A core-molecule-shell Au@PATP@Ag nanorod for nicotine detection based on surface-enhanced Raman scattering technology.

Nicotine is an addictive substance often found in tobacco and cigarette smoke and excessive exposure to it can cause various diseases. Herein, core-molecule-shell gold/4-aminothiophenol/silver nanorods (Au@PATP@Ag NRs) were prepared for quantitative detection of nicotine by using surface-enhanced Raman scattering (SERS) technology. The obtained Au@PATP@Ag NRs showed an outstanding SERS effect due to the plasticity of their morphology and the bimetallic synergistic effect between the excellent stability of Au and the highly enhanced effect of Ag. The Au@PATP@Ag NRs substrate exhibited an extremely high enhancement factor (EF) of 2.17 × 107. In addition, in-situ synthesized PATP was used as an internal standard to correct signal fluctuation and improve the reliability of quantitative nicotine detection. A wide linear dynamic range from 10-8 to 10-3 M was obtained and an ultra-low limit of detection (LOD) was about 3.12 × 10-9 M, which was superior to most of previously reported methods. This work has also been used for determining nicotine content in cigarettes and simulated environmental tobacco smoke by using a portable device. These results indicated that the developed SERS method had many potential applications in the quantitative determination of nicotine in real tobacco samples.

Systematic Analysis of the Grafting-Related Glucanase-Encoding GH9 Family Genes in Pepper, Tomato and Tobacco.

Grafting is an important agricultural practice to control soil-borne diseases, alleviate continuous cropping problems and improve stress tolerance in vegetable industry, but it is relatively less applied in pepper production. A recent study has revealed the key roles of ß-1, 4-glucanase in graft survival. We speculated that the GH9 family gene encoding glucanase may be involved in the obstacles of pepper grafting. Therefore, we performed a systematic analysis of the GH9 family in pepper, tomato and tobacco. A total of 25, 24 and 42 GH9 genes were identified from these three species. Compared with the orthologues of other solanaceous crops, the deduced pepper GH9B3 protein lacks a conserved motif (Motif 5). Promoter cis-element analysis revealed that a wound-responsive element exists in the promoter of tobacco NbGH9B3, but it is absent in the GH9B3 promoter of most solanaceous crops. The auxin-responsive related element is absent in CaGH9B3 promoter, but it presents in the promoter of tobacco, tomato, potato and petunia GH9B3. Tissue and induction expression profiles indicated that GH9 family genes are functionally differentiated. Nine GH9 genes, including CaGH9B3, were detected expressing in pepper stem. The expression patterns of NbGH9B3 and CaGH9B3 in grafting were different in our test condition, with obvious induction in tobacco but repression in pepper. Furthermore, weighted correlation network analysis (WGCNA) revealed 58 transcription factor genes highly co-expressed with NbGH9B3. Eight WRKY binding sites were detected in the promoter of NbGH9B3, and several NbWRKYs were highly co-expressed with NbGH9B3. In conclusion, the missing of Motif 5 in CaGH9B3, and lacking of wound- and auxin-responsive elements in the gene promoter are the potential causes of grafting-related problems in pepper. WRKY family transcription factors could be important regulator of NbGH9B3 in tobacco grafting. Our analysis points out the putative regulators of NbGH9B3, which would be helpful to the functional validation and the study of signal pathways related to grafting in the future.

Genetic mutation of TRPV2 induces anxiety by decreasing GABA-B R2 expression in hippocampus.

Transient receptor potential vanillic acid 2 (TRPV2) are well recognized for their contributions to neuronal development, cardiac function, immunity and cancer. However, the precise roles for this thermo TRPchannels in neurological disorder remain unknown. In this study, we employed the CRISPR/Cas9 system to generate genetic mutations of TRPV2. Genetic mutation of TRPV2 resulted in autistic-like phenotypes in mice accompanied with the disordered electrical signals recorded by multi-channels in vivo. To determine possible molecular mechanisms, western blotting was further used to assess the possible involvement of several autism-related proteins. The significantly decreased expression of the R2 subunit of the GABA-B receptor in the hippocampus was observed. Together, our findings suggest that genetic mutation of TRPV2 induces autism-like behavior, results in decreased expression of the R2 subunit of the GABA-B receptor.

Fluorescent Assay of FEN1 Activity with Nicking Enzyme-Assisted Signal Amplification Based on ZIF-8 for Imaging in Living Cells.

Flap endonuclease 1 (FEN1) participates in both DNA replication and repair to maintain the stability and integrity of the genome. As a potential tumor marker, detecting FEN1 activity could be an effective strategy for cancer diagnosis. In this work, a fluorescence assay was developed for sensitive detection of FEN1 using biomineralized metal-organic framework nanoparticles (ZIF-8 NPs) to codeliver the encapsulated proteins and DNA probes to living cells. After uptake into cells, the biomineralized ZIF-8 NPs were biodegraded to release proteins and DNA probes under an acid environment. In the presence of FEN1, the cleaved flap triggered by FEN1 hybridized with a hairpin probe to fabricate a double-stranded DNA structure which had a cleavage site of the nicking enzyme, causing the fluorophore to move away from the quencher. Assisting the nicking enzyme, an amplified fluorescence signal was obtained after several recycling. Confocal imaging indicated that this fluorescence assay could distinguish cancer cells from normal cells. Therefore, this strategy would contribute to the prediction and diagnosis in early-stage cancer.

Cancer Cell-Membrane Biomimetic Boron Nitride Nanospheres for Targeted Cancer Therapy.

PURPOSE: Nanomaterial-based drug-delivery systems allowing for effective targeted delivery of smallmolecule chemodrugs to tumors have revolutionized cancer therapy. Recently, as novel nanomaterials with outstanding physicochemical properties, boron nitride nanospheres (BNs) have emerged as a promising candidate for drug delivery. However, poor dispersity and lack of tumor targeting severely limit further applications. In this study, cancer cell-membrane biomimetic BNs were designed for targeted anticancer drug delivery. METHODS: Cell membrane extracted from HeLa cells (HM) was used to encapsulate BNs by physical extrusion. Doxorubicin (Dox) was loaded onto HM-BNs as a model drug. RESULTS: The cell-membrane coating endowed the BNs with excellent dispersibility and cytocompatibility. The drug-release profile showed that the Dox@HM-BNs responded to acid pH, resulting in rapid Dox release. Enhanced cellular uptake of Dox@HM-BNs by HeLa cells was revealed because of the homologous targeting of cancer-cell membranes. CCK8 and live/dead assays showed that Dox@HM-BNs had stronger cytotoxicity against HeLa cells, due to self-selective cellular uptake. Finally, antitumor investigation using the HeLa tumor model demonstrated that Dox@HM-BNs possessed much more efficient tumor inhibition than free Dox or Dox@BNs. CONCLUSION: These findings indicate that the newly developed HM-BNs are promising as an efficient tumor-selective drug-delivery vehicle for tumor therapy.

Deletion of Kv10.2 Causes Abnormal Dendritic Arborization and Epilepsy Susceptibility.

The abnormal function of the voltage-gated potassium channel Kv10.2 can induce epilepsy. However, the physiological function of Kv10.2 in the central nervous system remains unclear. In this study, we found that Kv10.2 knockout (KO) increased the complexity of neurons in the CA3 subarea of hippocampus. Kv10.2 KO led to enlarged somata, elongated dendritic length, and increased the number of dendritic tips in cultured rat hippocampus neurons. Kv10.2 KO also increased Synapsin I and PSD95 protein density in cultured rat hippocampal neurons. Whole cell patch-clamp recordings of brain slices in the CA3 subarea of hippocampus revealed that Kv10.2 KO increased the amplitude of spontaneous excitatory postsynaptic currents (sEPSC) and miniature excitatory postsynaptic currents (mEPSC), depolarized the resting membrane potential and increased the action potential firing, reduced the rheobase and increased the input resistance, which results in enhanced neuronal excitability. Furthermore, we made electroencephalogram (EEG) recordings of brain activity in freely moving rats before and after inducing seizures by pentylenetetrazole (PTZ) injection. Kv10.2 KO rats dramatically increased the EEG amplitude during epilepsy. Behavioral observation after seizure induction revealed that Kv10.2 KO rats demonstrated shortened onset latency, prolonged duration, and increased seizure severity when compared with wild type rats. Therefore, this study provides a new link between Kv10.2 and neuronal morphology and higher intrinsic excitability.

Label-Free Imaging of Flap Endonuclease 1 in Living Cells by Assembling Original and Multifunctional Nanoprobe.

Flap endonuclease 1 (FEN1) becomes a potential tumor marker since it is closely related to cancer occurrence and development. Here, a poly dA20-mediated nanoprobe (AuNPs-poly dA20-poly dT20) was designed for FEN1 detection. Poly dA20 segment at the 3'- end of ssDNA adsorbed on AuNPs due to its strong affinity interaction with Au (stronger than Au-S bond), while the poly dT20 segment at the 5'- end overhangs. This nanoprobe not only worked as effective fluorescence quencher but also as the original nanosubstrate of FEN1. OliGreen adsorbed on poly dT20 emits strong green fluorescence because of its high sensitivity and selectivity toward thymine. However, it is quenched on the nanoprobe. In the presence of FEN1, it recognizes the overhanging poly dT20 segment and cleaves it efficiently, turning on the fluorescence of OliGreen. This indicates that the assembled nanoprobe is an effective artificial substrate to FEN1, although it is completely different from previously reported substrates that are all composed of dsDNA with a flap strand. This proposed nanoprobe was used to detect FEN1 not only in vitro but also in vivo. The method was simple, which avoided complex labeling procedures. It had a wide linear range from 0.05 U to 2 U, with the lowest detection limit of 0.007 U. Confocal imaging can distinguish cancer cells from normal cells, demonstrating its potential in clinical diagnostic and therapeutic monitoring.

Increased Expression of Kv10.2 in the Hippocampus Attenuates Valproic Acid-Induced Autism-Like Behaviors in Rats.

The role of potassium channels provides suggestive evidence for the etiology of autism. The voltage-gated potassium channel Kv10.2 (KCNH5) is widely expressed in the brain. However, the inherent relationship between Kv10.2 and autism is still unclear. Herein, a rat valproic acid (VPA)-induced autism spectrum disorder model was established. The expression level of Kv10.2 was obviously decreased in the hippocampus of VPA rats. Kv10.2 was mainly localized in neurons. Subsequently, a recombinant lentivirus expressing Kv10.2 was used to upregulate the expression of Kv10.2 in the hippocampus of VPA-exposed rats. The results were promising as injection of the Kv10.2 lentivirus in the hippocampus relieved anxiety and stereotypical behavior, and improved the social and exploratory abilities of rats that were prenatally exposed to VPA. In addition, spectral analysis of electroencephalogram data revealed that animals exposed to VPA exhibited increased high-frequency activity compared with the control rats, and this activity recovered to a certain extent after upregulation of Kv10.2 expression by lentivirus injection. These results suggest that changes in Kv10.2 may play an important role in the etiology of autism, thus providing a promising direction for further research on autism.

A visible light photoelectrochemical sandwich aptasensor for adenosine triphosphate based on MgIn2S4-TiO2 nanoarray heterojunction.

This work designed a MgIn2S4-TiONA heterojunction by growing MgIn2S4 nanoplates on TiO2 nanowire array (TiONA) for preparation of visible light photoelectrochemical (PEC) sensing platform. The heterojunction exhibited strong absorption of visible light, large surface area and high loading of biomolecules, leading to high sensing sensitivity. Using adenosine triphosphate (ATP), a marker of cell vitality, as the target model, a PEC sandwich aptasensor was constructed by immobilizing capture DNA1 on MgIn2S4 surface. In the presence of ATP and signal DNA2 with terminal ferrocene as the electron donor, a sandwiched DNA1-ATP-DNA2 complex could be formed on the PEC aptasensor. The aptasensor showed excellent performance with a wide linear range from 50 fM to 100 nM and a detection limit of 20 fM. The sensing performance including specificity, reproducibility, stability and practical use were also evaluated, showing promising application of the MgIn2S4-TiONA heterojunction in PEC biosensing.

A glassy carbon electrode modified with a composite consisting of gold nanoparticle, reduced graphene oxide and poly(L-arginine) for simultaneous voltammetric determination of dopamine, serotonin and L-tryptophan.

A glassy carbon electrode (GCE) was modified with poly(L-arginine) (P-Arg), reduced graphene oxide (rGO) and gold nanoparticle (AuNP) to obtain an electrode for simultaneous determination of dopamine (DA), serotonin (5-HT) and L-tryptophan (L-Trp) in the presence of ascorbic acid (AA). The modified GCE was prepared via subsequent 'layer-by-layer' deposition using an electrochemical technique. The surface morphology of the modified electrode was studied by scanning electron microscopy, and electrochemical characterizations were carried out via cyclic voltammetry and electrochemical impedance spectroscopy. The modified electrode showed excellent electrocatalytic activity toward DA, 5-HT and L-Trp at pH 7.0. Figures of merit for the differential pulse voltammetric reponse are as follows: (a) Response to DA is linear in two intervals, viz. 1.0-50 nM and 1.0-50 µM DA concentration range, the typical working voltage is 202 mV (vs. Ag/AgCl), and the detection limit is 1 nM (at an S/N ratio of 3). For 5-HT, the respective data are 10 to 500 nM and 1.0 to 10 µM, 381 mV, and 30 nM. For L-Trp, the respective data are 10-70 nM and 10-100 µM, 719 mV, and 0.1 µM. The modified GCE is fairly selective. It was successfully applied to the simultaneous determination of DA, 5-HT, and L-Trp in spiked urine samples, and high recovery rates were found. Graphical abstract Schematic presentation of the voltammetric sensor based on a glassy carbon electrode modified with poly(L-arginine), reduced graphene oxide (rGO) and gold nanoparticle (GCE/P-Arg/ErGO/AuNP) for simultaneous determination of dopamine (DA), serotonin (5-HT) and L-tryptophan (L-Trp).

Ultra-sensitive electrochemical detection of oxidative stress biomarker 8-hydroxy-2'-deoxyguanosine with poly (L-arginine)/graphene wrapped Au nanoparticles modified electrode.

An innovative electrochemical sensor assembly relying on a simple "green" electrochemical reduction route is presented for the sensitive detection of 8-hydroxy-2'-deoxyguanosine (8-OHdG), the most abundant oxidative product of DNA. The sensing film consisted of poly (L-arginine) and graphene wrapped Au nanoparticles was fabricated on glassy carbon electrode (GCE/P-Arg/ErGO-AuNPs) using subsequent 'layer-by-layer' regime through electrochemical technique. The proposed method was also successfully applied for the quantification of 8-OHdG in the presence of interfering biomolecules like ascorbic acid and uric acid. Scanning electron microscopy (SEM) was utilized to characterize the surface morphology of the composite electrode. Electrochemical characterizations of the bare and modified electrodes were carried out via cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). According to differential pulse voltammetry (DPV) results, there were linear relationships between the peak currents and the concentrations in the ranges of 1.0-100 nM (R2 = 0.996), and 0.5-10 µM (R2 = 0.990), with a detection limit (S/N = 3) of 1.0 nM. Furthermore, the proposed sensor was successfully applied for the determination of target analyte in human urine samples and a very high recovery percentage was obtained.

TiO2 nanowire arrays modified with a simultaneous "etching, doping and deposition" technique for ultrasensitive amperometric immunosensing.

In this work, an ultrasensitive immunosensing scaffold was structured with TiO2 nanowire (TiNW) arrays modified with molybdenum (Mo) and MoS2 flakes by a triplex "etching, doping and deposition" technique. The triply modification of TiNW arrays improved their electron transfer, and the decoration of MoS2 flakes on TiNW arrays increased both the conductivity and the specific surface area of TiNW. Accordingly, the triply modified TiNW arrays provided a biocompatible microenviroment for the biomolecules and high specific surface area to load big amount of biomolecules. The immunosensor was prepared by immobilizing capture antibody on the scaffold surface with double amino-reactive crosslinker, and the tracing labels were prepared by immobilizing signal antibody and horseradish peroxidase molecules on cylinder-shaped TiO2 nanorods. After sandwich-type immunoreaction, the tracing labels were quantitatively captured on the immunosensor surface for the detection of carcinoembryonic antigen as a model analyte. This amperometric method showed a linear range of 0.001 and 150ngmL-1 with a detection limit of 0.5pgmL-1. This work provided a promising platform for sensitive amperometric immunosensing of protein biomarkers.