The biological effects of terahertz wave radiation-induced injury on neural stem cells.

Terahertz (THz) is an electromagnetic wave with a radiation wavelength range of 30-3000 µm and a frequency of 0.1-10 THz. With the development of new THz sources and devices, THz has been widely applied in various fields. However, there are few studies on biological effects of THz irradiation on the human neural stem cells (hNSCs) and mouse neural stem cells (mNSCs), which need to be further studied. We studied the biological effects of THz radiation on hNSCs and mNSCs. The effects of THz irradiation time and average output power on the proliferation, apoptosis, and DNA damage of NSCs were analyzed by flow cytometry and immunofluorescence. The results showed that the proliferation and apoptosis of NSCs were dose-dependently affected by THz irradiation time and average output power. The proliferation of hNSCs was more vulnerable to damage and apoptosis was more serious under the same terahertz irradiation conditions compared to those of mNSCs.

Enhanced Electrochemiluminescence of A Macrocyclic Tetradentate Chelate Pt(II) Molecule through Its Collisional Interactions with the Electrode.

A macrocyclic tetradentate chelate Pt(II) molecule (Pt1) served as an excellent luminophore in electrochemiluminescence (ECL) processes. The blue ECL of Pt1/S2 O8 2- coreactant system in N,N'-dimethylformamide was found to be 46 times higher than that of the Ru(bpy)2+ /S2 O8 2- system or 30 times higher than that of the 9,10-diphenylanthracene/S2 O8 2- system. The unprecedented high ECL quantum efficiencies were caused by the cyclic generation of monomer excited states through collisional interactions of Pt1 molecules with the electrode at an elevated frequency. The ECL is tunable from bright blue to pure white by simply changing the solvent from N,N'-dimethylformamide to dichloromethane. The white ECL of Pt(II) molecule was reported for the first time and the mechanism was proposed to be the simultaneous emissions from the monomer excited state (blue) and excimer (red).

Analytical and clinical evaluation of the light-initiated chemiluminescent assays for measurement of human thyroid hormones.

BACKGROUND: Light-initiated chemiluminescent assay (LiCA) is a new homogeneous immunoassay. The aim of this study was to evaluate the analytical and clinical performance of the assays for the detection of thyroid hormones based on the fully automated LiCA 800 analyzer. METHODS: Analytical validations of the LiCA thyroid assays (TSH, FT3, FT4, T3, and T4) included precision, linearity, analytical sensitivity, interference, and method comparison applying the protocols of the Clinical and Laboratory Standards Institute (CLSI). The diagnostic performance was assessed by the receiver operating characteristic (ROC) curve analysis with different assay schemes for the diagnosis of hyperthyroidism and hypothyroidism. RESULTS: Within-run and within-lab precisions (%CV) of the five assays ranged from 1.06 to 6.40% at all concentrations evaluated. A satisfactory linearity was verified over the entire measuring range for TSH, T3, and T4 (R > 0.99, change in recovery <10%, p = 0.000 all). Paired-comparison measurements presented a comparable assay for each of the five assays (R > 0.96, median bias <5%, p < 0.0001 all) between LiCA and Cobas across three institutes. The diagnostic accuracy of the LiCA assays for hyperthyroidism or hypothyroidism was quantified by the areas under curves (AUC) as 0.925 or 0.832 with the five-assay panel (TSH, FT3, FT4, T3, and T4) and as 0.921 or 0.811 with the three-assay panel (TSH, FT3, and FT4), respectively. No significant difference was found between the AUC of LiCA and that of DxI, Cobas, or Centaur (p > 0.3 all). CONCLUSION: LiCA 800 provides a precise and high-throughput immunoassay platform for detection of thyroid hormones. It is acceptable for clinical use.

PAMAM/polyhedral nanogold-modified probes with DNAase catalysis for the amperometric electrochemical detection of metastasis-associated lung adenocarcinoma transcript 1.

ABSTRACT: Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), a long non coding RNA (lncRNA) present in serum, is an important biomarker for detecting hepatocellular carcinoma (HCC). However, there are some shortcomings in current detection methods. So developing other novel MALAT1 detection methods is necessary. Electrochemical biosensors using different types of nanomaterials with various advantages may provide a suitable method for detection. Here, a new strategy for MALAT1 detection was proposed based on polyhedral nanogold-polyamide-amine dendrimers (PNG-PAMAMs). The SWCNH/Au composite was used as a capture probe immobilization matrix, and PNG-PAMAM was used as a trace label for the detection probe (DP). The strategy takes advantage of the ability of the surface of PNG to bind a capture probe whose sequence contains (GGG)3 trimer that can bind DNAzyme hemin. Moreover, PNG may carry abundant horseradish peroxidases (HRPs) to block excess nonspecific adsorption sites, with synergistic hemin catalysis. The results show that the biosensor provides ultrasensitive detection of MALAT1 with a remarkable catalytic effect. The enhanced biosensor has a detection limit of 0.22 fmol·mL- 1 for MALAT1, and the linear calibration of the biosensor ranged from 1 fmol·mL- 1 to 100 pmol·mL- 1. In addition, the electrochemical biosensor has desirable qualities compared to other detectors; for instance, it is inexpensive, highly stable, and sensitive and has good reproducibility. This assay was also successfully applied to the detection of MALAT1 in serum samples, demonstrating that the technology has potential application in the detection of MALAT1 for clinical HCC diagnosis. GRAPHICAL ABSTRACT: The schematic presentation ilustrates MALATI detection by biosensor with differential pulse stripping voltammetry. Polyhedral nanogold-PAMAM/horseradish peroxidases (PNG-PAMAM/HRP) detection probe with DNAzyme (hemin) sites was applied to determine MALATI. Signal was amplified by hemin/HRP/H2O2 catalytic system.

Regulation of Embryonic Stem Cell Self-Renewal and Differentiation by MicroRNAs.

MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression. They play an important role in various cellular processes such as apoptosis, differentiation, secretion, and proliferation. Embryonic stem cells (ESCs) are derived from the inner cell mass of the blastocyst stage of the embryo. miRNAs are critical factors for the self-renewal and differentiation of ESCs. In this review, we will focus on the role of miRNAs in the self-renewal and directional differentiation of ESCs. We will present the current knowledge on key points related to miRNA biogenesis and their function in ESCs.

Common functional polymorphism within miR-146a and miR-196a-2 as susceptibility loci for hepatocellular carcinoma: An updated meta-analysis.

BACKGROUND: Mutations or single nucleotide polymorphisms (SNPs) within the gene region of microRNAs play an important role for the development of hepatocellular carcinoma (HCC). Extensive studies have tried to investigate the susceptibility role of miR-146a rs2819164 and miR-196a-2 rs11614913. However, these results are still inconsistent and inconclusive. We undertook a meta-analysis containing primarily Asian studies to assess the associations of the two SNPs with HCC risk. METHODS: 19 studies including miR-146a (7170 cases and 9443 controls) and 15 studies including miR-196a-2 (6417 cases and 7627 controls) were used for meta-analysis. Odds ratios and 95% CI were calculated to assess the association in five different genetic models. RESULTS: For the rs2910164 polymorphism of miR-146a, significantly increased risks for HCC were observed when all studies were pooled under two models (CG vs CC: OR = 1.11, 95% CI = 1.02-1.21, P = 0.021; GG + CG vs CC: OR = 1.11, 95% CI = 1.01-1.22, P = 0.035). For the rs11614913 polymorphism of miR-196a-2, significant increased risks for HCC development were observed when all studies were pooled under four models (C vs T: OR = 1.14, 95% CI = 1.06-1.23, P = 0.001; CC vs TT: OR = 1.31, 95% CI = 1.12-1.53, P = 0.001; CC + TC vs TT: OR = 1.16, 95% CI = 1.03-1.31, P = 0.018; CC vs TC + TT: OR = 1.14, 95% CI = 1.00-1.30, P = 0.043). CONCLUSION: Our results show that the two common SNPs within the miRNAs were associated with modest increased risk of HCC (OR < 1.6), especially in the Asian population. Larger population-based studies validating these results are needed.

An electrochemical aptasensor for detection of IFN-γ using graphene and a dual signal amplification strategy based on the exonuclease-mediated surface-initiated enzymatic polymerization.

Tuberculosis is one of the major health problems in the world. The cytokine interferon γ (IFN-γ) is associated with the disease-specific immune responses and is used as a tuberculosis diagnosis marker. In this study, a novel electrochemical aptasensor was developed for IFN-γ detection based on the exonuclease-catalyzed target recycling and the TdT-mediated cascade signal amplification. To construct the aptasensor, a previously hybridized double-stranded DNA (capture probe hybridization with a complementary IFN-γ binding aptamer) was immobilized on a gold nanoparticle-graphene (Au-Gra) nanohybrid film-modified electrode. In the presence of IFN-γ, the formation of an aptamer-IFN-γ complex leads to the liberation of the aptamer from the double-stranded DNA (dsDNA). Using exonuclease, the aptamer was selectively digested, and IFN-γ was released for the target recycling. A large amount of single-stranded capture probes formed and led to the hybridization with signal probe-labelled Au@Fe3O4. Then, the labelled signal probe sequences were catalyzed at the 3'-OH group by terminal deoxynucleotidyl transferase (TdT) to form a long single-stranded DNA structure. As a result, the electron mediator hexaammineruthenium(III) chloride ([Ru(NH3)6](3+)) electrostatically adsorbed onto DNA producing a strong electrochemical signal which can be used to quantitatively measure the IFN-γ levels. With the conducting nanomaterial Au-Gra as a substrate and the target recycling-based surface-initiated enzymatic polymerization-mediated signal amplification strategy, the proposed aptasensor displayed a broad linearity with a low detection limit of 0.003 ng mL(-1). Moreover, the resulting aptasensor exhibited good specificity, acceptable reproducibility and stability, which makes this method versatile and suitable for detecting IFN-γ and other biomolecules.

Ultrasensitive strategy based on PtPd nanodendrite/nano-flower-like@GO signal amplification for the detection of long non-coding RNA.

Highly up-regulated in liver cancer (HULC) is a novel promising noninvasive biomarker for hepatocellular carcinoma (HCC), which is a kind of long non-coding RNAs (lncRNAs). But traditional methods limited HULC clinical detection for ownself drawbacks. Development a new HULC detection approach is urgent and necessary. Electrochemical nucleic acid sensor based on different signal amplification strategies with high sensitivity, fast, simple, and convenient, may solve this problem. Herein, we propose a novel strategy based on Pt-Pd bimetallic nanodendrites/nanoflower-like clusters on graphene oxide/Au/horseradish peroxidase (PtPd BND/BNF@GO/Au/HRP) to enhance the catalytic efficiency and sensitivity. And Au particles were simultaneously and separately capped with thionine or detection probe, which increase the binding amount of detection probe and decrease the electronic background. The results indicated that the catalytic effect was noticeably elevated and that the biosensor provides ultrasensitive detection for the lncRNA HULC. The linear calibration of the biosensor ranged from 1.00×10(-3) to 1.00×10(3) pM/mL, and the limit of detection was 0.247 fM/mL. The lncRNA biosensor based on the PtPd BND/BNF@GO/Au/HRP/Au/thionine exhibited acceptable reproducibility and clear selectivity. This strategy may provide a new alternative for clinical HCC diagnosis through the detection of HULC.

Assessment of endogenous reference gene suitability for serum exosomal microRNA expression analysis in liver carcinoma resection studies.

Serum exosomal microRNAs (miRNAs) have received considerable attention as potential biomarkers for tumor diagnosis. Reverse transcription­quantitative polymerase chain reaction (RT­qPCR) is commonly used to detect miRNA expression levels in various types of cancer. One prerequisite for valid RT­qPCR data is the correct normalization of miRNAs to stably expressed endogenous reference genes (RGs). The study of liver carcinoma resection requires the use of reliable RGs in order to assess the expression levels of serum exosomal target miRNAs. However, the assessment of RG suitability for optimum serum exosomal miRNA expression analysis has yet to be investigated. The present study investigated the expression stability of 10 candidate RGs. The candidate genes included eight miRNAs (miR­16, miR­103, miR­191, let­7a, miR­26a, miR­221, miR­181a, and miR­451) and two small RNAs (5S and U6). The stability values of the candidate genes were calculated using the following algorithms: geNorm, NormFinder, BestKeeper, and the comparative ΔCt method. The overall ranking obtained from these analyses revealed that miR­221, let­7a, and miR­26a were appropriate internal RGs for analysis of serum miRNAs in patients with hepatocellular carcinoma. In addition, normalization with miR­221 and let­7a combined, as recommended by geNorm, or with miR­26a, as recommended by NormFinder, increased the accuracy of interpretation of the target miRNA expression levels in hepatopathy studies.

Identification of endogenous controls for analyzing serum exosomal miRNA in patients with hepatitis B or hepatocellular carcinoma.

Serum exosomal microRNAs (miRNAs) have received considerable attention as potential biomarkers for diagnosing cancer. The canonical technique for measuring miRNA transcript levels is reverse transcription quantitative polymerase chain reaction (RT-qPCR). One prerequisite for validating RT-qPCR data is proper normalization with respect to stably expressed endogenous reference genes. However, genes that meet all of the criteria of a control gene for exosomal miRNAs have not yet been identified. To find out the control gene for exosomal miRNAs, we evaluated the expression stability of 11 well-known reference genes in circulating exosomes. In this study, we found that the combination of miR-221, miR-191, let-7a, miR-181a, and miR-26a can be an optimal gene reference set for normalizing the expression of liver-specific miRNAs. This combination enhanced the robustness of the relative quantification analyses. These findings highlight the importance of validating reference genes before quantifying target miRNAs. Furthermore, our findings will improve studies that monitor hepatitis progression and will aid in the discovery of noninvasive biomarkers to diagnose early stage HCC.

An electrochemical DNA biosensor for the detection of Mycobacterium tuberculosis, based on signal amplification of graphene and a gold nanoparticle-polyaniline nanocomposite.

Due to its low growth rate and its fastidious nature, Mycobacterium tuberculosis is difficult to identify. Its rapid and sensitive detection is, however, critical for the control of tuberculosis. Molecular biology, and more recently electrochemical technology, have been exploited for the detection of this pathogen. In the present study, a novel DNA biosensor was developed for the highly sensitive detection of the specific DNA insertion sequence IS6110 of M. tuberculosis, using reduced graphene oxide-gold nanoparticles (rGO-AuNPs) as a sensing platform and gold nanoparticles-polyaniline (Au-PANI) as a tracer label for amplification. Reduced graphene oxide, which has a large surface area, provided a biocompatible matrix. Gold nanoparticles were electrodeposited on the surface of the rGO modified electrode, which not only increased immobilisation of the capture probe but also promoted electronic transfer. The Au-PANI nanocomposite exhibited good biocompatibility and excellent electrochemical activity. It was therefore used as a tracer label for electrochemical detection, which provided a simple preparation process for a signal-on DNA biosensor. With the excellent electroactivity of the Au-PANI nanocomposite, the resulting DNA biosensor exhibited high sensitivity for the detection of M. tuberculosis over a broad linear range, between 1.0 × 10(-15) and 1.0 × 10(-9) M. The DNA biosensor showed good stability and high specificity and provides a new strategy for clinical M. tuberculosis diagnostics and probably also for pathogenic bacteria in general.

Procalcitonin sensitive detection based on graphene-gold nanocomposite film sensor platform and single-walled carbon nanohorns/hollow Pt chains complex as signal tags.

Septicemia is a serious disease that requires early diagnosis, and procalcitonin (PCT) serves as a diagnostic biomarker for this disease. Traditional clinical detection (via immune-gold chips) remains difficult and expensive. An electrochemical immunosensor based on new nanomaterials may provide a solving approach. Herein, an ultrasensitive sandwich electrochemical strategy for PCT detection was developed. Firstly, reduced graphene oxide (rGO)-gold (Au) nano-composite film was used as the immunosensor platform to increase the amount of PCT antibody 1(Ab1) immobilized. Next, single-walled carbon nanohorns (SWCNHs)/hollow Pt chains (HPtCs) complex was firstly utilized to label PCT Ab2 as signal tags. For SWCNHs with few side effects, high surface area and HPtCs with higher specific surface, better catalytic activity, complex synthesized from both may provide more advantages. Moreover, to amplify signal, HPtC catalytic activity with H2O2 was enhanced by horseradish peroxidase (HRP) for dual synergy amplification. The whole results demonstrated that the proposed immunosensor exhibited fast operation, high sensitivity, good reproducibility, acceptable stability and ideal selectivity compared with traditional method. The linear calibration of the immunosensor ranged from 1.00 pg/mL to 2.00 × 10(1)ng/mL with a detection limit of 0.43 pg/mL. Analytical application results revealed that the immunosensor matched with the real concentrations of serum samples. Overall this immunosensor may provide a new alternative strategy for PCT detection.

Hypoxia promotes epithelial--mesenchymal transition of hepatocellular carcinoma cells via inducing GLIPR-2 expression.

Glioma pathogenesis related-2 (GLIPR-2) belongs to pathogenesis related-1 (PR-1) family whose function remains unknown. In our previous studies, GLIPR-2 was found to be a novel potent stimulator of epithelial-to-mesenchymal transition (EMT) in renal fibrosis which has been classified as type 2 EMT. However, whether GLIPR-2 could induce type 3 EMT in carcinogenesis needs further investigation. In this study, we showed that GLIPR-2 was expressed in hepatocellular carcinoma (HCC) tissues, hypoxia could upregulate the expression of GLIPR-2 in HepG2 and PLC/PRF/5 cells in vitro, overexpression of this protein promoted migration and invasion via EMT, knockdown of GLIPR-2 attenuated migration and invasion of HepG2 and PLC/PRF/5 cells in hypoxia. Moreover, extracellular signal-regulated kinases 1 and 2 (ERK1/2) are positively regulated by GLIPR-2. Taken together, we provide evidence for a hypoxia/GLIPR-2/EMT/migration and invasion axis in HCC cells and it provides novel insights into the mechanism of migration and invasion of hepatocellular carcinoma cells in hypoxia condition.

Development of a real-time resistance measurement for Vibrio parahaemolyticus detection by the lecithin-dependent hemolysin gene.

The marine bacterium Vibrio parahaemolyticus (V. parahaemolyticus) causes gastroenteritis in humans via the ingestion of raw or undercooked contaminated seafood, and early diagnosis and prompt treatment are important for the prevention of V. parahaemolyticus-related diseases. In this study, a real-time resistance measurement based on loop-mediated isothermal amplification (LAMP), electrochemical ion bonding (Crystal violet and Mg(2+)), real-time monitoring, and derivative analysis was developed. V. parahaemolyticus DNA was first amplified by LAMP, and the products (DNA and pyrophosphate) represented two types of negative ions that could combine with a positive dye (Crystal violet) and positive ions (Mg(2+)) to increase the resistance of the reaction liquid. This resistance was measured in real-time using a specially designed resistance electrode, thus permitting the quantitative detection of V. parahaemolyticus. The results were obtained in 1-2 hours, with a minimum bacterial density of 10 CFU.mL(-1) and high levels of accuracy (97%), sensitivity (96.08%), and specificity (97.96%) when compared to cultivation methods. Therefore, this simple and rapid method has a potential application in the detection of V. parahaemolyticus on a gene chip or in point-of-care testing.

Expression of hPOT1 in HeLa cells and the probability of gene variation of hpot1 Exon14 in endometrial cancer are much higher than in other cancers.

To investigate the expression of hPOT1 in the HeLa cell line and screen point mutations of hpot1 in different tumor tissues a two step osmotic method was used to extract nuclear proteins. EMSA was performed to determine the expression of hPOT1 in the HeLa cell line. PCR was also employed to amplify the exon14 sequence of the hpot1 gene in various of cancer tissues. A SV gel and PCR clean-up system was performed to enrich PCR products. DNAStar was used to analyse the exon14 sequence of the hpot1 gene. hPOT1 was expressed in the HeLa cell line and the signal was gradually enhanced as the amount of extracted nuclear proteins increased. The DNA fragment of exon14 of hpot1 was successfully amplified in the HeLa cell line and all cancer tissues, point mutations being observed in 2 out of 3 cases of endometrial cancer (66.7%) despite the hpot1 sequence being highly conserved. However, the sequence of hpot1 exon14 do not demonstrate point mutations in most cancer tissues. Since hPOT1 was expressed in HeLa cell and the probability of gene point variants was obviously higher in endometrial cancer than other cancers, it may be involved in the pathogenesis of gynecological cancers, especially in cervix and endometrium.

Deletion of yncD gene in Salmonella enterica ssp. enterica serovar Typhi leads to attenuation in mouse model.

TonB-dependent transporters (TBDTs) are bacterial outer membrane proteins that are usually involved in the uptake of certain key nutrients, for example iron. In the genome of Salmonella enterica ssp. enterica serovar Typhi, the yncD gene encodes a putative TBDT and was identified recently as an in vivo-induced antigen. In the present study, a yncD-deleted mutant was constructed to evaluate the role of the yncD gene in virulence. Our results showed that the mutant is attenuated in a mouse model by intraperitoneal injection and its virulence is restored by the transformation of a complement plasmid. The competition experiments showed that the survival ability of the yncD-deleted mutant decreases significantly in vivo. To evaluate its vaccine potential, the yncD-deleted mutant was inoculated intranasally in the mouse model. The findings demonstrated a significant immunoprotection against the lethal wild-type challenge. The regulation analysis showed that yncD gene promoter is upregulated under acidic condition. The present study demonstrates that the yncD gene plays an important role in bacterial survival inside the host and is suitable for the construction of attenuated vaccine strains as a candidate target gene.

Protective effect of atorvastatin on radiation-induced vascular endothelial cell injury in vitro.

Vascular endothelial cells are very sensitive to ionizing radiation, and it is important to develop effective prevent agents and measures in radiation exposure protection. In the present study, the protective effects of atorvastatin on irradiated human umbilical vein endothelial cells (HUVEC) and the possible mechanisms were explored. Cultured HUVEC were treated by atorvastatin at a final concentration of 10 µ mol/ml for 10 minutes, and then irradiated at a dose of 2 Gy or 25 Gy. Twenty-four hours after irradiation, apoptosis of HUVEC was monitored by flow cytometry, and the expression of thrombomodulin (TM) and protein C activation in HUVEC was respectively assessed by flow cytometry and spectrophotometry. After treatment with atorvastatin for 24 h, the rate of cell apoptosis decreased by 6% and 16% in cells irradiated with 2 Gy and 25 Gy, respectively. TM expression increased by 77%, 59%, and 61% in untreated cells, 2 Gy irradiation-treated cells, and 25 Gy irradiation-treated cells, respectively. The protein C levels in 2 Gy and 25 Gy irradiation-treated cells were reduced by 23% and 34% when compared with untreated cells, but up-regulated by 79% and 76% when compared with cells which were irradiated and treated with atorvastatin. In conclusion, these data indicate that atorvastatin exerts protective effects on irradiated HUVEC by reducing apoptosis by up-regulating TM expression and enhancing protein C activation in irradiated HUVEC.

A novel, label-free immunosensor for the detection of alpha-fetoprotein using functionalised gold nanoparticles.

BACKGROUND AND OBJECTIVES: Novel immunoassay methods based on electrochemical sensors have been developed, but most of these immunosensors are unsuitable for clinical detection because their preparation requires complicated chemical procedures and because their detection sensitivity is restricted. In order to develop a highly sensitive, label-free amperometric sensor for immunoassays, we synthesised novel, functionalized gold nanoparticles (SV-GNP) by covalently capping the surface of gold nanoparticles (GNP) with 1,1'-bis-(2-mercapto)-4,4'-bipyridinium dibromide, a kind of sulfhyrdryl viologen (SV). DESIGN AND METHODS: We fabricated an immunosensor in a multi-step fashion, by first coating the SV-GNP onto a glassy carbon electrode surface; the resulting electrode core could then adsorb a suitable antibody in a second step to afford the desired immunosensor. alpha-fetoprotein (AFP) was used as a model analyte in this work. RESULTS: The anti-AFP/SV-GNP-modified electrode was sensitive to AFP with a linear relationship between 1.25 and 200 ng/mL and a correlation coefficient of 0.9983; the detection limit at a signal to noise ratio of 3 was 0.23 ng/mL under optimal conditions. In addition, the proposed immunosensor exhibited good sensitivity, selectivity, stability and long-term maintenance of bioactivity. CONCLUSION: The described immunosensor preparation and immunoassay methods offer promise for label-free, simple, and cost-effective analysis of biological samples.