Detection of Cadmium(II) in Aquatic Products Using a Rolling-Circle Amplification-Coupled Ratio Fluorescent Probe Based on an Aptamer-Peptide Conjugate.

The existing aptamers for cadmium (Cd2+), the common toxic heavy metal contaminant in food, cannot meet the requirements for detecting Cd2+ in rapid detection methods. In previous work, we found that coupling aptamer-peptide conjugates (APCs) with peptides and aptamers can provide a less disruptive method with a significantly improved affinity. Moreover, we found that the spatial conformation of aptamers and peptides is crucial for obtaining proper affinity in APC. Therefore, we describe a simple design strategy to obtain a series of APCs with different affinities by designing peptide orientations (N-terminal, C-terminal). The best affinity was found for APC(C1-N) with a binding constant (Ka) of 2.23 × 106 M-1, indicating that the APC(C1-N) affinity was significantly increased by 829.17% over aptamer. Finally, a rolling-circle amplification (RCA)-coupled ratio fluorescence-based biosensor for Cd2+ detection was established with a detection limit of 0.0036 nM, which has great potential for practical aquatic product detection.

RAB3D, upregulated by aryl hydrocarbon receptor (AhR), promotes the progression of prostate cancer by activating the PI3K/AKT signaling pathway.

Many patients with prostate cancer (PCa) cannot be diagnosed until an advanced stage, which make PCa become a large threat to human health. It is an urgent need to explore novel biomarkers for accurate diagnosis and targets for the effective treatment of PCa. This study aimed to investigate the effects of RAB3D (which belongs to the secretory Rab GTPases) on the progression of PCa. The results showed that RAB3D was highly expressed in PCa tissues compared to normal tissues according to the gene expression omnibus dataset. Consistent with the bioinformatics results, RAB3D exhibited a higher expression in PCa cells. Overexpression of RAB3D promoted the proliferation, migration, and invasion of PCa cells, whereas the knockdown of RAB3D led to the opposite results. The procancer effects of RAB3D were further confirmed by the in vivo growth of xenograft model. Subsequently, RAB3D upregulated the PI3K/AKT signaling pathway both in vivo and in vitro. LY294002 (a PI3K inhibitor) rescued the RAB3D upregulation-induced promotion of malignant phenotypes of PCa cells. Furthermore, the transcription activity of RAB3D was found to be enhanced by aryl hydrocarbon receptor (AhR; a transcription factor). The AhR silencing-induced inhibition of the proliferation and migration of PCa cells was reversed by the overexpression of RAB3D. Taken together, RAB3D, upregulated by AhR, promotes the PCa progression by activating the PI3K/AKT signaling pathway.

An aptasensor for the detection of Pb2+ based on photoinduced electron transfer between a G-quadruplex-hemin complex and a fluorophore.

Due to the threat to health of heavy metal contamination, simple and rapid detection methods for heavy metals are an urgent needed in environment protection and food safety. In this work, we have developed a fluorescent aptasensor for the 'turn-off' model detection of Pb2+ . The key feature of the aptasensor is that the dye-labelled nucleic acid strand can be folded into a G-quadruplex structure in the presence of Pb2+ . This conformational change induces photoinduced electron transfer (PET) between a G-quadruplex-hemin complex and 6-carboxyrhodamine X (ROX), which results in fluorescence quenching of ROX. We systematically investigated the interaction mechanism between Pb2+ and the aptasensor and the effects of several environmental parameters were also studied. Under the optimum conditions, the proposed method exhibited a good liner relationship between the concentration of Pb2+ and fluorescence quenching efficiency in the range 25-500 nM and the limit of detection was 1.02 nM. In addition, this method also manifested good performance in spiked lettuce samples with satisfactory recoveries of 87.10-109.6%. This target-induced PET platform can be further expanded to other heavy metal analysis.

[Dynamics of soil respiration and its influencing factors in urban forests under nitrogen addition]. / æ°®æ·»åŠ ä¸‹åŸŽå¸‚æ£®æž—åœŸå£¤å‘¼å¸åŠ¨æ€å˜åŒ–åŠå ¶å½±å“å› ç´ .

Urban forest is an important carbon pool, soil respiration of which is an important part of terrestrial carbon cycle. To understand the dynamics and influencing factors of soil respiration in urban forest under the background of increasing nitrogen deposition, we conducted dynamic observation on soil respiration rate, temperature, moisture and chemical properties by adding 0 (CK), 50 (LN), 100 (HN) kg N·m-2·a-1 ammonium nitrate to a typical urban forest. The results showed that soil respiration had significant seasonal variation, which was not affected by nitrogen addition. Soil respiration was significantly correlated with soil temperature. The interaction between soil temperature and soil moisture could better explain the variation of soil respiration. Nitrogen addition changed temperature sensitivity of soil respiration, with the order of Q10 values as LN (2.12) > CK (2.10) > HN (2.05). Soil nitrate concentration, soil soluble organic carbon, pH, soil carbon to nitrogen ratio had significant correlation with soil respiration. The positive effect of nitrogen deposition on soil respiration was mainly in the growing season, with slightly inhibitive effect in the non-growing season.

The Dual Function of Aryl Hydrocarbon Receptor in Bladder Carcinogenesis.

BACKGROUND/AIM: Although Aryl hydrocarbon receptor (AhR) may be critical to several types of cancers, the function of AhR for carcinogenesis of bladder cancer (BC) is still inconclusive. We, therefore, sought to examine the involvement of AhR in bladder carcinogenesis. MATERIALS AND METHODS: We examined the AhR expression of human BC and N-butyl-N-(4-hydroxybutyl)-induced bladder carcinogenesis in AhR-deficient mice. RESULTS: There was a significantly higher expression of AhR in non-muscle-invasive BC compared to normal tissue and muscle-invasive BC (MIBC). The incidence of MIBC in AhR-deficient mice (87.5%) was significantly higher than wild-type mice (9.5%, p<0.01). In cell invasion assay, the induction of AhR signaling resulted in attenuation of BC cell invasiveness and proliferation. CONCLUSION: These results suggest that AhR may be essential for the initiation of carcinogenesis and attenuated the invasion of BC cells; this signaling may have a dual function in bladder carcinogenesis.

Target-induced in-situ formation of fluorescent DNA-templated copper nanoparticles by a catalytic hairpin assembly: application to the determination of DNA and thrombin.

A fluorometric method is described for nucleic acid signal amplification through target-induced catalytic hairpin assembly with DNA-templated copper nanoparticles (Cu NPs). The toehold-mediated self-assembly of three metastable hairpins is triggered in presence of target DNA. This leads to the formation of a three-way junction structure with protruding mononucleotides at the 3' terminus. The target DNA is released from the formed branched structure and triggers another assembly cycle. As a result, plenty of branched DNA becomes available for the synthesis of Cu NPs which have fluorescence excitation/emission maxima at 340/590 nm. At the same time, the branched structure protects the Cu NPs from digestion by exonuclease III. The unreacted hairpins are digested by exonuclease III, and this warrants a lower background signal. The method can detect ssDNA (24 nt) at low concentration (44 pM) and is selective over single-nucleotide polymorphism. On addition of an aptamer, the strategy can also be applied to the quantitation of thrombin at levels as low as 0.9 nM. Graphical abstractSchematic representation of target-induced catalytic hairpin assembly to form branched DNA template for the in situ synthesis of fluorescent Cu nanoparticles.

A "turn-on" fluorometric assay for kanamycin detection by using silver nanoclusters and surface plasmon enhanced energy transfer.

A rapid method is described for the determination of the antibiotic kanamycin. It integrates a kanamycin-binding aptamer and surface plasmon enhanced energy transfer (SPEET) between DNA-templated silver nanoclusters (AgNCs) and gold nanoparticles (AuNPs). The AgNCs and AuNPs were selected as energy donor and energy acceptor, respectively. The aptamer was designed to regulate the energy transfer between AgNCs and AuNPs. The aptamer was adsorbed on the AuNPs. Upon addition of kanamycin, the aptamer-kanamycin complex is formed, and this results in the aggregation of the AuNPs in high salt concentration, the formation of a blue coloration, and in the suppression of the SPEET process. The fluorescence of the AgNCs (with excitation/emission peaks at 560/600 nm) is quenched by the aptamer protected AuNPs in absence of kanamycin. The fluorescence on addition of kanamycin increases linearly in the 5 to 50 nM concentration range, with a lower detection limit of 1.0 nM (at S/N = 3). The assay can be performed within 30 min. It was successfully applied to the determination of kanamycin in spiked milk samples, and recoveries ranged between 90.2 and 95.4%. Conceivably, the strategy has a wide potential for screening by simply changing the aptamer. Graphical abstract Schematic presentation of the aptamer regulated surface plasmon enhance energy transfer (SPEET) between silver nanoclusters (Ag NCs) and gold nanoparticles (Au NCs) in high salt concentration buffer, and of the procedure for the detection of kanamycin.

Aryl hydrocarbon receptor enhances the expression of miR-150-5p to suppress in prostate cancer progression by regulating MAP3K12.

It has been reported that mircoRNAs (miRNAs) can act as tumor inhibitors in multiple malignant tumors. As a tumor suppressor, miR-150-5p has been reported in some cancers. However, the biological impacts of miR-150-5p in prostate cancer is not fully elaborated. This study aims to explore the biological role and mechanism of miR-150-5p in prostate cancer. The expression level of miR-150-5p was examined with Quantitative real time polymerase chain reaction (qRT-PCR). Moreover, Kaplan Meier analysis revealed that downregulation of miR-150-5p predicted unfavorable prognosis for patients with prostate cancer. To identify the inhibitory effects of miR-150-5p on the cellular processes of prostate cancer, gain-of function assay was conducted. Next, the inhibitory effects of Tetrachlorodibenzo-p-dioxin (TCDD) and 3,3'-Diindolylmethane (DIM) on the proliferation and invasion of prostate cancer cells were demonstrated. Knockdown of Ahr reversed the TCDD/DIM-mediated proliferation and invasion. The expression level of CYP1A1 also was measured to confirm that Ahr was activated by TCDD or DIM in prostate cancer cells. Mechanism experiments revealed that MAP3K12 is a target mRNA of miR-150-5p in prostate cancer cells. In conclusion, Aryl hydrocarbon receptor enhances the expression of miR-150-5p to suppress cell proliferation and invasion in prostate cancer by regulating MAP3K12.

Aryl Hydrocarbon Receptor Suppresses the Prostate Cancer LNCaP Cell Growth and Invasion by Promoting DNA Damage Response Under Oxidative Stress.

Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that interacts with multiple signaling pathways during prostate development. In the present study, LNCaP cells were knocked down of AhR by siRNA, or treated with the AhR agonist 3-methylcholanthrene (3MC). The effects of AhR on LNCaP cells and the associated mechanisms were studied both under normal condition and under hydrogen peroxide (H2O2)-induced oxidative stress. MTT, transwell chamber assays and flow cytometry were employed to investigate cell proliferation, invasion, and apoptosis, respectively, whereas the DNA damage response (DDR) signaling (phosphorylation of ataxia-telangiectasia mutated [ATM], check-point kinase 2 [Chk2], histone H2AX, p53, and cleaved poly-ADP-ribose polymerase [PARP]) was detected by western blotting. Exposure of LNCaP cells to H2O2 inhibited their viability and migration, and induced apoptosis, at a greater extent compared with the culture under normal conditions. In addition, the oxidative stress increased p-ATM, p-Chk2, p-p53, and p-H2AX expression levels significantly. Knockdown of AhR attenuated the aforementioned effects caused by H2O2-induced oxidative stress. Activation of AhR by 3MC treatment, further aggravated these changes of LNCaP cells on oxidative stress. The findings indicated that AhR suppresses the viability and migration of LNCaP cells notably under oxidative stress, and this process is associated with positive regulation of the responses to oxidative DNA damage.

Growth and Modeling of Staphylococcus aureus in Flour Products under Isothermal and Nonisothermal Conditions.

This study was conducted to investigate the growth of Staphylococcus aureus in traditional Chinese flour products under isothermal (10, 15, 20, 25, 30, and 37°C) and nonisothermal (10 to 20, 20 to 30, and 25 to 37°C) conditions. Then, models for the growth of S. aureus in flour products as a function of storage temperature, pH, and water activity (aw) were developed, and the goodness of fit of models was evaluated using the determination coefficient (R2), root mean square error (RMSE), bias factor (Bf), and accuracy factor (Af). Based on the above information, S. aureus growth in steamed bread under nonisothermal conditions was predicted from experiments performed under isothermal conditions. It was shown that different combinations of temperature and aw in flour products have a strong influence on the growth of S. aureus . The modified Gompertz model was found to be more suitable for describing the growth data of S. aureus in flour products, with an R2 of >0.99 and an RMSE of <0.37. The newly developed secondary models were validated, and for the specific growth rate and the lag time, the R2 values were 0.96 and 0.97, Af was 1.12 and 1.06, and Bf was 1.13 and 1.05, respectively. The predicted nonisothermal growth curves of S. aureus were in agreement with the reported experimental ones, with RMSE <0.29, Af value 1.02 to 1.09, and Bf value 0.92 to 0.99. These results indicated that the predictive models provided useful information for the establishment of safety standards and a risk assessment for S. aureus in flour products.

Aryl hydrocarbon receptor signaling involved in the invasiveness of LNCaP cells.

There is now mounting evidence that the aryl hydrocarbon receptor (AhR) plays an important role in physiologic responses such as development, cell cycle regulation, immune function and also malignant transformation in various tissues. The strong nuclear AhR expression is observed in the invasive phenotype, and an elevated nuclear AhR expression is associated with a poor prognosis of human prostate cancer. On the other hand, there are conflicting results that the AhR deficiency results in increased susceptibility to prostate tumors in mouse model. In the present study, we investigated AhR expression and its role in the growth and invasiveness of human prostate cancer cells. The AhR protein expression was detected in prostate cancer cell lines and human prostate cancer tissues. A small interfering RNA targeting AhR, constitutive active AhR expression vector, and AhR agonist and antagonist were used to moderate its expression and signaling. The induction of AhR signaling attenuated invasiveness of prostate cancer cells without affecting the cellular growth rate. These results suggest that AhR signaling in prostate cancer cells facilitates invasion of these cells, and modulation with this signaling can be a potential therapeutic target of invasive tumors.

Coupling purification and in situ immobilization process of monoclonal antibodies to clenbuterol for immunosensor application.

Clenbuterol (CL), which promotes the growth of muscular tissue and the reduction of body fat in pigs and cattle, has been confirmed to be a potential hazard to human health. In this study, a monoclonal antibody to clenbuterol (CL mAb) from a hybridoma culture supernatant was purified by an aqueous two-phase system (ATPS) at different polyethylene glycol (PEG) concentrations, PEG molecular weights, pH values, and NaCl concentrations. Then the CL mAb was immobilized in situ by directly adding polystyrene microspheres (PSMSs) into a PEG phase containing CL mAb. Using the immobilized antibody, an immunosensor was constructed to detect the CL residues in pork samples. The results showed that using an ATPS composed of 15% (w/w) PEG6000, 15% (w/w) phosphate, and 15% (w/w) NaCl at pH 8.0, the partition coefficient was 7.24, the activity recovery was 87.86%, and the purification fold was 2.88. The PEG-CL mAb-PSMS retained approximately 98% of its initial activity after 30-ml phosphate buffer (PBS) washings. After 30days of storage, the CL mAb-PSMS lost nearly 75% of its activity, whereas the PEG-CL mAb-PSMS retained as much as 95% of its initial activity. Furthermore, the constructed immunosensor obtained recoveries of 90.5 to 102.6% when applied to pork samples spiked with CL.

Antitumor effects of mutant endostatin are enhanced by Bcl-2 antisense oligonucleotides in UM-UC-3 bladder cancer cell line.

BACKGROUND: Endostatin is a potent inhibitor of tumor angiogenesis. In the preliminary studies, we developed a mutant endostatin containing Arg-Gly-Asp-Arg-Gly-Asp (RGDRGD) sequences. In this study, we compared the antitumor effects of mutant endostatin and Bcl-2 antisense oligonucleotides both in combination and individually. METHODS: The artificially synthesized Bcl-2 ASODN (antisense oligonucleotides) included a translation-initiation site and was transfected into the bladder cancer cells by Lipofectamine. Cell growth was investigated by the tumor cell growth chart, MTT assay, caspase-3 activity detection assay, AO/EB fluorescein stain, and the annexin V-FITC apoptosis detection assay. In the in vivo study, UM-UC-3 bladder cancer cells were subcutaneously implanted into nude mice and the growth of tumor was examined. The ultrastructure of the tumor tissues in the treated and control groups were observed. RESULTS: The cell growth chart showed that the cell population of the treated combination group decreased by 52.04% compared to the control group. The inhibition rate of the treated combination group was (79.66 ± 6.79)%, whereas those of the individual ASODN and ES groups were (53.39 ± 3.22)% and (50.22 ± 5.46)% respectively. In the caspase-3 activity detection using AO/EB fluorescein stain and annexin V-FITC apoptosis detection assay, the co-inhibitory effect was higher than the individual inhibitory effects (P < 0.05). There were significant differences in the inhibition of the solid tumor growth in the in vivo study. CONCLUSIONS: Our findings indicated that Bcl-2 antisense oligonucleotides enhance the antitumor effects of mutant endostatin both in vitro and in vivo. We noted the synergistic effects of Bcl-2 antisense oligonucleotides combined with mutant endostatin.

Clinical Usefulness of Corticotropin Releasing Hormone Testing in Subclinical Cushing's Syndrome for Predicting Cortisol Replacement after Adrenalectomy.

PURPOSE: The purpose of this study was to investigate the clinical and hormonal features of patients with incidentally discovered adrenal adenomas in relation to corticotropin releasing hormone (CRH) testing and the clinical outcome of adrenalectomy. MATERIALS AND METHODS: Twenty-three consecutive patients with incidentally detected adrenal adenomas were included in this retrospective study. All the patients underwent abdominal computed tomography scans and hormonal assays, including assessment of circadian rhythms of plasma cortisol and corticotropin (adrenocorticotropic hormone, ACTH), a corticotropin stimulation test, and low-dose and high-dose dexamethasone tests. The patients were reevaluated at regular intervals (6, 12, and 24 months) for a median period of 24 months. Subclinical Cushing's syndrome (SCS) was diagnosed in patients with subtle hypercortisolism who did not present clinical signs of Cushing's syndrome. RESULTS: We calculated the responsive index (peak value of ACTH in CRH test/baseline value of ACTH in CRH test). Of 23 patients, 6 had Cushing's syndrome, 8 had SCS, and 9 had a non-functioning tumor. All patients underwent laparoscopic adrenalectomy. Several patients (5 of 6 with Cushing's syndrome and 2 of 8 with SCS) required cortisol replacement therapy after surgery. The remaining patients required no hormonal replacement after surgery. Those who required hormone replacement had a responsive index of less than 1.2. Those who did not need hormone replacement therapy had a responsive index of more than 2.0. CONCLUSIONS: In our limited experience, the responsive index of the CRH test might be a valuable tool for predicting the need for cortisol replacement after surgery in patients with SCS.

Testosterone promotes DNA damage response under oxidative stress in prostate cancer cell lines.

BACKGROUND: Sustained chronic inflammation and oxidative stress in the prostate promote prostate carcinogenesis. The process of oncogenic transformation leads to enhanced DNA damage and activates the checkpoint network that functions as an inducible barrier against cancer progression. Here, we analyzed the effects of testosterone on the DNA damage response in prostate cancer cells to assess whether testosterone functions a barrier to cancer progression under the oxidative stress. METHODS: We examined the effects of testosterone on components of the DNA damage response pathway, including ATM (ataxia-telangiectasia-mutated kinase), H2AX (histone H2AX variant), and Chk2 (checkpoint kinase2) in prostate cancer cell lines, treated with various concentration of hydrogen peroxide (H(2) O(2) ). Cellular apoptosis was quantified by poly (ADP-ribose) polymerase (PARP) cleavage and flow cytometry. RESULTS: H(2) O(2) induced apoptosis and phosphorylation of ATM, Chk2, and H2AX in LNCaP cells. An ATM inhibitor, Ku55933, reduced H(2) O(2) -induced apoptosis in LNCaP and 22Rv1 cells. Androgen treatments increased H(2) O(2) -induced activation of the DNA damage response and PARP cleavage, but not when the H(2) O(2) -treated cells were also treated with the anti-androgen flutamide. The ATM inhibitor Ku55933 inhibited androgen-induced phosphorylation of ATM and PARP cleavage. CONCLUSIONS: DNA damage responses play important roles in the maintenance of the cell homeostasis in response to oxidative stress. Our results indicated that under oxidative stress androgen signaling may induce apoptosis by activating the DNA damage response.

Testosterone augments polyphenol-induced DNA damage response in prostate cancer cell line, LNCaP.

Recently, we reported that combined ingestion of soy isoflavones and curcumin significantly decreased the serum level of prostate-specific antigen based on a randomized placebo-controlled double-blind clinical study. We investigated whether these polyphenols inhibited the proliferation of prostate cancer cells by activating a DNA damage response. The effects of isoflavones and curcumin on the expression and phosphorylation of ataxia-telangiectasia-mutated kinase (ATM), histone H2AX variant (H2AX) and checkpoint kinase2 (Chk2) were examined in LNCaP cells. The induction of apoptosis in LNCaP cells was evaluated by poly(ADP-ribose) polymerase (PARP) cleavage. Furthermore, the effects of a testosterone supplement on modulation of the DNA damage response were examined. Combined treatment of isoflavones and curcumin additively suppressed cellular proliferation and induced phosphorylation of ATM, histone H2AX, Chk2 and p53. Testosterone augmented the activation of the DNA damage response and PARP cleavage induced by curcumin. Our results indicate that activation of the DNA damage response by polyphenols might suppress the malignant transformation of prostate cancer. In addition, testosterone, when combined with curcumin, may have suppressive effects on the progression of prostate cancer.