assembly peptides (Mito-FF) have been demonstrated. This study intended to determine the anticancer effects of Mito-FF against sorafenib-resistant Huh7 (Huh7-R) cells. Compared to sorafenib, Mito-FF led to the generation of relatively higher amounts of mitochondrial reactive oxygen species (ROS) as well as the greater reduction in the expression of antioxidant enzymes (P < 0.05). Mito-FF was found to significantly promote cell apoptosis while inhibiting cell proliferation of Huh7-R cells. Mito-FF also reduces the expression of antioxidant enzymes while significantly increasing mitochondrial ROS in Huh7-R cells. The pro-apoptotic effect of Mito-FFs for Huh7-R cells is possibly caused by their up-regulation of mitochondrial ROS, which is caused by the destruction of the mitochondria of HCC cells.

Development of pyrene-based fluorescent ether lipid as inhibitor of SK3 ion channels.

We report the synthesis of three bioactive pyrene-based fluorescent analogues of Ohmline which is the most efficient and selective inhibitor of SK3 ion channel. The interaction of these Ohmline-pyrene (OP1-3) with liposomes of different composition reveals that only OP2 and OP3 are readily integrated into liposomes. Fluorescence measurements indicate that, depending on their concentration, OP2 and OP3 exist either as monomer or as a mixture of monomer and excimers within the liposome bilayer. Among the three Ohmline Pyrene compounds (OP1-3) only OP2 is able to reduce SK3 currents and is the first efficient fluorescent modulator of SK3 channel as revealed by patch clamp measurements (- 71.3 ± 13.3% at 10 µM) and by its inhibition of SK3-dependent cancer cell migration at (-32.5% ± 4.8% at 1 µM). We also report the first fluorescence study on living breast cancer cells (MDA-MB-231) showing that OP2 is rapidly integrated in bio-membranes followed by cell internalization.

Phase-selective staining of model and cell membranes, lipid droplets and lipoproteins with fluorescent solvatochromic pyrene probes.

The push-pull solvatochromic pyrene derivatives PA and PK have been applied to the study of model membrane vesicles, cells and purified human serum lipoproteins, using both confocal fluorescence microscopy and fluorescence spectroscopy. These polarity-sensitive probes provide information similar to that obtained by Laurdan or Prodan, i.e. mainly lipid order in biomembranes, but they have the essential advantage of being excitable by a standard 405 nm laser light, bypassing the use of multiphoton excitation. In addition, they are brighter and much more photostable than those dimethylamino naphthalene derivatives. Our results with model membrane spectroscopy (multilamellar vesicles) and with microscopy (giant unilamellar vesicles) showed the capacity of PA and PK to report differently on liquid-disordered, liquid-ordered and gel phase bilayers. Moreover, a ratiometric parameter, the Red/Blue Intensity Ratio (RBIR) could be used for inter-domain, inter-vesicle and even inter-technique comparison, and the appropriate microscopy-spectroscopy conversion coefficients could be estimated. In studies at the cellular level, PA probe stained almost exclusively the plasma membrane of red blood cells, revealing its high degree of lipid order. Using Chinese Hamster Ovary cells PA was shown to be an excellent probe for the detection of cytoplasmic lipid droplets, superior to Nile Red in that PA provides simultaneously a detailed information of membrane order in the whole cell, in which the lipid droplets appear with a very good contrast. Moreover, spectrofluorometric data of PA-stained serum lipoproteins indicated an essentially identical value of RBIR for lipid droplets and for high-density lipoproteins.

1-Methylpyrene induces chromosome loss and mitotic apparatus damage in a Chinese hamster V79-derived cell line expressing both human CYP1A2 and sulfotransferase 1A1.

1-Methylpyrene (1-MP) is a ubiquitous environmental pollutant and rodent carcinogen. Its mutagenic activity depends on sequential activation by various CYP and sulfotransferase (SULT) enzymes. Previously we have observed induction of micronuclei and mitotic arrest by 1-MP in a Chinese hamster (V79)-derived cell line expressing both human CYP1A2 and SULT1A1 (V79-hCYP1A2-hSULT1A1), however, the mode of chromosome damage and the involvement of mitotic tubulin structures have not been clarified. In this study, we used immunofluorescent staining of centromere protein B (CENP-B) with the formed micronuclei, and that of ß- and γ-tubulin reflecting the structures of mitotic spindle and centrioles, respectively, in V79-hCYP1A2-hSULT1A1 cells. The results indicated that 1-MP induced micronuclei in V79-hCYP1A2-hSULT1A1 cells from 0.125 to 2 µM under a 24 h/0 h (exposure/recovery) regime, while in the parental V79-Mz cells micronuclei were induced by 1-MP only at concentrations ≥ 8 µM; in both cases, the micronuclei induced by 1-MP were predominantly CENP-B positive. Following 54 h of exposure, 1-MP induced mitotic spindle non-congression and centrosome amplification (multipolar mitosis) in V79-hCYP1A2-hSULT1A1 cells, and anaphase/telophase retardation, at concentrations ≥ 0.125 µM with concentration-dependence; while in V79-Mz cells it was inactive up to 8 µM. This study suggests that in mammalian cells proficient in activating enzymes 1-MP may induce chromosome loss and mitotic disturbance, probably by interfering with the mitotic spindle and centrioles.

Novel tetranuclear PdII and PtII anticancer complexes derived from pyrene thiosemicarbazones.

Cyclometallated palladium(ii) and platinum(ii) pyrenyl-derived thiosemicarbazone (H2PrR) complexes of the type [M4(µ-S-PrR-κ3-C,N,S)4] (M = PdII, PtII; R = ethyl, cyclohexyl) have been synthesised in good yields and fully characterised. X-ray crystallography showed that the tetranuclear complex [Pt4(µ-S-PrCh-κ3-C,N,S)4](CH3)2COCHCl3 contains an eight-membered ring of alternating M-S atoms. The ethyl derivatives [M4(µ-S-PrEt-κ3-C,N,S)4] exhibit potent antiproliferative activity towards A2780 human ovarian cancer cells, with IC50 values of 1.27 µM (for M = PdII) and 0.37 µM (for M = PtII), the latter being an order of magnitude more potent than the anticancer drug cisplatin (IC50 1.20 µM). These promising complexes had low toxicity towards non-cancerous human MRC5 cells, which points towards an early indication of differential toxicity between cancer and normal cells. Experiments that investigated the effects of these tetranuclear complexes on the cell cycle, integrity of the cell membrane, and induction of apoptosis, suggested that their mechanism of action of does not involve DNA targeting, unlike cisplatin, and therefore could be promising for combatting cisplatin resistance.

Intriguing H-Aggregates of Heptamethine Cyanine for Imaging-Guided Photothermal Cancer Therapy.

Organic small-molecule-based photothermal agents such as cyanine dyes have received increasing attention in developing novel cancer therapies with potential clinical utility but suffer from poor stability, low photothermal efficiency, and limited accumulation at tumor sites in molecular forms. Self-assembly of small-molecule dyes into supramolecular assemblies may address these concerns by controlling the molecular organization of dye monomers to form structures of a higher order. Among them, H-aggregates of dyes favor face-to-face contacts with strongly overlapping areas, which always have a negative connotation to exhibit low or no fluorescence in most cases but may emanate energy in nonradiative forms such as heat for photothermal cancer therapy applications. Here, the synergistic self-assembly of cyanine dyes into H-aggregates is developed as a new supramolecular strategy to fabricate small-molecule-based photothermal nanomaterials. Compared to the free cyanine dyes, the H-aggregates assembled from pyrene or tetraphenylethene (TPE) conjugating cyanine exhibit the expected absorption spectral blue shift and fluorescence self-quenching but unique photothermal properties. Remarkably, the obtained H-aggregates are saucer-shaped nanoparticles that exhibit passive tumor-targeting properties to induce imaging-guided photothermal tumor ablation under irradiation. This supramolecular strategy presented herein may open up new opportunities for constructing next-generation small-molecule-based self-assembly nanomaterials for PTT cancer therapy in clinics.

Intercalating pyrene with polypeptide as a novel self-assembly nano-carrier for colon cancer suppression in vitro and in vivo.

Giving patients right dosage is an essential concept of precision medicine. Most of nanocarriers lack of flexible drug capacity and structural stability to be customized for specific treatment, resulting in low therapeutic efficacy and unexpected side effects. Thus, a growing need emerges for fast and rigorous approaches to develop nanoparticles with properties of adjustable dosage and controllable particle size. Poly-l-Lysine is known for its enhanced bioadhesivity and pH-triggered structural swelling effect, which is utilized as the main agent to activate the multistage drug releasing. Inspired by natural bio-assembly system, we report a simple method to self-assemble Poly-l-Lysine-based nanoparticles via supramolecular recognitions of cross-linked pyrenes, which provides noncovalent force to flexibly encapsulate Doxorubincin and to construct robust nanostructures. Pyrene-modified polypeptide self-assemblies are able to adjust drug payload from 1: 10 to 2:1 (drug: polypeptide) without changing its uniform nano-spherical morphology. This nanostructure remained the as-made morphology even after experiencing the long-term (~ 10 weeks) storage at room temperature. Also, the nanoparticles displayed multi-step drug release behaviours and exhibited great in vitro and in vivo cytotoxicity towards colon cancer cells. The as-mentioned nanoparticles provide a novel perspective to compensate the clinical needs of specific drug feedings and scalable synthesis with advantages of simple-synthesis, size-adaptivity, and morphology reversibility.

Indeno[1,2,3-cd]pyrene and picene mediate actions via estrogen receptor α signaling pathway in in vitro cell systems, altering gene expression.

Currently, the environmental impact of ubiquitous plastic debris triggered quite some public attention. However, the global impact of microplastic on human health is by and large either unknown or neglected. By looking at the underlying biochemical mechanisms leading to the global health threat microplastic was discovered to carry persistent organic pollutants, such as polycyclic aromatic hydrocarbons (PAH), to marine life. The effect of microplastic-ingestion in the human body remains unfortunately somewhat elusive as of yet. For this reason, we screened for compounds binding to the human estrogen receptor α (ERα) and identified the PAH compounds indeno[1,2,3-cd]pyrene (Indpy) and picene (Pice) with a high binding affinity. We applied next generation sequencing to analyze the differentially expressed genes in MCF-7 cells after treatment with Indpy and Pice. We found 8 upregulated genes: ABCC5, CCNG2, CYP1A1, DDIT4, IER3, RUNX2, STC2, and SLC7A5 and 14 downregulated genes: ADORA1, CEBPB, CELSR2, CTSD, CXCL12, KRT19, PGR, PKIB, RARA, RET, SEMA3B, SIAH2, TFAP2C, and XBP1 induced by both ligands and associated with ESR1-regulation. The altered gene expression may influence cell proliferation and metastasis, favoring cancer development with a poor response to therapy. In addition, we confirmed the binding of Indpy and Pice to ERα using molecular docking and microscale thermophoresis. ERα activation was measured with ESR1-overexpressing HEK293 (HEK-ESR1) cells and confirmed for Indpy. In conclusion, we showed an ESR1-mediated influence of the PAH compounds Indpy and Pice on the gene expression pattern of MCF-7 cells, possibly also promoting breast cancer development in patients.

The effect of hypoxia on the induction of strand breaks in plasmid DNA by alpha-, beta- and Auger electron-emitters 223Ra, 188Re, 99mTc and DNA-binding 99mTc-labeled pyrene.

INTRODUCTION: Radiation-induced DNA damage occurs from direct and indirect effects. The induction is influenced by the physical characteristics of the radionuclide, especially its linear energy transfer. Hypoxia reduces the effect of irradiation treatment in tumor cells and leads to poor patient outcomes. High linear energy transfer emitters can overcome this obstacle. Our aim is to demonstrate the influence of hypoxia on the interaction of different radiation qualities with isolated DNA. METHODS: PuC19 Plasmid DNA was irradiated with 223Ra, 188Re, 99mTc and 99mTc-labeled pyrene with and without DMSO under hypoxia or normoxic conditions. DNA damages in form of single-(SSB) and double-strand breaks (DSB) were analyzed by gel electrophoresis. RESULTS: Radiation doses up to 200 Gy of 223Ra, 188Re and 99mTc led to maximal yields of 80% SSB and 30%, 28% and 32% DSB, respectively. Hypoxia had minor effects on damages from 223Ra, but caused a small enhancement in DSB for 188Re and 99mTc. DMSO prevented DSB completely and reduced SSB from the "free" radionuclides to comparable levels. DNA-binding 99mTc-labeled pyrene induced less SSB and DSB compared to [99mTc]TcO4-. However, the incubation with DMSO could prevent the SSB and DSB induction only to a minor extent. CONCLUSIONS: Hypoxia does not limit DNA damage induced by 223Ra, 188Re, 99mTc and 99mTc-labeled pyrene. Dose-dependent radiation effects were comparable for alpha-emitters and both high- and low-energy electron emitters. The radioprotection by DMSO was not influenced by hypoxia. The results indicate the contribution of mainly indirect radiation effects for 99mTc, 188Re and 223Ra. 99mTc-labeled pyrene caused direct DNA damages and Auger-electrons from 99mTc-labeled pyrene are more effective than high-energy electrons or alpha particles. ADVANCES IN KNOWLEDGE: Without the consideration of DNA repair mechanisms, oxygen has no direct influence in radiation-induced DNA damages by different radiation qualities. IMPLICATIONS FOR PATIENT CARE: The short-time stimulation with oxygen during patient radiation could have minor influence compared to constant oxygen flooding to overcome hypoxic barriers.

Effects of Pyrene on Human Liver HepG2 Cells: Cytotoxicity, Oxidative Stress, and Transcriptomic Changes in Xenobiotic Metabolizing Enzymes and Inflammatory Markers with Protection Trial Using Lycopene.

Pyrene is one of the major polycyclic aromatic hydrocarbons formed during heat treatment of meat and in car exhausts; however, few studies have investigated pyrene-induced adverse effects on human cell lines. This study aimed at the investigation of pyrene-induced cytotoxicity and oxidative damage in human liver HepG2 cells at environmentally relevant concentrations. Pyrene-induced changes in mRNA expression of xenobiotic metabolizing enzymes (XMEs), xenobiotic transporters, antioxidant enzymes, and inflammatory markers were investigated using real-time PCR. As a protection trial, the ameliorative effects of lycopene, a carotenoid abundantly found in tomato, were investigated. The possible mechanisms behind such effects were examined via studying the co exposure effects of pyrene and lycopene on regulatory elements including the aryl hydrocarbon receptor (Air) and elytroid 2-related factor 2 (RF). The achieved results indicated that pyrene caused significant cytotoxicity at 50 n, with a clear production of reactive oxygen species (ROS) in a dose-dependent manner. Pyrene upregulated mRNA expression of phase I enzymes including CYP1A1, 1A2, and CYP1B1 and inflammatory markers including TNFα and Cox2. However, pyrene significantly downregulated phase II enzymes, xenobiotic transporters, and antioxidant enzymes. Interestingly, lycopene significantly reduced pyrene-induced cytotoxicity and ROS production. Moreover, lycopene upregulated detoxification and antioxidant enzymes, probably via its regulatory effects on Air- and RF-dependent pathways.

Impact of temperature and pyrene exposure on the functional response of males and females of the copepod Calanus finmarchicus.

We know very little about the effects of two global stressors, elevated temperature and contaminants, on the grazing of marine copepods. To address this issue, we tested the hypotheses that the individual and combined effects of these two stressors may reduce grazing rates and may depend on food availability and gender. We exposed male and female Calanus finmarchicus copepods to pyrene at two temperatures (10 and 14 °C) and six food concentrations (25-800 µg C Rhodomonas baltica L-1) and measured fecal pellet size, and grazing rate (GR) from pellet production. Males had smaller fecal pellets and lower GR than did females. Temperature and pyrene exposure had no effect on pellet size. Temperature alone had no effect on GR of males, but females had lower GR at elevated temperature. Pyrene-exposed males and females had lower GR only at the food concentrations of 200-800 µg C R. baltica L-1 and those patterns were independent of temperature. Pyrene-induced reduction in GR was stronger in females than in males. The negative effects of both elevated temperature and pyrene may reduce the abundance and trophic success of C. finmarchicus in a warmer, more polluted future.

Soluble epoxide hydrolase inhibitor, TUPS, attenuates isoproterenol/angiotensin II-induced cardiac hypertrophy through mammalian target of rapamycin-mediated autophagy inhibition.

OBJECTIVES: To investigate the potential role and mechanism of TUPS, a soluble epoxide hydrolase inhibitor, in cardiac hypertrophy. METHODS: Rat and H9C2 cell models of cardiac hypertrophy were induced by isoproterenol and angiotensin II, respectively, followed by TUPS treatment. The expression of hypertrophic markers, ANP and BNP, was determined by quantitative real-time PCR. The abundance of Beclin-1, LC3, p-AMPK and phosphorylated-mammalian target of rapamycin (p-mTOR) proteins was analysed by Western blot and immunohistocytology. Cell morphology and viability were evaluated by F-actin staining and MTS. H9C2 cells were transfected with GFP-LC3 to evaluate autophagy flux. KEY FINDINGS: TUPS significantly inhibited rat heart size, heart weight-to-body weight ratio, heart wall thickness, hypertrophic H9C2 cell swelling and viability suppression as well as the expression of ANP and BNP genes in hypertrophic models. In addition, autophagic markers Beclin-1 and LC3 were elevated in both cellular and animal models, which were suppressed by TUPS, with corresponding changes of autophagy flux. The abundance of p-AMPK was increased, while p-mTOR was decreased in hypertrophic cells, which were abolished by TUPS. Rapamycin decreased p-mTOR level, increased Beclin-1 and LC3 expression and induced cell size enlargement and cell viability inhibition in hypertrophic H9C2 cells treated with TUPS. CONCLUSIONS: TUPS inhibits cardiac hypertrophy by regulating mTOR/autophagy axis.

In vitro and in silico approaches of antibiofilm activity of 1-hydroxy-1-norresistomycin against human clinical pathogens.

In the present study, an attempt has been made to explore the antibiofilm activity of bioactive compound 1-hydroxy-1-norresistomycin (HNM) derived from coral mucus associated actinomycete Streptomyces variabilis. Initially, different concentration of HNM inhibited the biofilm formation of human clinical pathogens Escherichia coli, Vibrio cholerae and Staphylococcus aureus was determined using crystal-violet staining assay. The light microscopic analysis showed that HNM reduced the biofilm formation and adherence of bacterial cells on the surface of coverslip. HNM also damages the 3D architecture with reduced thickness as well as cell aggregation of biofilm forming bacteria analysed by confocal laser scanning microscopy (CLSM). In addition, HNM also demonstrated the efficiency in inhibiting theoretical adhesion by altering the surface hydrophobicity that can potentially hamper cellular adhesion and prevent biofilm formation. Furthermore, the molecular docking showed the significant interaction between HNM and key biofilm forming proteins proved an excellent antibiofilm activity of HNM. Together, these results suggest that the HNM can serve as potential antibiofilm agent in controlling the infections of E. coli, V. cholerae and S. aureus.

Evidence of selective activation of aryl hydrocarbon receptor nongenomic calcium signaling by pyrene.

In its classical genomic mode of action, the aryl hydrocarbon receptor (AhR) acts as a ligand activated transcription factor regulating expression of target genes such as CYP1A1 and CYP1B1. Some ligands may also trigger more rapid nongenomic responses through AhR, including calcium signaling (Ca2+). In the present study we observed that pyrene induced a relatively rapid increase in intracellular Ca2+-concentrations ([Ca2+]i) in human microvascular endothelial cells (HMEC-1) and human embryonic kidney cells (HEK293) that was attenuated by AhR-inhibitor treatment and/or transient AhR knockdown by RNAi. In silico molecular docking based on homology models, suggested that pyrene is not able to bind to the human AhR in the agonist conformation. Instead, pyrene docked in the antagonist conformation of the AhR PAS-B binding pocket, although the interaction differed from antagonists such as GNF-351 and CH223191. Accordingly, pyrene did not induce CYP1A1 or CYP1B1, but suppressed CYP1-expression by benzo[a]pyrene (B[a]P) in HMEC-1 cells, confirming that pyrene act as an antagonist of AhR-induced gene expression. Use of pharmacological inhibitors and Ca2+-free medium indicated that the pyrene-induced AhR nongenomic [Ca2+]i increase was initiated by Ca2+-release from intracellular stores followed by a later phase of extracellular Ca2+-influx, consistent with store operated calcium entry (SOCE). These effects was accompanied by an AhR-dependent reduction in ordered membrane lipid domains, as determined by di-4-ANEPPDHQ staining. Addition of cholesterol inhibited both the pyrene-induced [Ca2+]i-increase and alterations in membrane lipid order. In conclusion, we propose that pyrene binds to AhR, act as an antagonist of the canonical genomic AhR/Arnt/CYP1-pathway, reduces ordered membrane lipid domains, and activates AhR nongenomic Ca2+-signaling from intracellular stores.

PyA-cluster system for the detection and imaging of miRNAs in living cells through double-three-way junction formation.

Herein, we describe an extended version of a fluorescence probe for detecting miRNAs through the novel application of a PyA-cluster system. By testing various (CG)n sequences in the middle of the oligonucleotide strand of the probe, we obtained an optimal sequence that formed a double-three-way-junction structure, with two PyA units positioned close together, in the presence of the target miRNA. This system readily detected the locations of target miRNAs in living cells and allowed visualization of structural changes through variations in the color of the fluorescence.