Metallothioneins (MTs), low molecular mass cysteine-rich proteins, which are able to bind up to 20 monovalent and up to 7 divalent heavy metal ions are widely studied due to their functions in detoxification of metals, scavenging free radicals and cells protection against the oxidative stress. It was found that the loss of the protective effects of MT leads to an escalation of pathogenic processes and carcinogenesis. The most extensive area is MTs expression for oncological applications, where the information about gene patterns is helpful for the identification biological function, resistance to drugs and creating the correct chemotherapy. In other medical applications the effect of oxidative stress to cell lines exposed to heavy metals and hydrogen peroxide is studied as well as influence of drugs and cytokines on MTs expression and MTs expression in the adipose tissue. The precise detection of low metallothionein concentrations and its isoforms is necessary to understand the connection between quantity and isoforms of MTs to size, localization and type of cancer. This information is necessary for well-timed therapy and increase the chance to survival. Microarray chips appear as good possibility for finding all information about expression of MTs genes and isoforms not only in cancer, but also in other diseases, especially diabetes, obesity, cardiovascular diseases, ageing, osteoporosis, psychiatric disorders and as the effects of toxic drugs and pollutants, which is discussed in this review.
- Keywords
- Drug, Gene expression, Metallothionein, Microarray, Resistance, Tumour diseases,
- MeSH
- Humans MeSH
- Metallothionein analysis genetics metabolism MeSH
- Microarray Analysis methods MeSH
- Neoplasms diagnosis genetics metabolism MeSH
- Oxidative Stress physiology MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Review MeSH
- Names of Substances
- Metallothionein MeSH
The combination of microarray technologies with microfluidic sample delivery and real-time detection methods has the capability to simultaneously monitor 10-1000 s of biomolecular interactions in a single experiment. Despite the benefits that microfluidic systems provide, they typically operate in the laminar flow regime under mass transfer limitations, where large analyte depletion layers act as a resistance to analyte capture. By locally stirring the fluid and delivering fresh analyte to the capture spot, the use of passive mixing structures in a microarray environment can reduce the negative effects of these depletion layers and enhance the sensor performance. Despite their large potential, little attention has been given to the integration of these mixing structures in microarray sensing environments. In this study, we use passive mixing structures to enhance the mass transfer of analyte to a capture spot within a microfluidic flow cell. Using numerical methods, different structure shapes and heights were evaluated as means to increase local fluid velocities, and in turn, rates of mass transfer to a capture spot. These results were verified experimentally via the real-time detection of 20-mer ssDNA for an array of microspots. Both numerical and experimental results showed that a passive mixing structure situated directly over the capture spot can significantly enhance the binding rate of analyte to the sensing surface. Moreover, we show that these structures can be used to enhance mass transfer in experiments regarding an array of capture spots. The results of this study can be applied to any experimental system using microfluidic sample delivery methods for microarray detection techniques.
- Keywords
- Biosensors, Mass transfer, Microarrays, Microfluidic mixing, Microfluidics,
- MeSH
- Biosensing Techniques instrumentation MeSH
- Equipment Design MeSH
- Diffusion MeSH
- DNA, Single-Stranded analysis MeSH
- Microarray Analysis instrumentation MeSH
- Microfluidic Analytical Techniques instrumentation MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- DNA, Single-Stranded MeSH
An oligonucleotide microarray for the detection of some fruit-tree viruses was designed and its theoretical detection limit was assessed using Cy3-labelled oligonucleotides. The real sensitivity of the microarray was compared for different kinds of fluorescently labelled targets: (a) cDNA and PCR amplified targets, (b) PCR amplified targets labelled using three different labelling methods. In the first case (a), the number of viral cDNA molecules was below the assessed detection limit of the microarray and only PCR amplified targets were detected. A second comparison (b), done on 3 selected viruses, included indirect labelling, the direct incorporation of labelled-dUTPs, and the use of Cy3-labelled primer. The targets labelled most intensively were produced by the Cy3-primer labelling (2 of 3 viruses) or by the indirect labelling method (1 of 3 viruses), the weakest signal showed targets labelled directly (all 3 viruses). The use of Cy3-primer labelling involved the simplest preparation and the lowest cost, however occasional weak cross-hybridization appeared. The indirect labelling method was of the highest specificity. The probes hybridizing near the 3-end of the targets showed the lowest intensities of fluorescent signal.
- MeSH
- False Positive Reactions MeSH
- Fluorescence MeSH
- Microarray Analysis methods MeSH
- Plant Diseases virology MeSH
- Plant Viruses isolation & purification MeSH
- Oligonucleotide Array Sequence Analysis MeSH
- Sensitivity and Specificity MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Research Support, Non-U.S. Gov't MeSH
The left and right ventricle originate from distinct parts of the cardiac tube, and several genes are known to be differentially expressed in these compartments. The aims of this study were to determine developmental differences in gene expression between the left and right ventricle, and to assess the effect of altered hemodynamic loading. RNA was extracted from isolated left and right normal chick embryonic ventricles at embryonic day 6, 8, and 10, and from day 8 left atrial ligated hearts with hypoplastic left and dilated right ventricles. cRNA was hybridized to Affymetrix Chicken Genome array according to manufacturer protocols. Microarray analysis identified 302 transcripts that were differentially expressed between the left and right ventricle. Comparative analysis detected 91 genes that were different in left ventricles of ligated hearts compared to age-matched ventricles, while 66 were different in the right ones. A large number of the changes could be interpreted as a delay of normal maturation. The approach described in this study could be used as one of the measures to gauge success of surgical procedures for congenital heart disease and help in determining the optimal time frame for intervention to prevent onset of irreversible changes.
- MeSH
- Hemodynamics MeSH
- Chick Embryo MeSH
- Microarray Analysis MeSH
- Myocardium metabolism MeSH
- Heart Ventricles embryology metabolism MeSH
- Heart Atria embryology metabolism MeSH
- Transcriptome MeSH
- Animals MeSH
- Check Tag
- Chick Embryo MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Research Support, N.I.H., Extramural MeSH
BACKGROUND: The full maturational capability of mammalian oocytes is accompanied by nuclear and cytoplasmic modifications, which are associated with proliferation and differentiation of surrounding cumulus cells. These events are regulated on molecular level by the expression of target genes involved in signal transduction pathways crucial for folliculogenesis and oogenesis. Transforming growth factor beta signaling includes several molecules that are involved in the regulation of oogenesis and embryo growth, including bone morphogenetic protein (BMP). However, the BMP-related gene expression profile in oocytes at different maturational stages requires further investigation. METHODS: Oocytes were isolated from pubertal crossbred Landrace gilts follicles, selected with a use of BCB staining test and analyzed before and after in vitro maturation. Gene expression profiles were examined using an Affymetrix microarray approach and validated by RT-qPCR. Database for Annotation, Visualization, and Integrated Discovery (DAVID) software was used for the extraction of the genes belonging to a BMP-signaling pathway ontology group. RESULTS: The assay revealed 12,258 different transcripts in porcine oocytes, among which 379 genes were down-regulated and 40 were up-regulated. The DAVID database indicated a "BMP signaling pathway" ontology group, which was significantly regulated in both groups of oocytes. We discovered five up-regulated genes in oocytes before versus after in vitro maturation (IVM): chordin-like 1 (CHRDL1), follistatin (FST), transforming growth factor-beta receptor-type III (TGFβR3), decapentaplegic homolog 4 (SMAD4), and inhibitor of DNA binding 1 (ID1). CONCLUSIONS: Increased expression of CHRDL1, FST, TGFβR3, SMAD4, and ID1 transcripts before IVM suggested a subordinate role of the BMP signaling pathway in porcine oocyte maturational competence. Conversely, it is postulated that these genes are involved in early stages of folliculogenesis and oogenesis regulation in pigs, since in oocytes before IVM increased expression was observed.
- Keywords
- In vitro maturation, Microarray, Oocytes, Pig,
- MeSH
- In Vitro Oocyte Maturation Techniques * MeSH
- Bone Morphogenetic Proteins genetics metabolism MeSH
- Cumulus Cells cytology metabolism physiology MeSH
- Microarray Analysis MeSH
- Oocytes cytology metabolism physiology MeSH
- Oogenesis genetics MeSH
- Swine genetics metabolism MeSH
- Signal Transduction genetics MeSH
- Gene Expression Profiling MeSH
- Transcriptome MeSH
- Gene Expression Regulation, Developmental MeSH
- Animals MeSH
- Check Tag
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Bone Morphogenetic Proteins MeSH
Microarrays are one of the new emerging methods in plant virology currently being developed by various laboratories. In this study, a new approach is described on the detection of plant viruses using short synthetic single-stranded oligomers (40 nt) instead of PCR products as capture probes. A microchip detecting potato viruses, PVA, PVS, PVM, PVX, PVY and PLRV, in both single and mixed infections was developed and tested. The chip was also designed to distinguish between the main strains of PVY and PVS. Results of initial tests with PVY(NTN) and PVY(O) strains using several different probes for one virus are presented. Possibilities and advantages of the new oligonucleotide-based microarray approach for plant viral diagnosis are discussed.
- MeSH
- Species Specificity MeSH
- Plant Diseases virology MeSH
- Oligonucleotide Probes * MeSH
- Oligonucleotides * chemical synthesis chemistry genetics MeSH
- Plant Viruses classification genetics isolation & purification MeSH
- Oligonucleotide Array Sequence Analysis methods MeSH
- Sensitivity and Specificity MeSH
- Solanum tuberosum virology MeSH
- Publication type
- Journal Article MeSH
- Evaluation Study MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Oligonucleotide Probes * MeSH
- Oligonucleotides * MeSH
DNA microarrays (collections of DNA probes arranged on a shared base) have recently enlarged the spectrum of commercially available laboratory-ready kits in molecular biology. They are powerful new tools for the investigation of global changes in gene expression profiles in cells and tissues. Their assembly process is automatized and the DNA microarrays are further miniaturized. The DNA microarrays are used in search for various specific genes (e.g. connected with an infectious agent) or in gene polymorphism and expression analysis. They will be widely used to investigate expression of various genes connected with various diseases in order to find causes of these diseases and to enable their accurate treatment. Since the DNA microarray assembly technology has been based on methods widely used in the semiconductor industry, we can expect a rapid onset of the routine use of this revolutionary device.
- MeSH
- Biotechnology MeSH
- DNA genetics MeSH
- Gene Expression MeSH
- Humans MeSH
- Base Sequence MeSH
- Oligonucleotide Array Sequence Analysis * instrumentation methods trends MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Review MeSH
- Names of Substances
- DNA MeSH
INTRODUCTION: This study aimed to evaluate the occurrence of clinically relevant (sub)microscopic chromosomal aberrations in fetuses with the nuchal translucency (NT) range from 3.0 to 3.4 mm, which would be potentially missed by cfDNA testing. METHODS: A retrospective data analysis of 271 fetuses with NT between 3.0 and 3.4 mm and increased first trimester combined test (CT) risk in five cohorts of pregnant women referred for invasive testing and chromosomal microarray was performed. RESULTS: A chromosomal aberration was identified in 18.8% fetuses (1:5; 51/271). In 15% (41/271) of cases, trisomy 21, 18, or 13 were found. In 0.7% (2/271) of cases, sex chromosome aneuploidy was found. In 1.1% (3/271) of cases, CNV >10 Mb was detected, which would potentially also be detected by genome-wide cfDNA testing. The residual risk for missing a submicroscopic chromosome aberration in the presented cohorts is 1.8% (1:54; 5/271). CONCLUSION: Our results indicate that a significant number of fetuses with increased CT risk and presenting NT of 3.0-3.4 mm carry a clinically relevant chromosomal abnormality other than common trisomy. Invasive testing should be offered, and counseling on NIPT should include the test limitations that may result in NIPT false-negative results in a substantial percentage of fetuses.
- Keywords
- Chromosomal microarray, Increased nuchal translucency, Noninvasive prenatal testing, Prenatal diagnosis,
- MeSH
- Chromosome Aberrations MeSH
- Chromosome Disorders diagnosis genetics MeSH
- Adult MeSH
- Cohort Studies MeSH
- Humans MeSH
- Nuchal Translucency Measurement * MeSH
- Microarray Analysis MeSH
- Noninvasive Prenatal Testing methods MeSH
- Retrospective Studies MeSH
- Pregnancy MeSH
- Cell-Free Nucleic Acids blood genetics MeSH
- Check Tag
- Adult MeSH
- Humans MeSH
- Pregnancy MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Multicenter Study MeSH
- Names of Substances
- Cell-Free Nucleic Acids MeSH
BACKGROUND: In a prospective study, we measured plasma markers of myocardial damage induced by radiofrequency catheter ablation (RFA) with the protein biochip microarray system. METHODS: A total of 32 consecutive patients undergoing RFA for atrioventricular nodal re-entry tachycardia (AVNRT), right atrial flutter (AFL) and atrial fibrillation (AF) were included in the study. Cardiac troponin I (cTnI), creatine kinase isoenzyme MB (CK-MB), heart-type fatty acid binding protein (hFABP) and glycogen phosphorylase BB (GPBB) were measured using biochip array technology at baseline and 24 h after the procedure. RESULTS: Values for all markers increased 24 h after RFA (cTnI: 0.92+/-0.49 microg/L vs. 0.33+/-0.06 microg/L, p<0.001; CK-MB: 3.79+/-2.04 microg/L vs. 1.85+/-0.55 microg/L, p<0.001; hFABP: 2.82+/-0.95 microg/L vs. 2.00+/-0.95 microg/L, p<0.001; GPBB: 9.07+/-5.83 microg/L vs. 4.70+/-2.50 microg/L, p<0.001). The correlations between plasma marker levels and RFA time were cTnI: r=0.63, p<0.01; CK-MB: r=0.75, p<0.01; hFABP: r=0.55, p<0.05, GPBB: r=0.51, p<0.05; the correlation between RFA time and number of RF applications was significant (r=0.81, p<0.001). Patients with RFA due to AF or flutter had elevated cTnI, CK-MB and hFABP levels compared to patients with AVNRT (cTnI: 1.14+/- 0.49 microg/L vs. 0.59+/-0.25 microg/L, p<0.05; CK-MB: 4.46+/- 2.07 microg/L vs. 2.81+/-1.54 mug/L, p<0.05; hFABP: 3.21+/- 0.98 microg/L vs. 2.25+/-0.54 microg/L, p<0.01). CONCLUSIONS: Myocardial injury induced by RFA can be detected by cTnI, CK-MB, hFABP and GPBB. Plasma cTnI, CK-MB and hFABP levels significantly increased in patients with AFL and AF compared to patients with AVNRT. The increase of cTnI, CK-MB and GPBB levels correlates with the total duration of RFA.
- MeSH
- Biomarkers analysis blood MeSH
- Adult MeSH
- Myocardial Infarction diagnosis etiology MeSH
- Catheter Ablation adverse effects MeSH
- Middle Aged MeSH
- Humans MeSH
- Microarray Analysis methods MeSH
- Young Adult MeSH
- Prospective Studies MeSH
- Check Tag
- Adult MeSH
- Middle Aged MeSH
- Humans MeSH
- Young Adult MeSH
- Male MeSH
- Female MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Biomarkers MeSH
Wilson disease (WD) is an autosomal recessive inherited disorder of copper metabolism that is caused by mutations in the ATP7B gene. To date, more than 300 mutations have been described in this gene. Molecular diagnostics of WD utilizes restriction enzyme digestion, multiplex ligation-dependent probe amplification or a direct sequencing of the whole gene. To simplify and speed up the screening of ATP7B mutations, we have developed a genotyping microarray for the simultaneous detection of 87 mutations and 17 polymorphisms in the ATP7B gene based on the arrayed primer extension reaction. The patient's DNA is amplified in four multiplex polymerase chain reactions, fragmented products are annealed to arrayed primers spotted on a chip, which enables DNA polymerase extension reactions with fluorescently labeled dideoxynucleotides. The Wilson microarray was validated by screening 97 previously genetically confirmed WD patients. In total, we detected 43 mutations and 15 polymorphisms that represent a majority of the common mutations occurring in the Czech and Slovak populations. All screened sequence variants were detected with 100% accuracy. The Wilson chip appears to be a rapid, sensitive and cost-effective tool, representing the prototype of a disease chip that facilitates and speeds up the screening of potential WD patients.
- MeSH
- Adenosine Triphosphatases genetics MeSH
- Copper-Transporting ATPases MeSH
- Point Mutation * MeSH
- Genetic Carrier Screening methods MeSH
- Genotype MeSH
- Hepatolenticular Degeneration diagnosis genetics MeSH
- Heterozygote MeSH
- Humans MeSH
- Microarray Analysis instrumentation methods MeSH
- Mutation MeSH
- DNA Mutational Analysis MeSH
- Cation Transport Proteins genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Adenosine Triphosphatases MeSH
- ATP7A protein, human MeSH Browser
- Copper-Transporting ATPases MeSH
- Cation Transport Proteins MeSH
