硒化壳聚糖对K 562和K 562/ADM作用的比较研究

应用MTT法和细胞集落形成率法观察了硒化壳聚糖对K562和K562／ADM肿瘤细胞株生长的影响，结果发现硒化壳聚糖可有效地抑制两种K562细胞生长，呈量效关系。同等剂量的硒化壳聚糖对K562细胞的作用强度高于对K562／ADM细胞。硒化壳聚糖对耐药的细胞株可产生抑制作用，若与化疗药合用则可减慢甚至部分逆转临床上K562细胞耐药性的发生。     

Sensing lymphoma cells based on a cell-penetrating/apoptosis-inducing/electron-transfer peptide probe

To electrochemically sense lymphoma cells (U937), we fabricated a multifunctional peptide probe that consists of cell-penetrating/apoptosis-inducing/electron-transfer peptides. Electron-transfer peptides derive from cysteine residue combined with the C-terminals of four tyrosine residues (Y₄). A peptide whereby Y₄C is bound to the C-terminals of protegrin 1 (RGGRLCYCRRRFCVCVGR-NH₂) is known to be an apoptosis-inducing agent against U937 cells, and is referred to as a peptide-1 probe. An oxidation response of the peptide-1 probe has been observed due to a phenolic hydroxyl group, and this response is decreased by the uptake of the peptide probe into the cells. To improve the cell membrane permeability against U937 cells, the RGGR at the N-terminals of the peptide-1 probe was replaced by RRRR (peptide-2 probe). In contrast, RNRCKGTDVQAWY₄C (peptide-3 probe), which recognizes ovalbumin, was constructed as a control. Compared with the other probes, the change in the peak current of the peptide-2 probe was the greatest at low concentrations and occurred in a short amount of time. Therefore, the cell membrane permeability of the peptide-2 probe was increased based on the arginine residues and the apoptosis-inducing peptides. The peak current was linear and ranged from 100 to 1000 cells/ml. The relative standard deviation of 600 cells/ml was 5.0% (n = 5). Furthermore, the membrane permeability of the peptide probes was confirmed using fluorescent dye.     

Electrochemical DNA sensing based on gold nanoparticle amplification

A hybridization signal-amplified method based on a gold nanoparticle-supported DNA sequence for electrochemical DNA sensing has been investigated by cyclic voltammetry, differential-pulse voltammetry, and atomic-force microscopy (AFM). Quantitative analysis showed that the peak current increment (Ip) is linearly dependant on the concentration of the gold nanoparticle-supported DNA sequence Au2 over the range 0.51–8.58 pmol L^–1. AFM results indicated that the extent of surface hybridization was dependent on the concentration of the gold-nanoparticle-supported DNA sequence. Moreover, a new pair of peaks, which might arise from the special configuration of the gold-nanoparticle-supported DNA sequence, appeared in the cyclic voltammogram after hybridization. Although quite sensitive, this DNA sensing surface was not easily regenerated, so this kind of amplified method was suitable for disposable DNA sensors and chip-based gene diagnosis sensors.     

硒化壳聚糖抗K562细胞的实验研究

用MTT法和AO/EB荧光染色法观察了硒化壳聚糖对K 562肿瘤细胞株生长的影响。结果发现,硒化壳聚糖可有效地抑制K 562细胞生长,并呈量效、时效关系。经硒化壳聚糖作用后的细胞可明显出现核固缩、碎裂等凋亡形态改变。硒化壳聚糖可诱导K 562细胞凋亡,抑制其生长。     

Antitumor Activity and Mechanism of Action of the Antimicrobial Peptide AMP-17 on Human Leukemia K562 Cells

Cancer is one of the most common malignant diseases in the world. Hence, there is an urgent need to search for novel drugs with antitumor activity against cancer cells. AMP-17, a natural antimicrobial peptide derived from Musca domestica, has antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria, and fungi. However, its antitumor activity and potential mechanism of action in cancer cells remain unclear. In this study, we focused on evaluating the in vitro antitumor activity and mechanism of AMP-17 on leukemic K562 cells. The results showed that AMP-17 exhibited anti-proliferative activity on K562 cells with an IC₅₀ value of 58.91 ± 3.57 μg/mL. The membrane integrity of K562 was disrupted and membrane permeability was increased after AMP-17 action. Further observation using SEM and TEM images showed that the cell structure of AMP-17-treated cells was disrupted, with depressions and pore-like breaks on the cell surface, and vacuolated vesicles in the cytoplasm. Furthermore, further mechanistic studies indicated that AMP-17 induced excessive production of reactive oxygen species and calcium ions release in K562 cells, which led to disturbance of mitochondrial membrane potential and blocked ATP synthesis, followed by activation of Caspase-3 to induce apoptosis. In conclusion, these results suggest that the antitumor activity of AMP-17 may be achieved by disrupting cell structure and inducing apoptosis. Therefore, AMP-17 is expected to be a novel potential agent candidate for leukemia treatment.     

DNA detection based on Mn-doped ZnS quantum dots/methylene blue nanohybrids

Nanohybrids were formed from 3-mercaptopropionic acid(MPA)-coated Mn-doped ZnS quantum dots(QDs) and methylene blue(MB) via electrostatic interaction, and then used in the detection of trace DNA.The principle of detection is as follows: MB binds with Mn-doped ZnS QDs via electrostatic interaction,and then quenches the room temperature phosphorescence(RTP) of the QDs through photoinduced electron-transfer(PIET). After the addition of DNA, MB binds with DNA through intercalation and electrostatic interaction, and desorbs from the surfaces of Mn-doped ZnS QDs, which recovers the RTP of the QDs. On this basis, a DNA detection method based on the properties of RTP was set up. This method shows a detection range of 0.2–20 mg/L, and a detection limit of 0.113 mg/L. Since this method is based on the RTP of QDs, it is not interfered by the background fluorescence or scattering light in vivo, and thus,avoids complex sample pretreatment. Thus, this method is very feasible for detection of trace DNA in biofluids.     

Identification of methylated regions with peak search based on Poisson model from massively parallel methylated DNA immunoprecipitation-sequencing data

DNA methylation is one of the most important epigenetic modification types, which plays a critical role in gene expression. High efficient surveying of whole genome DNA methylation has been aims of many researchers for long. Recently, the rapidly developed massively parallel DNA‐sequencing technologies open the floodgates to vast volumes of sequence data, enabling a paradigm shift in profiling the whole genome methylation. Here, we describe a strategy, combining methylated DNA immunoprecipitation sequencing with peak search to identify methylated regions on a whole‐genome scale. Massively parallel methylated DNA immunoprecipitation sequencing combined with methylation DNA immunoprecipitation was adopted to obtain methylated DNA sequence data from human leukemia cell line K562, and the methylated regions were identified by peak search based on Poison model. From our result, 140 958 non‐overlapping methylated regions have been identified in the whole genome. Also, the credibility of result has been proved by its strong correlation with bisulfite‐sequencing data (Pearson R²=0.92). It suggests that this method provides a reliable and high‐throughput strategy for whole genome methylation identification.     

Design of an electroactive peptide probe for sensing of a protein

We designed a new electroactive peptide probe that has a molecular recognition function for the sensing of a protein. Ovalbumin (OVA) was the model protein, and when RNRCKGTDVQAW interacted with OVA, it conjugated with a tyrosine-rich peptide (Y₄C). This peptide is electroactive, has a high degree of biocompatibility, and offers the possibility of gene expression. To measure the effect of a number of the tyrosine residues, voltammetric measurements were conducted using a series of tyrosine-rich peptides (Y_nC, n = 3–7) with sensitivities that ranged from 10⁻⁹ to 10⁻⁸ M. The electrode response of Y₅C was the maximum value in the series. However, the peak current did not increase when the number of tyrosine residues was increased in a linear fashion. This may have been due to the micelles that are formed by a tyrosine-rich surfactant peptide. Thus, Y₄C was suitable as an electroactive label for the construction of the peptide probe. The electrode response of Y₄CRNRCKGTDVQAW obtained by a glassy carbon electrode was 100-fold that of tyrosine alone. The measurement of OVA via the peptide probe resulted in a detection on the order of 10⁻¹² M. In contrast, the sensitivity of OVA using RCKGTDVQAWY₄C probe was at the 10⁻¹¹ M level, because the hydrophobic moiety gave it a molecular recognition function. The recoveries of the OVA using Y₄CRNRCKGTDVQAW in a solution containing fetal bovine serum ranged between 98 and 101%. Consequently, the combination of a specific peptide and an electroactive element could be a powerful probe for the sensing of proteins.     

Electrochemical sensing of DNA immobilization and hybridization based on carbon nanotubes/nano zinc oxide/chitosan composite film

A novel electrochemical DNA biosensor based on zinc oxide （ZnO） nanoparticles and multi-walled carbon nanotubes （MWNTs） for DNA immobilization and enhanced hybridization detection is presented. The MWNTs/nano ZnO/chitosan composite film modified glassy carbon electrode （MWNTs/ZnO/CHIT/GCE） was fabricated and DNA probes were immobilized on the electrode surface. The hybridization events were monitored by differential pulse voltammetry （DPV） using methylene blue （MB） as an indicator. The sensor can effectively discriminate different DNA sequences related to PAT gene in the transgenic corn, with a detection limit of 2.8× 10^-12 mol/L of target sequence.     

Electrochemical biosensor for detection of BCR/ABL fusion gene based on hairpin locked nucleic acids probe

This communication reports on a novel biosensor to study the hybridization specificity by using thiolated hairpin locked nucleic acids (LNA) as the capture probe. The LNA probe was immobilized on the gold electrode through sulfur–Au interaction and could selectively hybridize with its target DNA. Differential pulse voltammetry (DPV) was used to monitor the hybridization reaction on the probe electrode. The decrease of the peak current of methylene blue, an electroactive indicator, was observed upon hybridization of the probe with the target DNA. The results indicated this new method has excellent specificity for single-base mismatch and complementary after hybridization, and a high sensitivity. This LNA probe has been used for assay of fusion gene in Chronic Myelogenous Leukemia (CML) of the real sample with satisfactory result.     

Ethyl lucidenates A reverses P-glycoprotein mediated vincristine resistance in K562/A02 cells

Multidrug resistance is a major unresolved obstacle to successful cancer chemotherapy. It is often associated with an elevated efflux of a variety of anticancer drugs by ATP-binding cassette transporters including P-glycoprotein, BCRP and MRP1. In this study, the reversal effect of Ethyl lucidenates A on K562/A02 cells was investigated. At concentrations of 10 μM, Ethyl lucidenates A could reverse the resistance of K562/A02 to vincristine up to 7.59 folds. Mechanistically, Ethyl lucidenates A could increase the intracellular accumulation of vincristine in K562/A02 cells through inhibiting the P-glycoprotein mediated drug-transport activity by rhodamine accumulation assay and cell cycle analysis. Further mechanistic investigation found that Ethyl lucidenates A did not alter P-glycoprotein expression. In conclusion, Ethyl lucidenates A could reverse the multidrug resistance of K562/A02 cells via its influence on P-glycoprotein drug-transport activity and thus, be a potential multidrug resistance reversal agent.     

Chemical constituents from Cicuta virosa Linnaeus and their reversal effects on doxorubicin-resistant human myelogenous leukemia (K562/A02) cells

Two new compounds,11,11 '-dimer of scopoletin(1) and 11-O-β-glucopyranosylhamaudol(2),together with seven known compounds were isolated and identified from the whole grass of Cicuta virosa.The chemical structures of the isolated compounds were elucidated using different spectroscopic methods.In addition,the chemical constituents were evaluated for multidrug resistance reversing activity towards doxorubicin-resistant K562/A02 cells.Compounds 1,8,and 9 were endowed with remarkable MDR reversing effects.     

Furocoumarins photolysis products induce differentiation of human erythroid cells

Psoralens, also known as furocoumarins, are a well-known class of photosensitizers largely used in the therapy of various skin disease. In this study we have evaluated the effects of crude pre-irradiated solutions of furocoumarins derivatives on (a) erythroid differentiation and apoptosis of human leukemic K562 cells and (b) hemoglobin synthesis in cultures of human erythroid progenitors derived from the peripheral blood. To prove the activity of a mixture of photoproducts generated by UVA irradiation of the three psoralen derivatives 5-methoxypsoralen (5-MOP) 8-methoxypsoralen (8-MOP), and angelicin (ANG), we employed the human leukemic K562 cell line and the two-phase liquid culture procedure for growing erythroid progenitors. The results obtained demonstrate that pre-irradiated solutions of psoralen derivatives significantly induce erythroid differentiation of K562 cells irrespective of the type of derivative used, suggesting that the active photoproduct(s) share a common structure. Interestingly, solutions of psoralens irradiated in anaerobic conditions do not exhibits erythroid inducing ability, indicating that the effect is mostly due to photooxidized psoralen products. In erythroid precursor cells, psoralens photolysis products stimulates at low concentrations an increase of hemoglobin A and hemoglobin F. Altogether, these data suggest that photoproducts of psoralen warrant further evaluation as potential therapeutic drugs in beta-thalassaemia and sickle cell anaemia.     

Electrochemical biosensor based on nanogold-modified poly-eriochrome black T film for BCR/ABL fusion gene assay by using hairpin LNA probe

A novel electrochemical biosensor is described for detection of breakpoint cluster region gene and a cellular abl (BCR/ABL) fusion gene in chronic myelogenous leukemia (CML) by using thiolated-hairpin locked nucleic acids (LNA) as the capture probe. The hairpin LNA probe was immobilized on the nanogold (NG)/poly-eriochrome black T (EBT) film-modified glassy carbon electrode (GCE). The immobilized LNA probe could selectively hybridize with its target DNA on LNA/NG/EBT/GCE surface. The immobilization and hybridization of the LNA probe were characterized with cyclic voltammetry and electrochemical impedance spectroscopy. The hybridization of the immobilized LNA probe with the target DNA was detected by differential pulse voltammetry with the electroactive methylene blue as an indicator. The results indicated this new method has excellent specificity for single-base mismatch and complementary after hybridization, and a high sensitivity. This novel electrochemical biosensor has been used for assay of PCR real sample with satisfactory result.     

Electrochemical sensing of gases based on liquid collection interfaces

Although many electrochemical gas sensors have been reported, electrochemical gas sensors based on liquid collection constitute a smaller subset. Minimally, a liquid interface based electrochemical gas sensor is composed of two electrodes and an ion conducting electrolyte. There is a large number of possible arrangements of these parts, and many choices exist for their composition and preparation methods. This results in a diverse and rich technology now available for gas sensing. The measurement of some analyte gases of interest, notably ozone, nitrogen oxides, hydrogen peroxide, formaldehyde, ammonia, sulfur dioxide and hydrogen sulfide are specifically discussed. Finally, the recent reviews that are likely to be the most relevant to the further development of electrochemical detection approaches for gases with a liquid collection interface are cited and discussed.     

Gene sensors based on peptide nucleic acid (PNA) probes: relationship between sensor sensitivity and probe/target duplex stability

Gene sensors based on peptide nucleic acid (PNA) probes were prepared and the relationship between sensor sensitivity and the duplex stability of the probe PNAs and target complementary DNAs was studied using five synthesized PNAs (10-, 15-, 17-, 20-, and 22-mers). It was found that the association constants for the probe PNA/target DNA pairs depend not only on the length but also on the base pair sequence, and that the trend in the sensor responses was the same as that in the association constants for the corresponding pairs. In addition, by using two kinds of probe PNAs with different lengths, it was demonstrated that fabrication of sensors based on probe PNAs with comparable association constants yielded similar response curves and sensor sensitivities.     

Electrochemical gene sensor based on a glassy carbon electrode modified with hemin-functionalized reduced graphene oxide and gold nanoparticle-immobilized probe DNA

Microchimica Acta - The authors describe an electrochemical DNA biosensor based on a glassy carbon electrode modified with gold nanoparticles (AuNPs) and reduced graphene oxide that was...     

The effect of irregularity on the dielectric dispersion characteristics of spherical cellular suspension

The dielectric dispersion characteristics of cellular suspensions are fundamentally determined based on the analogy to composite dielectric materials when periodically and discrete arrangement of cells is assumed. However, under native physiological conditions, when flocculation and clamping events usually occur, those assumptions are usually not valid. In the framework of this study, an examination of irregularity effect on the dispersion characteristics of spherical cellular suspensions is presented. Here, the permittivity spectra of the suspensions have been determined by both measurements of living K562 cell suspensions and finite numerical simulations. Based on the measured and simulated spectra, the dispersion characteristics of the suspensions, for several destinies and arrangements of cells, have been quantitatively analyzed using the Havriliak–Negami empirical formula. Generally, a strong correlation between the low dispersion characteristics was observed as the concentration and density of the cells was increased. In addition, all characteristics found to be significantly deviated in comparison to the characteristics of a periodically arrayed suspension. However, when low-dense arrangement was assumed, the correlation found to be much lower when all characteristics found to be less perturbated. Based on a simple model of interacting cells, it is suggested that those deviations are related to intercellular interactions between adjacent cells.     

Mechanism of mercury detection based on interaction of single-strand DNA and hybridized DNA with gold nanoparticles

Mechanisms of interaction of single-strand DNA and hybridized DNA on gold nanoparticles in the presence of Hg²⁺ was studied in this work. Recently the detection of Hg²⁺ using unmodified gold nanoparticles (AuNPs) combined with DNA is becoming a promising technique with the advantages of simplicity, cost-effectiveness and high sensitivity. However, few studies focused on the interaction of ssDNA and hybridized DNA on AuNPs to date. In the present work, we compared the interactions of different DNA probes on AuNPs using both absorption and fluorescence detection. It was found that there were only small partial dsDNA dissociated from the surface of AuNPs after hybridization in the presence of Hg²⁺. Moreover, we found that the aggregated AuNPs/DNA system tended to be dispersed again with increasing Hg²⁺ concentration up to 250 μM. Based on these results, the mechanisms of mercury detection based on interaction between DNA-conjugated gold nanoparticles were investigated. Positively charged dsDNA could bind to the surface of AuNPs and dominate the electrostatic interactions and consequently aggregation of the AuNPs/DNA system.