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Because ADAM17 promotes colonic tumorigenesis, we investigated potential miRNAs regulating ADAM17; and examined effects of diet and tumorigenesis on these miRNAs. We also examined pre-miRNA processing and tumour suppressor roles of several of these miRNAs in experimental colon cancer. Using TargetScan, miR-145, miR-148a, and miR-152 were predicted to regulate ADAM17. miR-143 was also investigated as miR-143 and miR-145 are co-transcribed and associated with decreased tumour growth. HCT116 colon cancer cells (CCC) were co-transfected with predicted ADAM17-regulating miRNAs and luciferase reporters controlled by ADAM17-3’UTR. Separately, pre-miR-143 processing by colonic cells was measured. miRNAs were quantified by RT-PCR. Tumours were induced with AOM/DSS in WT and transgenic mice (Tg) expressing pre-miR-143/miR-145 under villin promoter. HCT116 transfection with miR-145, −148a or −152, but not scrambled miRNA inhibited ADAM17 expression and luciferase activity. The latter was suppressed by mutations in ADAM17-3’UTR. Lysates from colonocytes, but not CCC, processed pre-miR-143 and mixing experiments suggested CCC lacked a competency factor. Colonic miR-143, miR-145, miR-148a, and miR-152 were downregulated in tumours and more moderately by feeding mice a Western diet. Tg mice were resistant to DSS colitis and had significantly lower cancer incidence and tumour multiplicity. Tg expression blocked up-regulation of putative targets of miR-143 and miR-145, including ADAM17, K-Ras, XPO5, and SET. miR-145, miR-148a, and miR-152 directly suppress colonocyte ADAM17 and are down-regulated in colon cancer. This is the first direct demonstration of tumour suppressor roles for miR-143 and miR-145 in an in vivo model of colonic tumorigenesis.

DNA 5-hydroxymethylcytosine (5hmC) modification is known to be associated with gene transcription and frequently used as a mark to investigate dynamic DNA methylation conversion during mammalian development and in human diseases. However, the lack of genome-wide 5hmC profiles in different human tissue types impedes drawing generalized conclusions about how 5hmC is implicated in transcription activity and tissue specificity. To meet this need, we describe the development of a 5hmC tissue map by characterizing the genomic distributions of 5hmC in 19 human tissues derived from ten organ systems. Subsequent sequencing results enabled the identification of genome-wide 5hmC distributions that uniquely separates samples by tissue type. Further comparison of the 5hmC profiles with transcriptomes and histone modifications revealed that 5hmC is preferentially enriched on tissue-specific gene bodies and enhancers. Taken together, the results provide an extensive 5hmC map across diverse human tissue types that suggests a potential role of 5hmC in tissue-specific development; as well as a resource to facilitate future studies of DNA demethylation in pathogenesis and the development of 5hmC as biomarkers., DNA 5-hydroxymethylcytosine (5hmC) modification is associated with gene transcription and used as a mark of mammalian development. Here the authors report a comprehensive 5hmC tissue map and analysis of 5hmC genomic distributions in 19 human tissues derived from 10 organ systems, thus providing insights into the role of 5hmC in tissue-specific development.

Carbon fiber reinforced polymer (CFRP) plates are widely used to retrofit or reinforce steel structures, and the debonding damage between the steel structure and the CFRP plate is a typical failure in strengthening steel structures. This paper proposes a new approach to detecting debonding between a steel beam and a reinforcing CFRP plate by using removable lead zirconate titanate (PZT)-based transducers and active sensing. The removable PZT-based transducers are used to implement the active sensing approach, in which one transducer, as an actuator, is used to generate stress wave, and another transducer, as a sensor, is used to detect the stress wave that propagates across the bonding between the steel beam and the reinforcing CFRP plate. The bonding condition significantly influences the received sensor signal, and a wavelet-packet-based energy index (WPEI) is used to quantify the energy of the received signal to evaluate the severity of debonding between the steel beam and the reinforcing CFRP plate. To validate the proposed approach, experimental studies were performed, and two removable PZT-based transducers were designed and fabricated to detect the debonding between a steel beam and the reinforcing CRFP plate. The experimental results demonstrate the feasibility of the proposed method in detecting the debonding between a steel beam and the reinforcing CFRP plate using removable PZT-based transducers.

In this work, we develop a ratiometric two-photon fluorescent probe, ATD@QD-E2Zn2SOD (ATD = amino triphenylamine dendron, QD = CdSe/ZnSe quantum dot, E2Zn2SOD = Cu-free derivative of bovine liver copper-zinc superoxide dismutase), for imaging and sensing the changes of intracellular Cu(2+) level with clear red-to-yellow color change based on specific biomolecular recognition of E2Zn2SOD for Cu(2+) ion. The inorganic-organic nanohybrided fluorescent probe features two independent emission peaks located at 515 nm for ATD and 650 nm for QDs, respectively, under two-photon excitation at 800 nm. Upon addition of Cu(2+) ions, the red fluorescence of QDs drastically quenches, while the green emission from ATD stays constant and serves as a reference signal, thus resulting in the ratiometric detection of Cu(2+) with high accuracy by two-photon microscopy (TPM). The present probe shows high sensivity, broad linear range (10(-7)-10(-3) M), low detection limit down to ∼10 nM, and excellent selectivity over other metal ions, amino acids, and other biological species. Meanwhile, a QD-based inorganic-organic probe demonstrates long-term photostability, good cell-permeability, and low cytotoxicity. As a result, the present probe can visualize Cu(2+) changes in live cells by TPM. To the best of our knowledge, this is the first report for the development of a QD-based two-photon ratiometric fluorescence probe suitable for detection of Cu(2+) in live cells.

Horseradish peroxidase (HRP)-mediated crosslinking of poly(amido amine) (PAA) copolymers was successfully applied in the preparation of in situ forming degradable hydrogels under physiological conditions. PAA copolymers containing different amounts of tyramine residues (termed as pAEEOL/TA) could be synthesized through Michael-type addition between methylenebisacryamide and amine mixture of 2-(2-aminoethoxy) ethanol and tyramine (TA). Depending on the amount of TA residue, the HRP, and H2O2 concentration, the gelation times could be varied from about 50 to 350 s. The swelling and degradation experiments indicated under physiological conditions the pAEEOL/TA-based hydrogels are completely degradable within 6–8 days. Rheological analysis revealed that storage modulus of the hydrogels increased from 2500 to 4100 Pa when increasing HRP concentrations. Importantly, pAEEOL/TA copolymers have low cytotoxicity. Moreover, NIH 3T3 (mouse embryonic fibroblast) cells exposed in the degradation products of pAEEOL/TA-based hydrogels retained high cell viability, implying that the hydrogels are cyto-biocompatible. In vitro release of methylene blue and IgG protein from pAEEOL/TA-based hydrogels could be effectively sustained by encapsulation of the drug in the hydrogels. The results indicate that HRP-crosslinked, degradable pAEEOL/TA-based hydrogels are promising for biomedical applications. © 2012 Wiley Periodicals, Inc. J. Appl. Polym. Sci., 2013

This paper demonstrates a novel strategy for site-selective cell adhesion and in situ cultivation of living cells, integrated with real-time monitoring of cellular small biomolecules based on dual functional protein microarrays. The protein microarrays have been produced on the superhydrophobic|philic Au-TiO2 micropatterns, through further modification of L-cysteine (Cys) and followed by successive immobilization of a model protein, cytochrome c (cyt c). Experimental results have revealed that the created cyt c microarrays play dual functions: one is employed as a robust substrate for site-selective cell adhesion and in situ cultivation of living cells, because the protein microarrays exhibit high selectivity and bioaffinity toward cells, as well as long biostability under cell culture condition up to 7 days. Meanwhile, the cyt c microarrays can also serve as sensing elements for hydrogen peroxide (H2O2) due to the inherent enzymatic activity of the heme center in cyt c. Direct electron transfer of cyt c has been enhanced at the Cys-modified Au-TiO2 (Au-TiO2/Cys) microarrays, and the electrochemical behavior can be tuned by varying the width and spacing of the microband arrays. Furthermore, cyt c is stably immobilized on the Au-TiO2/Cys microarrays and maintains its enzymatic activity after confined on the microarrays. Thus, the optimized cyt c microarrays show striking analytical performance for H2O2 determination, e.g., high sensitivity and selectivity, broad linear range from 10(-9) M to 10(-2) M, low detection limit down to 2 nM, and short response time within 5 s. As a result, the excellent analytical properties of the cyt c microarrays, as well as the characteristic of the protein microarrays themselves, including high selectivity, long biostability, and good bioaffinity, opens up a method for selective in situ cultivation of cells integrated with real-time detection of signaling biomolecules such as H2O2 released from living cells, which shows potential for physiological and pathological investigations.

Here, we report on a novel superoxide anion (O2(*-)) biosensor based on direct electron transfer of copper, zinc-superoxide dismutase (Cu, Zn-SOD) at zinc oxide nanodisks surface for in vivo tracking of O2(*-) in bean sprouts. Direct electron transfer of SOD is achieved at ZnO nanodisks film prepared by a one-step electrodeposited method, with a high heterogeneous electron rate constant of 17 +/- 2 s(-1). Spectroscopic data demonstrate that SOD strongly immobilized onto the nanostructured ZnO surfaces processes its inherent activity toward O2(*-) dismutation. A combination of the facilitated direct electron transfer and the bifunctional enzymatic catalytic activities of the SOD substantially provides a dual electrochemical approach to determination of O2(*-) with high selectivity, wide linear range, long stability, and good reproducibility. In particular, SOD adsorbed on the ZnO nanodisks film is capable of sensing O2(*-) cathodically at a very positive potential, 0 mV (vs Ag|AgCl), where the common interfering species such as hydrogen peroxide, uric acid, ascorbic acid, and 3,4-dihydroxyphenylacetic acid were effectively avoided. The excellent analytical performance of the present O2(*-) biosensor, combined with the remarkable characteristics of nanostructured ZnO films, such as biocompatibility, ease of preparation, and facile to miniaturize, paves an electrochemical way for reliable and durable in vivo determination of O2(*-) in bean sprouts.

Based on the differences in titration spectra, mixtures of organic acids with similar p K a were determined simultaneously. Principal component regression calibration was used for the evaluation of titration spectra. The effects of differences in p K a and sampling interval of pH on accuracy were studied in detail. Combining precipitation and the acid–base reaction of organic acid, precipitation–acid–base titrimetry is proposed. The mixture of acids with almost the same p K a can be determined simultaneously and accurately only if their p K sp values are different. Synthetic mixtures of isomer of chlorobenzoic acid and homologue of phenol were determined simultaneously and acceptable results were obtained.

A novel and facile photoelectrochemical method has been developed to detect H(2)S in water samples with high sensitivity and selectivity. The protocol is based on the photocurrent generated by CdS nanoclusters which are deposited onto TiO(2) nanotubes exposing in CdSO(4) solution with the gradual addition of Na(2)S, with low-cost, environment-friendly, theoretical and technical simplicity. The developed method shows a very broad linear range from 10(-8) M to 10(-3) M and a low detection limit of 0.31 nM (9.92 ppt), far lower than the ceiling value in drinking water provided by WHO. Furthermore, the present method has been applied for determination of H(2)S in water samples. The concentrations of H(2)S in water samples determined by the present method are in a good agreement with those monitored by traditional spectrophotometry.

A newly developed hydrogen peroxide selective membrane electrode using lettuce seed meal as source biocatalyst is described. This amperometric-based probe exhibits high biocatalytic activity, long lifetime and favorable selectivity for the measurement of hydrogen peroxide.