Synthesis, DNA binding of bis-naphthyl ferrocene derivatives and the application as new electroactive indicators for DNA biosensor.

A series of bis-naphthyl ferrocene derivatives were synthesized and characterized. Based on the results obtained from UV-visible absorption titration and ethidium bromide (EB) displacement experiments, it was observed that the synthesized compounds exhibited a strong binding ability to dsDNA. In comparison to the viscosity curve of EB, the tested compounds demonstrated a bisintercalation binding mode when interacting with CT-DNA. Differential pulse voltammetry (DPV) was employed to assess the binding specificity of these indicators towards ssDNA and dsDNA. All tested indicators displayed more pronounced signal differences before and after hybridization between probe nucleic acids and target nucleic acids compared to Methylene Blue (MB). Among the evaluated compounds, compound 3j containing an ether chain showed superior performance as an indicator, making it suitable for constructing DNA-based biosensors. Under optimized conditions including probe ssDNA concentration and indicator concentration, this biosensor exhibited good sensitivity, reproducibility, stability, and selectivity. The limit of detection was calculated as 4.53 × 10-11 mol/L. Furthermore, when utilizing 3j as the indicator in serum samples, the biosensor achieved satisfactory recovery rates for detecting the BRCA1 gene.

Large scale uniform Ni-P plated carbon fiber for boosting urea electro-oxidation and electro-detection.

Seeking an excellent electrocatalyst is the trickiest issue for the application of urea electro-oxidation and electro-detection. Phosphorus-doped nickel plating on carbon fibers (Ni-P/CF) is synthesized by simple electroless plating. SEM results exhibit that the Ni-P densely and uniformly grows onto the surface of carbon fibers (CF), forming carbon fibers-like nanoarchitectures. Benefiting from the carbon fibers-like nano architectures with abundant exposed active sites on the surface of CF, electron transfer can be synchronously facilitated, and Ni-P/CF displays superior urea electrooxidation (UOR) performance with potentials of 1.40 V to reach 100 mA cm-2. Impressively, it can maintain at 20 mA cm-2 for 48 h without evident activity attenuation, demonstrating robust durability. Cycle stability shows that the voltage has only increased by 10 mV at 300 mA cm-2 from the 10th to 20000th cycles. Most importantly, Ni-P/CF at a length of 100 cm with good reproducibility was successfully synthesized, denoting great potential for large-scale industrial production. Therefore, this work not only affords cost-effective tactics for urea-rich wastewater degradation but also can achieve practical medical applications.

Genomic vulnerability to climate change in Quercus acutissima, a dominant tree species in East Asian deciduous forests.

Understanding the evolutionary processes that shape the landscape of genetic variation and influence the response of species to future climate change is critical for biodiversity conservation. Here, we sampled 27 populations across the distribution range of a dominant forest tree, Quercus acutissima, in East Asia, and applied genome-wide analyses to track the evolutionary history and predict the fate of populations under future climate. We found two genetic groups (East and West) in Q. acutissima that diverged during Pliocene. We also found a heterogeneous landscape of genomic variation in this species, which may have been shaped by population demography and linked selections. Using genotype-environment association analyses, we identified climate-associated SNPs in a diverse set of genes and functional categories, indicating a model of polygenic adaptation in Q. acutissima. We further estimated three genetic offset metrics to quantify genomic vulnerability of this species to climate change due to the complex interplay between local adaptation and migration. We found that marginal populations are under higher risk of local extinction because of future climate change, and may not be able to track suitable habitats to maintain the gene-environment relationships observed under the current climate. We also detected higher reverse genetic offsets in northern China, indicating that genetic variation currently present in the whole range of Q. acutissima may not adapt to future climate conditions in this area. Overall, this study illustrates how evolutionary processes have shaped the landscape of genomic variation, and provides a comprehensive genome-wide view of climate maladaptation in Q. acutissima.

Synthesis, electrochemistry, DNA binding and in vitro cytotoxic activity of tripodal ferrocenyl bis-naphthalimide derivatives.

A series of tripodal ferrocenyl bis-naphthalimide derivatives were synthesized and characterized. All of the bis-naphthalimide derivatives exhibited good DNA binding ability which was confirmed by ethidium bromide (EB) displacement experiment and ultraviolet (UV)-visible absorption titration. And the binding mode of these compounds was proved to be a hybrid binding mode by experiments. The cytotoxicity of synthesized compounds against 4 different human cancer cell lines (EC109, BGC823, SGC7901 and HEPG2) was evaluated by thiazolyl blue tetrazolium bromide (MTT) assay. All of the bis-naphthalimide derivatives exhibited good anticancer activity than the positive control drug (amonafide), which was due to the promotion of reactive oxygen species (ROS) level in test cancer cells by the reversible one-electron redox process of ferrocenyl bis-naphthalimide derivatives. Although there was no obvious relationship between the binding constants and the chain length, the structure cytotoxicity relationship revealed that the linker of n = 3, m = 1 was the best choice for the tested tripodol bis-naphthalimide derivatives. SYNOPSIS: A series of tripodal ferrocenyl bis-naphthalimide derivatives were synthesized to study the DNA binding ability and the cytotoxicity induced by reactive oxygen species. All of the compounds exhibited good DNA binding ability. And the structure cytotoxicity relationship revealed that the structure of 5h was the best choice.

LaMIR166a-mediated auxin biosynthesis and signalling affect somatic embryogenesis in Larix leptolepis.

Somatic embryogenesis (SE) involves complex molecular signalling pathways. Understanding molecular mechanism of SE in Larix leptolepis (L. leptolepis) can aid research on genetic improvement of gymnosperms. Previously, we obtained five LaMIR166a (miR166a precursor) -overexpression embryonic cell lines in the gymnosperm Larix leptolepis. The proliferation rates of pro-embryogenic masses in transgenic and wild-type lines were calculated. Overexpression of the miR166a precursor LaMIR166a led to slower proliferation. When pro-embryogenic masses were transferred to maturation medium, the relative expression of LaMIR166a and miR166a in the LaMIR166a-overexpression lines was higher than in the wild-type during SE, while LaHDZ31-34 expression levels also increased without negative control by miR166, suggesting that regulation of HD-ZIP III by miR166a exits stage-specific characteristics. The key indole-3-acetic acid (IAA) biosynthetic gene Nitrilase of L. leptolepis (LaNIT) was identified and the effects of miR166a on auxin biosynthesis and signalling genes were studied. During SE, LaNIT, Auxin response factor1 (LaARF1) and LaARF2 mRNA levels and IAA contents were markedly higher in LaMIR166a-overexpression lines, which revealed lower deformity rate of embryos, indicating endogenous IAA synthesis is required for somatic embryo maturation in L. leptolepis. Additionally, the IAA biosynthesis and signalling genes showed similar expression patterns to LaHDZ31-34, suggesting HD-ZIP III genes have a positive regulatory effect on LaNIT. Our results suggest miR166a and LaHDZ31-34 have important roles in auxin biosynthesis and signalling during SE, which might determine if the somatic embryo normally developed to mature in L. leptolepis.

Modulation of ovine SBD-1 expression by Saccharomyces cerevisiae in ovine ruminal epithelial cells.

BACKGROUND: The ovine rumen is involved in host defense responses and acts as the immune interface with the environment. The ruminal mucosal epithelium plays an important role in innate immunity and secretes antimicrobial innate immune molecules that have bactericidal activity against a variety of pathogens. Defensins are cationic peptides that are produced by the mucosal epithelia and have broad-spectrum antimicrobial activity. Sheep ß-defensin-1 (SBD-1) is one of the most important antibacterial peptides in the rumen. The expression of SBD-1 is regulated by the probiotic, Saccharomyces cerevisiae (S.c); however, the regulatory mechanism has not yet been elucidated. In the current study, the effects of S.c on the expression and secretion of SBD-1 in ovine ruminal epithelial cells were investigated using quantitative real-time PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA). In addition, specific inhibitors were used to block the nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB), p38, JNK, and ERK1/2 signalling pathways separately or simultaneously, to determine the regulatory mechanism(s) governing S.c-induced SBD-1 upregulation. RESULTS: Incubation with S.c induced release of SBD-1 by ovine ruminal epithelial cells, with SBD-1 expression peaking after 12 h of incubation. The highest SBD-1 expression levels were achieved after treatment with 5.2 × 107 CFU∙mL- 1 S.c. Treatment with S.c resulted in significantly increased NF-κB, p38, JNK, ERK1/2, TLR2, and MyD88 mRNA expression. Whereas inhibition of mitogen-activated protein kinases (MAPKs) and NF-κB gene expression led to a decrease in SBD-1 expression. CONCLUSIONS: S.c was induced SBD-1 expression and the S.c-induced up-regulation of SBD-1 expression may be related to TLR2 and MyD88 in ovine ruminal epithelial cells. This is likely simultaneously regulated by the MAPKs and NF-κB pathways with the p38 axis of the MAPKs pathway acting as the primary regulator. Thus, the pathways regulating S.c-induced SBD-1 expression may be related to TLR2-MyD88-NF-κB/MAPKs, with the TLR2-MyD88-p38 component of the TLR2-MyD88-MAPKs signalling acting as the main pathway.

Synthesis, DNA Binding, and Anticancer Properties of Bis-Naphthalimide Derivatives with Lysine-Modified Polyamine Linkers.

A series of bis-naphthalimide derivatives with different diamine linkers were designed and synthesized. All of the synthesized bis-naphthalimide derivatives were characterized by NMR and HRMS spectra. The binding ability between the compounds and CT DNA was evaluated by using UV-Vis titration experiments. The bis-naphthalimide compound with an ethylenediamine linker showed the largest binding constant with CT DNA. Hence, it was used as the model compound to study the DNA binding selectivity by UV-Vis titration aiming at different DNA duplexes. As a result, this compound showed binding preference to AT-rich duplexes. The DNA binding modes of the compounds were also measured by viscosity titration. The cytotoxicity of the compounds was evaluated by MTT assay. Compounds with 1,6-diaminohexane or 1,4-phenylenedimethanamine linkers showed higher cytotoxicity compared with other bis-naphthalimide derivatives.

Expression of hepaCAM inhibits bladder cancer cell proliferation via a Wnt/ß-catenin-dependent pathway in vitro and in vivo.

We previously established that hepatocyte cell adhesion molecule (hepaCAM), a typical structure of immunoglobulin (Ig)-like adhesion molecules, inhibited the proliferation and the progression of cultured human bladder cancer cells. As increasing evidence reveals that aberrant activation of canonical Wnt pathway is involved in the pathogenesis of bladder cancer, and ß-catenin serves as a pivotal molecule of Wnt pathway. Then, we explored whether the anti-proliferation effect of hepaCAM was associated with Wnt/ß-catenin pathway in human bladder cancer cells. The negative correlation between hepaCAM and ß-catenin in transitional cell carcinoma of bladder (TCCB) was found. Follow by, studied the effect of hepaCAM on the key elements of Wnt pathway. Here, Our researches showed that hepaCAM played a central role in modulating the Wnt/ß-catenin signaling pathway by interfering nuclear protein levels of ß-catenin, leading to down-regulate transcriptional activity of LEF/TCF and its target genes c-Myc and cyclinD1. Mechanistically, we demonstrated that hepaCAM-activated GSK3ß led to elevate the phosphorylation of ß-catenin, contributing to the aberrant translocation of ß-catenin. In addition, Anti-proliferation and associated molecular mechanisms of hepaCAM were demonstrated by using vivo experiment. In conclusion, our reports uncover that expression of hepaCAM suppresses the proliferation of bladder cancer cells through a Wnt/ß-catenin-dependent signaling pathway in vitro and in vivo.

Exosome-derived microRNA-29c induces apoptosis of BIU-87 cells by down regulating BCL-2 and MCL-1.

BACKGROUND: Aberrant expression of the microRNA-29 family is associated with tumorigenesis and cancer progression. As transport carriers, tumor-derived exosomes are released into the extracellular space and regulate multiple functions of target cells. Thus, we assessed the possibility that exosomes could transport microRNA- 29c as a carrier and correlations between microRNA-29c and apoptosis of bladder cancer cells. MATERIALS AND METHODS: A total of 28 cancer and adjacent tissues were examined by immunohistochemistry to detect BCL-2 and MCL-1 expression. Disease was Ta-T1 in 12 patients, T2-T4 in 16, grade 1 in 8, 2 in 8 and 3 in 12. The expression of microRNA-29c in cancer tissues was detected by quantitative reverse transcriptase PCR (QRT- PCR). An adenovirus containing microRNA-29c was used to infect the BIU-87 human bladder cancer cell line. MicroRNA-29c in exosomes was measured by QRT-PCR. After BIU-87 cells were induced by exosomes-derived microRNA-29c, QRT-PCR was used to detect the level of microRNA-29c. Apoptosis was examined by flow cytometry and BCL-2 and MCL-1 mRNA expressions were assessed by reverse transcription-polymerase chain reaction. Western blotting was used to determine the protein expression of BCL-2 and MCL-1. RESULTS: The expressions of BCL-2 and MCL-1 protein were remarkably increased in bladder carcinoma (p<0.05), but was found mainly in the basal and suprabasal layers in adjacent tissues. The expression of microRNA-29c in cancer tissues was negatively correlated with the BCL-2 and MCL-1. The expression level of microRNA-29c in exosomes and BIU-87 cells from the experiment group was higher than that in control groups (p<0.05). Exosome-derived microRNA-29c induced apoptosis (p<0.01). Although only BCL-2 was reduced at the mRNA level, both BCL-2 and MCL-1 were reduced at the protein level. CONCLUSIONS: Human bladder cancer cells infected by microRNA- 29c adenovirus can transport microRNA-29c via exosomes. Moreover, exosome-derived microRNA29c induces apoptosis in bladder cancer cells by down-regulating BCL-2 and MCL-1.

A new electrochemical sensor of nitro aromatic compound based on three-dimensional porous Pt-Pd nanoparticles supported by graphene-multiwalled carbon nanotube composite.

In this study, an electrochemical sensor of nitro aromatic compound based on three-dimensional porous Pt-Pd nanoparticles (Pt-Pd NPs) supported by reduced graphene oxide (rGO) nanosheets-multiwalled carbon nanotube (CNTs) nanocomposite (marked as Pt-Pd NPs/CNTs-rGO) was investigated for the first time. This hybrid nanocomposite has been prepared via a facile and versatile hydrothermal synthetic strategy while its structure and property are evaluated by X-ray diffraction (XRD), transmission electron microscopy (TEM) and electrochemical impedance spectroscopy (EIS). The result shows that 3D porous Pt-Pd NPs/CNTs-rGO nanocomposite has a large specific surface area of 326.6m(2)g(-1) and exhibited ultrahigh rate capability and good cycling properties at high rates. Electrochemical studies have been performed for the nitro aromatic compounds detection by using different pulse voltammetry (DPV) techniques. The proposed nanocomposite exhibited much enhanced elctrocatalytic activity and high sensitivity toward the detection of nitro aromatic compounds which compared with Pt-Pd NPs dispersed on functionalized rGO, Pt-Pd NPs dispersed on functionalized CNTs, rGO-CNTs and bare glass carbon electrode (GCE). On the basis of the above synergetic electrochemical sensing and synthesis procedure, the hybrid material can be recommended as a robust material for sensor-related applications. Moreover, the proposed sensor exhibits high reproducibility, long-time storage stability and satisfactory anti-interference ability.

Nuclear factor-κB signaling pathway is involved in phospholipase Cε-regulated proliferation in human renal cell carcinoma cells.

Phospholipase Cε (PLCε), a downstream effector of small GTPase superfamily, has been identified to play a crucial role in tumorigenesis. Previously, our studies have showed that PLCε promotes proliferation of renal cell carcinoma (RCC) cells. However, the molecular mechanisms by which PLCε enhances the survival phenotype of RCC cells are still not fully instructed. In the present study, we first demonstrated that PLCε was highly expressed and had a close correlation with Ki67 protein expression in RCC tissue samples. Further, we found that downregulation of PLCε expression repressed growth and induced apoptosis in RCC cells. In addition, we reported a mechanism by which knockdown of PLCε gene potently suppressed the nuclear factor kappa (NF-κB) signaling pathway through action on inhibitor of κB kinase. Moreover, silencing PLCε gene decreased vascular endothelial growth factor (VEGF) expression, which was a downstream growth factor of NF-κB signaling pathway. Finally, downregulation of VEGF was severely enhanced by treatment cells with NF-κB specific inhibitor BAY11-7028 in PLCε knockdown cells. Taken together, these findings suggest that PLCε promotes RCC cell growth via NF-κB-mediated upregulation of VEGF.

A new PKCα/ß/TBX3/E-cadherin pathway is involved in PLCε-regulated invasion and migration in human bladder cancer cells.

Although PLCε has been verified to enhance bladder cancer cell invasion, the signaling pathways responsible for this remain elusive. Protein kinase C (PKCα/ß), which is involved in cancer development and progression, has been demonstrated to be activated by PLCε. However, the roles of PKCα/ß in PLCε-mediated bladder carcinoma cell invasion and migration have not been clearly identified. In this study, to determine what role PKCα/ß plays in PLCε-mediated bladder cancer cell invasion and migration, we silenced PLCε gene by adenovirus-shPLCε in T24 and BIU-87 cells and then revealed that it significantly inhibited cell migration and invasion. Further research indicated that cell bio-function of PLCε-regulated was related with PKCα/ß activity. These in vitro findings were supported by data from bladder carcinoma patient samples. In 35 case bladder cancer tumor samples, PLCε-overexpressing tumors showed significantly higher positive rates of PKCα/ß membrane immunohistochemistry staining than PLCε-low-expressing tumors. Mechanistically, study further showed that PLCε knockdown gene induced E-cadherin expression and decreased TBX3 expression, both of which were dependent on PKCα/ß activity. In addition, we demonstrated that treatment cells with TBX3-specific shorting hairpin RNA (shRNA) up-regulated E-cadherin expression and inhibited cell invasion/migration. Moreover, in in vivo experiment, immunohistochemistry analysis of Ad-shPLCε-infected tumor tissue showed low expression levels of phospho-PKCα/ß and TBX3 and high expression levels of E-cadherin compared with those of the control group. In summary, our findings uncover that PKCα/ß is critical for PLCε-mediated cancer cell invasion and migration and provide valuable insights for current and future Ad-shPLCε and PKCα/ß clinical trials.

[Influence of hepatocyte cell adhesion molecule on gene expression profile of human bladder transitional cell carcinoma cell line].

OBJECTIVE: To investigate the changes of gene expression file in transitional cell carcinoma of bladder after hepatocyte cell adhesion molecule(hepaCAM) overexpression. METHODS: Affymetrix Human Genome U133 Plus 2.0 Array was used to investigate the changes of gene expression profile between adenovirus-green fluorescent protein(GFP) -hepaCAM group and GFP group in transitional cell carcinoma of bladder EJ cells.Significant Analysis of Microarray(SAM) was used to screen the differentially expressed genes, DAVID software was used to conduct gene ontology analysis and wikiPathway analysis based on the differentially expressed genes. Reverse transcription-polymerase chain reaction and Western blot were applied to verify microarray data. RESULTS: Compared with the GFP group, a total of 2469 genes were up-regulated or down-regulated by more than 2 times in the GFP-hepaCAM group. Among these genes, 1602 genes were up-regulated and 867 were down-regulated.Most of the differentially expressed genes were involved in the function of cell proliferation and cell cycle regulation. The mRNA expressions of nibrin, liver kinase B1, and cyclin D1 detected by reverse transcription-polymerase chain reaction in three different bladder cancer cell lines were consistent with the microarray data.The protein expressions of nibrin and liver kinase B1 in these three cell lines measured by Western blot were consistent with the mRNA expression. CONCLUSIONS: HepaCAM can alter the gene expression profile of bladder cancer EJ cells. The well-known anti-tumor effect of hepaCAM may be mediated by regulating the gene expression via multiple pathways.