Next-generation sequencing-based analysis of the effect of N6-methyldeoxyadenosine modification on DNA replication in human cells

N⁶-methyldeoxyadenosine（6 mdA） modification is considered as a new epigenetic mark that may play important roles in various biological processes.However,it remains unclear about the effect of 6 mdA on DNA replication in human cells.Herein,we combined next-generation sequencing with shuttle vector technology to explore how 6 mdA affects the efficiency and accuracy of DNA replication in human cells.Our results showed that 6 mdA neither blocked DNA replication nor induced mutations in hu...     

Specific detection and effective inhibition of a single bacterial species <Emphasis Type="Italic">in situ</Emphasis> using peptide mineralized Au cluster probes

Increasingly serious microbial infections call for the development of new simpler methods for the precise diagnosis and specific inhibition of such pathogens. In this work, a peptide mineralized Au cluster probe was applied as a new simplified strategy to both recognize and inhibit a single bacteria species of Staphylococcus aureus (S. aureus) simultaneously. The probes are composed of peptides and Au clusters. Moreover, the peptides specifically target S. aureus cells and the Au clusters provide fluorescent imaging and have an antibacterial effect. These new probes enable the simultaneous specific detection and effective destruction S. aureus cells in situ.     

Discovery of Leptulipin,a New Anticancer Protein from theIranian Scorpion,Hemiscorpius lepturus

Cancer is one of the leading causes of mortality in the world. Unfortunately, the present anticancer chemotherapeutics display high cytotoxicity. Accordingly, the discovery of new anticancer agents with lower side effects is highly necessitated. This study aimed to discover an anticancer compound from Hemiscorpius lepturus scorpion venom. Bioactivity-guided chromatography was performed to isolate an active compound against colon and breast cancer cell lines. 2D electrophoresis and MALDI-TOF were performed to identify the molecule. A partial protein sequence was obtained by mass spectrometry, while the full-length was deciphered using a cDNA library of the venom gland by bioinformatics analyses and was designated as leptulipin. The gene was cloned in pET-26b, expressed, and purified. The anticancer effect and mechanism action of leptulipin were evaluated by MTT, apoptosis, and cell cycle assays, as well as by gene expression analysis of apoptosis-related genes. The treated cells displayed inhibition of cell proliferation, altered morphology, DNA fragmentation, and cell cycle arrest. Furthermore, the treated cells showed a decrease in BCL-2 expression and an increase in Bax and Caspase 9 genes. In this study, we discovered a new anticancer protein from H. lepturus scorpion venom. Leptulipin showed significant anticancer activity against breast and colon cancer cell lines.     

Synthesis and Evaluation of Thymol-Based Synthetic Derivatives as Dual-Action Inhibitors against Different Strains of H. pylori and AGS Cell Line

Following a similar approach on carvacrol-based derivatives, we investigated the synthesis and the microbiological screening against eight strains of H. pylori, and the cytotoxic activity against human gastric adenocarcinoma (AGS) cells of a new series of ether compounds based on the structure of thymol. Structural analysis comprehended elemental analysis and ¹H/¹³C/¹⁹F NMR spectra. The analysis of structure–activity relationships within this molecular library of 38 structurally-related compounds reported that some chemical modifications of the OH group of thymol led to broad-spectrum growth inhibition on all isolates. Preferred substitutions were benzyl groups compared to alkyl chains, and the specific presence of functional groups at para position of the benzyl moiety such as 4-CN and 4-Ph endowed the most anti-H. pylori activity toward all the strains with minimum inhibitory concentration (MIC) values up to 4 µg/mL. Poly-substitution on the benzyl ring was not essential. Moreover, several compounds characterized by the lowest minimum inhibitory concentration/minimum bactericidal concentration (MIC/MBC) values against H. pylori were also tested in order to verify a cytotoxic effect against AGS cells with respect to 5-fluorouracil and carvacrol. Three derivatives can be considered as new lead compounds alternative to current therapy to manage H. pylori infection, preventing the occurrence of severe gastric diseases. The present work confirms the possibility to use natural compounds as templates for the medicinal semi-synthesis.     

In Vitro Pharmacological Screening of Essential Oils from Baccharis parvidentata and Lippia origanoides Growing in Brazil

In this study, the in vitro antimicrobial, antiparasitic, antiproliferative and cytotoxic activities of essential oil from Baccharis parvidentata Malag. (EO-Bp) and Lippia origanoides Kunth (EO-Lo) were explored. The relevant effects were observed against the parasitic protozoans Plasmodium falciparum, Trypanosoma cruzi, Trypanosoma brucei and Leishmania amazonensis (ranging 0.6 to 39.7 µg/mL) and malignant MCF-7, MCF-7/HT, 22Rv1, and A431 cell lines (ranging 6.1 to 31.5 µg/mL). In parallel, EO-Bp showed better selective indexes in comparison with EO-Lo against peritoneal macrophages from BALB/c mice and MRC-5 cell line. In conclusion, EO-Lo is known to show a wide range of health benefits that could be added as another potential use of this oil with the current study. In the case of EO-Bp, the wide spectrum of its activities against protozoal parasites and malignant cells, as well as its selectivity in comparison with non-malignant cells, could suggest an interesting candidate for further tests as a new therapeutic alternative.     

Effects of water-based gel electrolyte on the charge recombination and performance of dye-sensitized solar cells

A new water-based solution of ion-conductive polymeric gel electrolyte composed of polyethylene glycol and polyvinylpyrrolidone as gel-forming substances, I^?/I₃ ^? as reversible redox couple, and various ratios of acetonitrile/water solvents was prepared and used in the fabrication of dye-sensitized solar cells. The effects of water on the electrochemical behavior of the prepared electrolyte solutions were examined by the cyclic voltammetry and electrochemical impedance spectroscopy techniques. Electrochemical impedance spectroscopy was employed to quantify the charge-transfer resistance and the electron lifetime at the TiO₂ conduction band. The characteristic peak shifted to a lower frequency in the Bode phase plot, which is an indication of a longer electron lifetime for the cell containing more water content. Photovoltaic performance of the cells prepared by the new water-based gel electrolyte was studied. Changes in the current density–voltage (J–V) characteristics can be explained based on the effect of water on the energetics and kinetics of charge transport and charge recombination in the dye-sensitized solar cells (DSSCs). It was observed that the increase in open-circuit voltage (V _oc) and fill factor and decrease in J _SC were noticeable for cells containing water-based gel electrolyte. It was indicated that the charge recombination between injected electrons and electron acceptors (polyiodide) in the redox electrolyte was remarkably inhibited by the increase of water. The photovoltaic performance stability of the DSSC containing gel electrolyte solution including 50 wt% of water was examined, and it was shown that it is more stable than conventional cells considerably for 168 h. Energy conversion efficiency of 2.30 % was achieved, under illumination with a simulated solar light of 100 mW cm^?2.     

Novel benzimidazole derivatives as anti-cervical cancer agents of potential multi-targeting kinase inhibitory activity

Multi-target EGFR, HER2, VEGFR-2 and PDGFR is an improved strategy for the treatment of solid tumors. This work deals with synthesis of an array of new 6-benzoyl benzimidazole derivatives utlizing1-(6-benzoyl-2-(3,4-dimethoxyphenyl)-1H benzo[d] imidazol-1-yl)propan-2-one (1) as a starting compound. The new compounds were screened as cytotoxic agents against cervical cancer cells (Hela) and Doxorubicin served as a reference drug. Most of the tested compounds showed promising anticancer activity in addition to their safety towards the normal cell line. The most potent candidates were evaluated as EGFR, HER2, PDGFR-β and VEGFR2 inhibitors in comparison to Erlotinib. Compounds 9 and 13 exhibited promising suppression effects. Also, the latter compounds exhibited their ability to induce cellular apoptosis alongside cell cycle arrest at the G2/M phase and accumulation of cells in pre-G1 phase. Molecular docking analysis suggested that compounds 2c, 3f, 9, 12 and 13 tightly interacts with the amino acid residues in the active binding site of HER2 kinase.     

Real Time Anti-<em>Toxoplasma gondii </em>Activity of an Active Fraction of <em>Eurycoma longifolia</em> Root Studied by <em>in Situ</em> Scanning and Transmission Electron Microscopy

The inhibitory effect of active fractions of Eurycoma longifolia (E. longifolia) root, namely TAF355 and TAF401, were evaluated against Toxoplasma gondii (T. gondii). In our previous study, we demonstrated that T. gondii was susceptible to TAF355 and TAF401 with IC₅₀ values of 1.125 μg/mL and 1.375 μg/mL, respectively. Transmission (TEM) and scanning electron microscopy (SEM) observations were used to study the in situ antiparasitic activity at the IC₅₀ value. Clindamycin was used as positive control. SEM examination revealed cell wall alterations with formation of invaginations followed by completely collapsed cells compared to the normal T. gondii cells in response to the fractions. The main abnormality noted via TEM study was decreased cytoplasmic volume, leaving a state of structural disorganization within the cell cytoplasm and destruction of its organelles as early as 12 h of treatment, which indicated of rapid antiparasitic activity of the E. longifolia fractions. The significant antiparasitic activity shown by the TAF355 and TAF401 active fractions of E. longifolia suggests their potential as new anti-T. gondii agent candidates.      

Depsides and depsidones from the soil-derived fungus Aspergillus unguis PSU-RSPG204

Three new depsides, aspergisides A-C, and one new depsidone, aspergisidone, were isolated from the soil-derived fungus Aspergillus unguis PSU-RSPG204 together with one phthalide derivative and 11 depsidones including emeguisin A, aspersidone and folipastatin. The structures were determined by spectroscopic evidence. Aspersidone and emeguisin A showed remarkably antibacterial activity against Staphylococcus aureus and methicillin-resistance S. aureus with equal MIC values of 0.5?μg/mL. Emeguisin A, which displayed the highest activity with 87.06% inhibition of human colon carcinoma (HCT-116) cell viability, decreased numbers of live cells/numbers of dead cells in a dose-dependent manner with the IC₅₀ values of 34.8–84.7?μM in a 3D culture model depending on durations of incubation. In addition, folipastatin dose-dependently inhibited forskolin-stimulated chloride secretion in human intestinal epithelial (T84) cells with an IC₅₀ value of 0.5?μM. These depsidones were considered to be inactive against noncancerous Vero Cells with the IC₅₀ values of >10?μM.     

EGFR/cell membrane chromatography-online-high performance liquid chromatography/mass spectrometry method for screening EGFR antagonists from Radix Angelicae Pubescentis

The intracellular kinase domains of the epidermal growth factor receptor (EGFR) in some tumor cells are significant targets for drug discovery. We have developed a new EGFR cell membrane chromatography (EGFR/CMC)-online-high performance liquid chromatography/mass spectrometry (HPLC/MS) method for screening anti-EGFR antagonists from medicinal herbs such as Radix Angelicae Pubescentis. In this study, the HEK293 EGFR cells with high expression of EGFR were used to prepare cell membrane stationary phase (CMSP) in the EGFR/CMC model. The retention fractions on the EGFR/CMC model were directly analyzed by combining a 10 port columns switcher with a HPLC/MS system online. As a result, osthole from Radix Angelicae Pubescentis was found to be the active component acting on EGFR like dasatinib as the control drug. There was a good relationship between their inhibiting effects on EGFR secretion and HEK293 EGFR cell growth in vitro. This new EGFR/CMConline-HPLC/MS method can be applied for screening anti-EGFR antagonists from TCMs, for instance, Radix Angelicae Pubescentis. It will be a useful method for drug discovery with natural medicinal herbs as a leading compound resource.     

Chemical speciation of MeHg+ and Hg2+ in aqueous solution and HEK cells nuclei by means of DNA interacting fluorogenic probes

Selected new fluorogenic probes that interact in different ways with Hg²⁺ and MeHg⁺ have been prepared and used for the chemical speciation of both cations in aqueous solution as well as in HEK293 cells. The best selective speciation of Hg²⁺ and MeHg⁺ has been achieved by in vitro approaches based on fluorogenic probes supported in cultured cells, due to the particular sensitivity of the HEK293 cells to permeation by Hg²⁺, MeHg⁺ and the fluorogenic probes. In particular, MeHg⁺ was selectively detected in cell nuclei by probe JG45.     

Insight into the pathway for biosynthesis of sesquarterpenes in nonpathogenic Mycobacterium species: identification of heptaprenylcycli-14E,18E-diene

A new sesquarterpene, heptaprenylcycli-14E,18E-diene, was isolated from Mycobacterium chlorophenolicum cells grown up to the stationary phase. The absence of heptaprenylcycli-14Z,18E-diene indicates that only (E,E,E)-geranylgeranyl diphosphate may be utilized as an intermediate of sesquarterpene biosynthesis in the stationary phase, in contrast with the logarithmic growth phase in which both (E,E)-farnesyl diphosphate and (E,E,E)-geranylgeranyl diphosphate are used. Further, our findings suggest that the stepwise reduction of the polyprenyl group in sesquarterpene biosynthesis might proceed in a different order from that in chlorophyll biosynthesis.     

Synthesis,characterization, and biological evaluation of doxorubicin containing silk fibroin micro- and nanoparticles

The aim of this study was biological evaluation of doxorubicin containing silk fibroin micro- and nanoparticles (Dox-MF and Dox-NF). Dox-MF and Dox-NF were synthesized. Cell toxicity on MCF-7, Saos2, and HFF cells was assessed using MTT assay. Induced apoptosis was assessed using flow cytometry and staining with PI/annexin V. Production of reactive oxygen species (ROS) was measured. Gene expression of p53 was evaluated by real-time PCR. FTIR, SEM, and DLS confirmed the accurate synthesis. Cytotoxicity of Dox-MF and Dox-NF showed significant inhibition of cell growth compared with the controls. Regarding Dox-NF, a significant increase was seen in mRNA level of P53 in MCF-7 and SAOS-2 ​cells and a significant decrease in HFF cells compared to the controls. There was a significantly higher expression of P53 gene in MCF-7 and HFF cells treated by Dox-MF. However, a significant decrease in P53 gene expression was detected in SAOS-2 ​cells. Significant apoptotic induction of cell lines by Dox-MF and Dox-NF was observed in both early and late stages. Dox-MF and Dox-NF acted in the direction of cell death through the apoptotic pathway and changing p53 gene expression. So, Dox-MF and Dox-NF can be considered as a candidate for new anticancer agents.     

Repeated Batch Cell-Immobilized System for the Biotechnological Production of Xylitol as a Renewable Green Sweetener

The present paper studies the biotechnological production of xylitol using sugarcane bagasse hydrolysate in a repeated batch fermentation system with immobilized cells of Candida guilliermondii FTI20037. Immobilized cell system is considered as an attractive alternative to reuse the well-grown and adapted yeast cells in a new fresh fermentation media, without the need of the inoculum stage. In this work, seven repeated batches were performed in a fluidized bed bioreactor using immobilized cells in calcium alginate beads. According to the obtained results it was observed that the immobilized cells of C. guilliermondii can be reused for six successive batches maintaining an average xylitol yield (Y _p/s) of 0.7 g/L and a volumetric productivity (Q _p) of 0.42 g/L?h at the end of 432 h of fermentation. On the other hand, in the seventh batch (504 h), a decrease of 44 % in the final concentration of xylitol was observed. This reduction can be explained by the possible diffusion and accumulation of insoluble substances, found in the hemicellulosic hydrolysate, in the interior of the immobilization support resulting in substrate mass transfer limitations.     

Bioactivated in vivo assembly(BIVA) peptide-tetraphenylethylene(TPE) probe with controllable assembled nanostructure for cell imaging

The emergence of fluorescent light-up molecular probe, which can specifically turn on their fluorescent in the presence of stimulation factors, has open up a new opportunity to advance biosensing and bioimaging. In this work, we designed and synthesized a peptide-AIE conjugate probe for cell imaging with controlled in situ assembled nanostructures. The modular designed probe is consisted of a selfassembled peptide-tetraphenylethene(TPE) motif, a fibroblast activation protein alpha(FAP-α)responsive motif, a hydrophilic motif and a targeting motif. The probe exhibits typically turn-on fluorescence property specifically triggered by FAP-α, which is a significant overexpressed membrane protein on pancreatic tumor cells. Interestingly, the peptide modified the TPE dramatically impacts the assembled nanostructure, which can be modulated by peptide sequences. As a result, the peptide FF(PhePhe) modification of TPE as the self-assembled motif provides a suitable balance of the probe with lightup property and nanofiber assembled structure in situ. Finally, our probe could effectively detect the FAP-α on tumor cells with high specificity. Meantime, the nanofibers in situ assembled on the surface of CAFs enhanced the probe accumulation and prolonged the retention for cell imaging. We envision that this study may inspire new insights into the design of nanostructure controlled AIE light-up bio-probe.     

5-ALA Attenuates the Palmitic Acid-Induced ER Stress and Apoptosis in Bovine Mammary Epithelial Cells

The conservation of mammary gland physiology by maintaining the maximum number of mammary epithelial cells (MECs) is of the utmost importance for the optimum amount of milk production. In a state of negative energy balance, palmitic acid (PA) reduces the number of bovine MECs. However, there is no effective strategy against PA-induced apoptosis of MECs. In the present study, 5-aminolevulinic acid (5-ALA) was established as a remedial agent against PA-induced apoptosis of MAC-T cells (an established line of bovine MECs). In PA-treated cells, the apoptosis-related genes BCL2 and BAX were down- and upregulated, respectively. The elevated expression of major genes of the unfolded protein response (UPR), such as CHOP, a proapoptotic marker (C/EBP homologous protein), reduced the viability of PA-treated MAC-T cells. In contrast, 5-ALA pretreatment increased and decreased BCL2 and BAX expression, respectively. Moreover, cleaved caspase-3 protein expression was significantly reduced in the 5-ALA-pretreated group in comparison with the PA group. The downregulation of major UPR-related genes, including CHOP, extended the viability of MAC-T cells pretreated with 5-ALA and also reduced the enhanced intensity of the PA-induced expression of phospho-protein kinase R-like ER kinase. Moreover, the enhanced expression of HO-1 (antioxidant gene heme oxygenase) by 5-ALA reduced PA-induced oxidative stress (OxS). HO-1 is not only protective against OxS but also effective against ER stress. Collectively, these findings offer new insights into the protective effects of 5-ALA against PA-induced apoptosis of bovine MECs.     

Bioelectrochemical probing of intracellular redox processes in living yeast cells—application of redox polymer wiring in a microfluidic environment

Conventionally, microbial bioelectrochemical assays have been conducted using immobilized cells on an electrode that is placed in an electrochemical batch cell. In this paper, we describe a developed microfluidic platform with integrated microelectrode arrays for automated bioelectrochemical assays utilizing a new double mediator system to map redox metabolism and screen for genetic modifications in Saccharomyces cerevisiae cells. The function of this new double mediator system based on menadione and osmium redox polymer (PVI-Os) is demonstrated. “Wiring” of S. cerevisiae cells using PVI-Os shows a significant improvement of bioelectrochemical monitoring in a microfluidic environment and functions as an effective immobilization matrix for cells that are not strongly adherent. The function of the developed microfluidic platform is demonstrated using two strains of S. cerevisiae, ENY.WA and its deletion mutant EBY44, which lacks the enzyme phosphoglucose isomerase. The cellular responses to introduced glucose and fructose were recorded for the two S. cerevisiae strains, and the obtained results are compared with previously published work when using an electrochemical batch cell, indicating that microfluidic bioelectrochemical assays employing the menadione–PVI-Os double mediator system provides an effective means to conduct automated microbial assays.

Boesenberols I–K,new isopimarane diterpenes from Boesenbergia pandurata with TRAIL-resistance overcoming activity

TRAIL is a promising anticancer agent due to its unique ability to kill tumor cells selectively without harming non-malignant cells. However, some cancer cells are reported to be TRAIL-resistant. Searching potent TRAIL agonist to overcome resistance is an important strategy for its successful use as an anticancer agent. As a part of our continuous research on natural inhibitors, activity-guided fractionation of the MeOH extract of Boesenbergia pandurata rhizomes afforded three (1–3) new isopimarane diterpenes, boesenberols I–K, together with two (4–5) known compounds. The structures of these new compounds were elucidated using HRESIMS, IR, 1D and 2D NMR spectroscopy. All compounds showed TRAIL-resistance overcoming activity.     

Structural characterization of vanchrobactin, a new catechol siderophore produced by the fish pathogen Vibrio anguillarum serotype O2

Vanchrobactin, a new catecol-type siderophore produced by cells of the fish pathogen Vibrio anguillarum serotype O2, has been isolated from the supernatants of iron-deficient cultures. Its structure was characterized from spectral data and established as N-[N′-(2,3-dihydroxybenzoyl)-arginyl]-serine.     

Albatrellus confluens (Alb. & Schwein.) Kotl. & Pouz.: Natural Fungal Compounds and Synthetic Derivatives with In Vitro Anthelmintic Activities and Antiproliferative Effects against Two Human Cancer Cell Lines

Neglected tropical diseases affect the world’s poorest populations with soil-transmitted helminthiasis and schistosomiasis being among the most prevalent ones. Mass drug administration is currently the most important control measure, but the use of the few available drugs is giving rise to increased resistance of the parasites to the drugs. Different approaches are needed to come up with new therapeutic agents against these helminths. Fungi are a source of secondary metabolites, but most fungi remain largely uninvestigated as anthelmintics. In this report, the anthelmintic activity of Albatrellus confluens against Caenorhabditis elegans was investigated using bio-assay guided isolation. Grifolin (1) and neogrifolin (2) were identified as responsible for the anthelmintic activity. Derivatives 4–6 were synthesized to investigate the effect of varying the prenyl chain length on anthelmintic activity. The isolated compounds 1 and 2 and synthetic derivatives 4–6, as well as their educts 7–10, were tested against Schistosoma mansoni (adult and newly transformed schistosomula), Strongyloides ratti, Heligmosomoides polygyrus, Necator americanus, and Ancylostoma ceylanicum. Prenyl-2-orcinol (4) and geranylgeranyl-2-orcinol (6) showed promising activity against newly transformed schistosomula. The compounds 1, 2, 4, 5, and 6 were also screened for antiproliferative or cytotoxic activity against two human cancer lines, viz. prostate adenocarcinoma cells (PC-3) and colorectal adenocarcinoma cells (HT-29). Compound 6 was determined to be the most effective against both cell lines with IC₅₀ values of 16.1 µM in PC-3 prostate cells and 33.7 µM in HT-29 colorectal cells.