MicroRNA-331-3p Suppresses Cervical Cancer Cell Proliferation and E6/E7 Expression by Targeting NRP2

Aberrant expression of microRNAs (miRNAs) is involved in the development and progression of various types of cancers. In this study, we investigated the role of miR-331-3p in cell proliferation and the expression of keratinocyte differentiation markers of uterine cervical cancer cells. Moreover, we evaluated whether neuropilin 2 (NRP2) are putative target molecules that regulate the human papillomavirus (HPV) related oncoproteins E6 and E7. Cell proliferation in the human cervical cancer cell lines SKG-II, HCS-2, and HeLa was assessed using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay. Cellular apoptosis was measured using the TdT-mediated dUTP nick end labeling (TUNEL) and Annexin V assays. Quantitative RT-PCR was used to measure the messenger RNA (mRNA) expression of the NRP2, E6, E7, p63, and involucrin (IVL) genes. A functional assay for cell growth was performed using cell cycle analyses. Overexpression of miR-331-3p inhibited cell proliferation, and induced G2/M phase arrest and apoptosis in SKG-II, HCS-2 and HeLa cells. The luciferase reporter assay of the NRP2 3′-untranslated region revealed the direct regulation of NRP2 by miR-331-3p. Gene expression analyses using quantitative RT-PCR in SKG-II, HCS-2, and HeLa cells overexpressing miR-331-3p or suppressing NRP2 revealed down-regulation of E6, E7, and p63 mRNA and up-regulation of IVL mRNA. Moreover, miR-331-3p overexpression was suppressed NRP2 expression in protein level. We showed that miR-331-3p and NRP2 were key effectors of cell proliferation by regulating the cell cycle, apoptosis. NRP-2 also regulates the expression of E6/E7 and keratinocyte differentiation markers. Our findings suggest that miR-331-3p has an important role in regulating cervical cancer cell proliferation, and that miR-331-3p may contribute to keratinocyte differentiation through NRP2 suppression. miR-331-3p and NRP2 may contribute to anti-cancer effects.     

Nanostructured poly(3-hexylthiophene-2,5-diyl) films with tunable dimensions through self-assembly with polystyrene

Poly(3-hexylthiophene-2,5-diyl) is among the most widely used conjugated polymers for opto-electronic applications. To enhance its properties, researchers have attempted to nanostructure this polymer using various processes including breath figure arrays, nanolithography and elaborated organic synthesis. We here demonstrate a simple process to nanostructure the conjugated polymer using self-assembly with polystyrene and selective removal of one of the phases. The influence of the molecular weight of each polymer on the thin film morphology was systematically studied by atomic force microscopy. Using this approach, we observe two types of nanostructure, namely, nanoporous and nanoisland structures, of which the dimensions can be tuned by modifying the molecular weight of each polymer in the blend. This simple process introduces a cost-effective alternative to produce thin films of conjugated polymer with average nano-features from 100 nm up to 500 nm which could be used in a wide range of applications.     

Multilayered reasoning by means of conceptual fuzzy sets

The real world consists of instances of events and continuous numeric values, while people represent and process their knowledge in terms of symbols. Fuzzy sets provide a strong notation connecting the symbolic representation to the real world. In previously proposed Conceptual Fuzzy Sets (CFS), the meaning of a concept is represented by the distribution of activations of labels in a bidirectional associative memory. In particular, a multilayered structured CFS represents the meaning of the same concept as it is used in various expressions in each layer. The propagation of activations corresponds to reasoning. Therefore, we propose a multilayered reasoning method associated to a multilayered structured CFS, which has the following features: (1) capable of simultaneous symbolic and quantitative processing, (2) capable of simultaneous top-down and bottom-up processing. The effectiveness of the proposed method is illustrated by practical examples of decision regarding the amount of steering in the task of parking a car, and recognition of facial expressions for an image understanding system. © 1996 John Wiley & Sons, Inc.     

Synthesis and chemical properties of polyethyleneimines crosslinked by penta-2,4-dienylideneammonium unit

The reaction of N-(2,4-dinitrophenyl)pyridinium anion ( salt(A) ; A = Cl⁻, FeCl₄⁻, and (CN)₂N⁻) with linear polyethyleneimine (LPEI; M_n = 20 380) and branched polyethyleneimines (BPEI1; M_n = 600, BPEI2; M_n = 10 000) at various molar feed ratios without using a catalyst resulted in pyridinium ring opening to yield ionic LPEI and BPEIs that were crosslinked by conjugated penta-2,4-dienylideneammonium (PDA) units, LPEI-PDA , BPEI1-PDA , and BPEI2-PDA , respectively. A model compound was synthesized by the reaction of salt(Cl) with diethylamine. The solubilities of BPEI1-PDA and BPEI2-PDA depended on the feed ratios between salt(Cl) and BPEI1 or BPEI2. Dipping LPEI-PDA into water and methanol yielded hydro- and organogels, respectively. UV–vis and reflection measurements revealed an expanded π-conjugation length between the polymer chains due to the through-space orbital interaction of the electrons on the two nitrogen atoms at the crosslinked positions in LPEI-PDA , BPEI1-PDA , and BPEI2-PDA . Cyclic voltammetry analysis suggested that the polymers underwent electrochemical oxidation. Measurement using a superconducting quantum interference device (SQUID) indicated that LPEI-PDA having FeCl₄⁻ anions was paramagnetic. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2020 , 137, 48712.     

Effect of morphology on shear viscosity for binary blends of polycarbonate and polystyrene

The structure and rheological properties of binary blends of polycarbonate (PC) and polystyrene (PS) were investigated using various PS samples with different molecular weights, namely PS1k (M_w = 1,000), PS53k (M_w = 53,000), and PS240k (M_w = 240,000). The blends with PS53k and PS240k show phase-separated structures, whereas the blend with PS1k is miscible. The shear viscosity decreases greatly on addition of PS53k and PS240k, especially at high shear rates, which would be a great advantage at processing operations. Because the nonlinear response occurs in the small strain region for multilayered films of PC and PS240k, the origin of the significant viscosity drop for the phase-separated system is interfacial slippage at the phase boundary.     

Determination of histamine in fish and fish products by tandem solid-phase extraction

A simple and practical method was developed for the determination of histamine in fish and fish products by solid-phase extraction and fluorescence derivatization. Histamine was extracted with trichloroacetic acid. The extract was neutralized and diluted with phosphate buffer (pH 6.8), and cleaned up with a tandem-connected octadecyl silica (ODS) and strong cation exchange silica (SCX) cartridge. After removal of the solvent, histamine was derivatized with fluorescamine and analyzed by ion-paired reversed-phase high-performance liquid chromatography with fluorescence detection. Recovery tests of histamine from six kinds of fish and fish products showed acceptable recovery (83-92%) with low relative standard deviation (less than 5%). This method could be useful for determination of histamine in fish.     

Selective enumeration of viable Enterobacteriaceae and Pseudomonas spp. in milk within 7 h by multicolor fluorescence in situ hybridization following microcolony formation

Rapid and simultaneous enumeration of viable Enterobacteriaceae and viable Pseudomonas spp. in milk was achieved by using multicolor fluorescence in situ hybridization (FISH) with oligonucleotide probes based on 16S ribosomal RNA (rRNA) sequences in combination with a microcolony growth method (multicolor microcolony-FISH; MMC-FISH). The procedure of MMC-FISH method is rather simple; that is milk clearing, filtration of cells, incubation, hybridization and enumeration. Enumeration of targeted bacteria in logarithmic growth phase, stationary phase, or in a starved state in milk by MMC-FISH required 5-7 h, while it took 1-3 days to test for Escherichia coli and Pseudomonas putida by the conventional culture method. The numbers of E. coli and P. putida in each phase or in a starved state in milk determined by MMC-FISH were almost the same or greater than the number of colony forming units determined by the plate counting method. The MMC-FISH allows rapid examination of contamination in milk by viable Enterobacteriaceae and Pseudomonas spp. with growth potential.     

Degradation of cyanuric acid in soil by Pseudomonas sp. NRRL B-12227 using bioremediation with self-immobilization system

The rates of degradation of cyanuric acid, a key intermediate in a metabolic pathway of s-triazine herbicides, were measured for Pseudomonas sp. NRRL B-12227. The rate of degradation was affected by the rate of cyanuric acid transport through cell membranes and the activity of cyanuric acid amidohydrolase inside the cells. At low concentrations of cyanuric acid, the acclimation of cells to cyanuric acid and/or added nutrients effectively enhanced the degradation rate. The strain was also applied to bioremediation using a Bioremediation with Self-Immobilization System (BSIS), in which Pseudomonas sp. NRRL B-12227 cells were co-immobilized with Bacillus subtilis, the latter of which secretes a viscous polymer, in a shallow layer of soil packed in a column. More than 70% of the Pseudomonas sp. NRRL B-12227 cells were co-immobilized with the B. subtilis in a 7.5 cm layer of the packed soil by self-aggregation. More than 60% of the 1 mM cyanuric acid supplied to the packed soil was degraded in this layer during a 72 h period.     

Accumulation and elimination profiles of paralytic shellfish poison in the short-necked clam Tapes japonica fed with the toxic dinoflagellate Gymnodinium catenatum

The paralytic shellfish poison (PSP)-producing dinoflagellate Gymnodinium catenatum (Gc) was fed to the short-necked clam Tapes japonica, and the accumulation, transformation and elimination profiles of PSP were investigated by means of high-performance liquid chromatography with postcolumn fluorescence derivatization (HPLC-FLD). The short-necked clams ingested most of the Gc cells (4 x 10(6) cells) supplied as a bolus at the beginning of the experiment, and accumulated a maximal amount of toxin (181 nmol/10 clams) after 12 hr. The rate of toxin accumulation at that time was 16%, which rapidly decreased thereafter. During the rearing period, a variation in toxin composition, derived presumably from the transformation of toxin analogues in the clams, was observed, including a reversal of the ratio of C2 to C1, and the appearance of carbamate (gonyautoxin (GTX) 2, 3) and decarbamoyl (dc) derivatives (decarbamoylsaxitoxin (dcSTX) and dcGTX2, 3), which were undetectable in Gc cells. The total amount of toxin contained in clams and residue (remaining Gc cells and/or excrement in the rearing tank) gradually declined, and only about 1% of the supplied toxin was detected at the end of the experiment.     

Theasaponin E1 destroys the salt tolerance of yeasts

Cells of Zygosaccharomyces rouxii in a medium containing a high concentration of NaCl were killed during incubation for 2-4 h with a low concentration of a mixture of saponins from tea seeds (TSS). The higher the concentration of NaCl in the medium, the higher the inhibitory effect of TSS on the growth of the yeast. The above inhibitory effect of TSS on the growth of the yeast was not observed when cells were incubated in hypertonic media composed of nonionic substances such as sugars. The ATPase activity of plasma membrane preparations from the yeast cells was slightly affected by the addition of TSS. It is shown that TSS facilitates leakage of glycerol from the yeast cells under NaCl-hypertonic conditions. The major inhibitor in the mixture of saponins was isolated and identified as theasaponin E1. Its isomer, theasaponin E2, did not have any effect on the salt tolerance of Z. rouxii or Saccharomyces cerevisiae.