Evaluation of Boric Acid Treatment on microRNA-127-5p and Metastasis Genes Orchestration of Breast Cancer Stem Cells.

Coregulation of microRNAs (miRNAs) and cancer stem cells (CSCs) is very important in carcinogenesis. miR-127-5p is known to be downregulated in breast cancer. In this study, we aimed to investigate how boric acid (BA), known for its previously unstudied anti-cancer properties, would affect the expression of miR127-5p and genes responsible for breast cancer stem cells (BC-SCs) metastasis. BC-SCs were isolated from human breast cancer cells (MCF-7) by immunomagnetic cell separation and characterized with flow cytometry and sphere formation. The viability of BC-SCs and the determination of its IC50 value in response to boric acid (BA) were assessed via the MTT assay. Boric acid exhibited dose- and time-dependent inhibition of cell viability in cells. The IC50 doses of boric acid in MCF-7 cells and BC-SCs were 45.69 mM and 41.27 mM, respectively. The impact of BA on the expression of metastatic genes and miR127-5p was elucidated through RT-qPCR analysis. While the expression of the COL1A1 (p < 0.05) and VIM (p < 0.01) was downregulated, the expression of the miR-127-5p, ZEB1 (p < 0.01), CDH1 (p < 0.05), ITGB1 (p < 0.05), ITGA5 (p < 0.05), LAMA5 (p < 0.01), and SNAIL (p < 0.05), was up-regulated in dose-treated BC-SCs (p < 0.001) to the RT-qPCR results. Our findings suggest that boric acid could induce miR-127-5p expression. However, it cannot be said that it improves the metastasis properties of breast cancer stem cells.

High-throughput microfluidic chip with silica gel-C18 channels for cyclotide separation.

Over the past two decades, microfluidic-based separations have been used for the purification, isolation, and separation of biomolecules to overcome difficulties encountered by conventional chromatography-based methods including high cost, long processing times, sample volumes, and low separation efficiency. Cyclotides, or cyclic peptides used by some plant families as defense agents, have attracted the interest of scientists because of their biological activities varying from antimicrobial to anticancer properties. The separation process has a critical impact in terms of obtaining pure cyclotides for drug development strategies. Here, for the first time, a mimic of the high-performance liquid chromatography (HPLC) on microfluidic chip strategy was used to separate the cyclotides. In this regard, silica gel-C18 was synthesized and characterized by Fourier-transform infrared spectroscopy (FTIR) and proton nuclear magnetic resonance (1H-NMR) and then filled inside the microchannel to prepare an HPLC C18 column-like structure inside the microchannel. Cyclotide extract was obtained from Viola ignobilis by a low voltage electric field extraction method and characterized by HPLC and matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF). The extract that contained vigno 1, 2, 3, 4, 5, and varv A cyclotides was added to the microchannel where distilled water was used as a mobile phase with 1 µL/min flow rate and then samples were collected in 2-min intervals until 10 min. Results show that cyclotides can be successfully separated from each other and collected from the microchannel at different periods of time. These findings demonstrate that the use of microfluidic channels has a high impact on the separation of cyclotides as a rapid, cost-effective, and simple method and the device can find widespread applications in drug discovery research.

On chip microfluidic separation of cyclotides.

Cyclotides as a cyclic peptide produced by different groups of plants have been a very attractive field of research due to their exceptional properties in biological activities and drug design applications. The importance of cyclotides as new biological activities from nature caused to attract researchers to develop new separation systems. Recent growth and development on chip-based technology for separation and bioassay especially for anticancer having sparklingly advantages comparison with common traditional methods. In this study, the microfluidic separation of Vigno 1-5 cyclotides extracted from Viola ignobilis by using polar and nonpolar forces as a liquid-liquid interaction was investigated through modified microfluidic chips and then the results were compared with a traditional counterpart technique of high-performance liquid chromatography (HPLC). The traditional process of separating cyclotides from plants is a costly and time-consuming procedure. The scientific novelty of this study is to accelerate the separation of cyclotides using modified microfluidic chips with low cost and high efficiency. The results revealed that a novel and simple microfluidic chip concept is an effective approach for separating the Vigno groups in the violet extract. We believe that the concept could potentially be utilized for further drug development process especially for anticancer studies by coupling bioassay chips as online procedures via reducing in time and cost compared with traditional offline methods.

Revealing the single-channel characteristics of OprD (OccAB1) porins from hospital strains of Acinetobacter baumannii.

Nowadays, reports of antimicrobial resistance (AMR) against many antibiotics are increasing because of their misapplication. With this rise, there is a serious decrease in the discovery and development of new types of antibiotics amid an increase in multi-drug resistance. Unfermented Acinetobacter baumannii from gram-negative bacteria, which is one of the main causes of nosocomial infections and multi-drug resistance, has 4 main kinds of antibiotic resistance mechanism: inactivating antibiotics by enzymes, reduced numbers of porins and changing of their target or cellular functions due to mutations, and efflux pumps. In this study, characterization of the possible mutations in OprD (OccAB1) porins from hospital strains of A. baumannii were investigated using single channel electrophysiology and compared with the standard OprD isolated from wild type ATCC 19,606. For this aim, 5 A. baumannii bacteria samples were obtained from patients infected with A. baumannii, after which OprD porins were isolated from these A. baumannii strains. OprD porins were then inserted in an artificial lipid bilayer and the current-voltage curves were obtained using electrical recordings through a pair of Ag/AgCl electrodes. We observed that each porin has a characteristic conductance and single channel recording, which then leads to differences in channel diameter. Finally, the single channel data have been compared with the gene sequences of each porin. It was interesting to find out that each porin isolated has a unique porin diameter and decreased anion selectivity compared to the wild type.

Simple and low-cost antibiotic susceptibility testing for Mycobacterium tuberculosis using screen-printed electrodes.

One quarter of the global population is thought to be latently infected by Mycobacterium tuberculosis (TB) with it estimated that 1 in 10 of those people will go on to develop active disease. Due to the fact that M. tuberculosis (TB) is a disease most often associated with low- and middle-income countries, it is critical that low-cost and easy-to-use technological solutions are developed, which can have a direct impact on diagnosis and prescribing practice for TB. One area where intervention could be particularly useful is antibiotic susceptibility testing (AST). This work presents a low-cost, simple-to-use AST sensor that can detect drug susceptibility on the basis of changing RNA abundance for the typically slow-growing M. tuberculosis (TB) pathogen in 96 h using screen-printed electrodes and standard molecular biology laboratory reactionware. In order to find out the sensitivity of applied sensor platform, a different concentration (108 -103  CFU/mL) of M. tuberculosis was performed, and limit of detection and limit of quantitation were calculated as 103.82 and 1011.59  CFU/mL, respectively. The results display that it was possible to detect TB sequences and distinguish antibiotic-treated cells from untreated cells with a label-free molecular detection. These findings pave the way for the development of a comprehensive, low-cost, and simple-to-use AST system for prescribing in TB and multidrug-resistant tuberculosis.

Rapid fabrication of teflon apertures by controlled high voltage pulses for formation of free standing planar lipid bilayer membrane.

Free standing artificial lipid bilayers are widely used in the study of biological pores. In these types of studies, the free standing planar lipid bilayer is formed over a micron-sized aperture consisting of either polymer such as Polytetrafluoroethylene (PTFE, Teflon) or glass. Teflon is chemically inert, has a low dielectric constant, and has a high electrical resistance which combined allow for obtaining low noise recordings. This study investigates the reproducible generation of micropores in the range of 50-100 microns in diameter in a Teflon film using a high energy discharge set-up. The discharger set-up consists of a microprocessor, a transformer, a voltage regulator, and is controlled by a computer. We compared two approaches for pore creation: single and multi-pulse methods. The results showed that the multi-pulse method produced narrower aperture size distributions and is more convenient for lipid bilayer formation, and thus would have a higher success rate than the single-pulse method. The bilayer stability experiments showed that the lipid bilayer lasts for more than 33 h. Finally, as a proof-of-concept, we show that the single and multi-channel electrophysiology experiments were successfully performed with the apertures created by using the mentioned discharger. In conclusion, the described discharger provides reproducible Teflon-pores in a cheap and easy-to-operate manner.

Electrospun essential oil-polycaprolactone nanofibers as antibiofilm surfaces against clinical Candida tropicalis isolates.

OBJECTIVE: As an approach to prevent biofilm infections caused by Candida tropicalis on various surfaces, determination of effect of biodegradable polycaprolactone nanofibers (PCLNFs) with different concentrations of two different essential oils were tested in this study. RESULTS: Both of the tested essential oils exhibited antifungal effect (minimal inhibitory concentration; 0.25-0.49 µL/mL, minimal fungicidal concentration; 0.25-0.49 µL/mL, depending on the C. tropicalis strain) (Zone of inhibition caused by 500 µL/mL concentration of oils; 28-56 mm). 0, 2, 4% clove oil PCLNFs and 0, 2, 4% red thyme oil-PCLNFs were free from bead formation and uniform in diameter. Diameters of all essential oil containing PCLNFs were ranged from 760 to 1100 nm and were significantly different from 0% essential oil-PCLNF (P < 0.05). 0, 2, 4% clove oil-PCLNFs were significantly more hydrophobic compared to 8% clove oil-PCLNF (P < 0.01), whereas 0% and 2% red thyme oil-PCLNFs were significantly more hydrophobic compared to 4% and 8% red thyme oil PCLNFs (P < 0.01). Highest amount of biofilm inhibition was observed by 4% clove oil-PCLNF and by 4% red thyme oil-PCLNF. CONCLUSIONS: Clove and red thyme oils may be used not only as antifungals but also as biofilm inhibitive agents on surfaces of biomaterials that are frequently contaminated by C. tropicalis, when they are incorporated into PCLNFs.