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Imputation is a method that supplies missing information about genetic variants that could not be directly genotyped with DNA microarrays or low-coverage sequencing. Imputation plays a critical role in genome-wide association studies (GWAS). It leads to a significant increase in the number of studied variants, which improves the resolution of the method and enhances the comparability of data obtained in different cohorts and/or by using different technologies, which is important for conducting meta-analyses. When performing imputation, genotype information from the study sample, in which only part of the genetic variants are known, is complemented using the standard (reference) sample, which has more complete genotype data (most often the results of whole-genome sequencing). Imputation has become an integral part of human genomic research due to the benefits it provides and the increasing availability of imputation tools and reference sample data. This review focuses on imputation in human genomic research. The first section of the review provides a description of technologies for obtaining information about human genotypes and characteristics of these types of data. The second section describes the imputation methodology, lists the stages of its implementation and the corresponding programs, provides a description of the most popular reference panels and methods for assessing the quality of imputation. The review concludes with examples of the use of imputation in genomic studies of samples from Russia. This review shows the importance of imputation, provides information on how to carry it out, and systematizes the results of its application using Russian samples.

Staphylococcus aureus CC239-MRSA-III is an ancient pandemic strain of hospital-associated, methicillin-resistant S. aureus that spread globally for decades and that still can be found in some parts of the world. In Kuwait, microarray-based surveillance identified from 2019 to 2022 a series of isolates of a hitherto unknown variant of this strain that carried a second set of recombinase genes, ccrA/B-2. To elucidate the structure of its SCCmec element, two isolates were subjected to nanopore sequencing. This revealed, in addition to ccrA/B-2, several SCC-associated genes including speG (spermidine N acetyltransferase) and a gene encoding a large "E-domain containing protein" (dubbed as edcP-SCC). This gene contained three regions consisting of multiple repeating units. In terms of sequence and structure it was similar but not identical to the biofilm-related aap gene from S. epidermidis. A review of published sequences identified edcP-SCC in eighteen genome sequences of S. aureus, S. epidermidis and S. capitis, and frequently it appears in a similar cluster of genes as in the strains sequenced herein. Isolates also carried a prophage with the adhesion factor sasX/sesI and aminoglycoside resistance genes. This is consistent with an affiliation to the "South-East Asian" Clade of CC239. The emergence of edcP-SCC and sasX-positive CC239 strain shows that, against a global trend towards community-associated MRSA, the ancient pandemic CC239 hospital strain still continues to evolve and to cause outbreaks. [ABSTRACT FROM AUTHOR]

Objectives: The objective of the present study was to examine the diversity of Staphylococcus aureus from mastitis milk samples of cows in Rwanda. Methods: A total of 1080 quarter milk samples from 279 dairy cows were collected in 80 different farms from all five provinces of Rwanda. In total, 135 S. aureus isolates were obtained and subjected to genotyping (spa typing, DNA microarray, whole-genome sequencing (WGS)), antimicrobial susceptibility testing (AST) and phenotypic profiling by Fourier Transform Infrared (FTIR) spectroscopy (including capsular serotyping). Results: Resistance to penicillin and/or tetracycline was most frequently observed. Ten sequence types (STs) (ST1, ST151, ST152, ST5477, ST700, ST7110, ST7983, ST7984, ST8320, ST97) belonging to seven clonal complexes (CCs) (CC1, CC130, CC152, CC3591, CC3666, CC705, CC97) were detected. The Panton–Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83) and the human and bovine toxic shock syndrome toxin gene tst-1 variants were detected. FTIR-based capsular serotyping showed CC-specific differences. Most CC97 (cap5 allele) isolates were primarily nonencapsulated (82%), whereas isolates of CC3591 and CC3666 (cap8 allele) were mostly encapsulated (86.4% and 57.8%, respectively). Our results underline the widespread global distribution of cattle-adapted CC97. Conclusion: The presence of CC3591 and CC3666 in bovine mastitis suggests an important role in cattle health and dairy production in Rwanda. The results of the present study support the need for a rigorous One-Health Surveillance program of the bovine–human interface.

Background and Objectives: Immunocompromised patients are a high-risk group for developing mycobacterial infections with either pulmonary and/or extrapulmonary diseases. Low-cost/density DNA-microarray is considered an easy and efficient method for the detection of typical and atypical mycobacterial species.Materials and Methods: Thirty immunocompromised patients were recruited to provide their clinical specimens (sputum, serum, urine, and lymph node aspirates). Real-time polymerase chain reaction (PCR) and DNA-microarray techniques were performed and compared to the conventional methods of Ziehl-Neelsen staining and Lowenstein Jensen culturing.Results: Mycobacterium tuberculosis complex was detected in all 30 clinical specimens (100% sensitivity) by real-time PCR and DNA-microarray. Additionally, coinfection with 4 atypical species belonging to nontuberculous mycobacteria was identified in 7 sputum specimens. These atypical mycobacterial species were identified as M. kansasii 10% (n = 3), M. avium complex 6.6% (n = 2), M. gordanae 3.3% (n = 1), and M. peregrinum 3.3% (n = 1).Conclusion: This study documents the presence of certain species of atypical mycobacteria among immunocompromised patients in Egypt. [ABSTRACT FROM AUTHOR]

Epstein—Barr virus (EBV) is one of the etiological agents causing infectious mononucleosis. A severe form of the disease can result in developing serious complications, which risk might also depend on the state of patient’s immune system. To date, no specific tests for assessing a risk of developing severe disease form are available. Our study was aimed at identifying molecular genetic markers of severe EBV-infectious mononucleosis (EBV-IM) in immunocompetent peripheral blood cells. Expression of 483 genes and gene transcripts regulating apoptosis, proliferation and differentiation of immunocompetent cells was measured in the peripheral blood leukocytes from patients with severe and moderate EBV-IM as well as apparently healthy donors. A DNA-microarray designed by us and subsequent data processing were carried out by using custom-made “MiDA” software. To identify markers of a severe form of the pathology, expression of each gene and transcript was compared in EBV-IM patients and apparently healthy donors. For each gene and transcript, the level of expression fold change and significance for binary classification were determined. Genes and transcripts, characterized by the maximum values of two determined parameters while comparing patients with severe infection and healthy donors, as well as patients with severe and moderate EBV-IM forms, were selected as markers of severe EBV-IM. Genes and transcripts with differed expression in patients with moderate EBV-IM and healthy donors, were excluded from the list of markers. In addition, sex- and age-linked markers with differed expression were excluded as well. The markers for severe EBV-IM consisted of apoptosis regulators (BCL2L11, BIRC3 genes and XIAP.NM_001167 transcript) and splicing factors (CELF6 gene and SF1.NM_201995 transcript). Compared with donors and patients with a moderate form of the disease, a decreased expression of BCL2L11, BIRC3 genes, transcripts SF1.NM_201995 and XIAP.NM_001167, as well as increased expression of the CELF6 gene were detected in the blood of patients with severe EBV-IM. The functional role of identified molecular markers suggests that severe EBV-IM is characterized by suppressed mitochondrial and activated TRAF-dependent apoptosis pathways in immunocompetent cells. The expression pattern for select markers is specific for severe EBV-MI, not associated with patient sex and age. Thus, study data may be used to develop specific tools for assessing a risk of developing complications of EBV mononucleosis.

Objective of the study is to design the DNA-microarray for differentiation of Y. pestis strains of the main and non-main subspecies and biovars of the main subspecies. Materials and methods. Efficiency analysis for the devised means was conducted using 62 Y. pestis strains of various subspecies and biovars, isolated in the natural foci of Russia and neighboring countries. Results and conclusions. Selected have been the DNA-targets, probes and primers – calculated. Enhanced is the method of sub-specific and biovar differentiation of Y. pestis strains by means of DNA-microarray. DNA-chip with “Med24”, “glpD(-93)”, and “45” targets allows for prompt differentiation of the strains of the main and non-main subspecies and biovars of the main subspecies based on the presence and absence of fluorescent signal by the specific for the main subspecies and its biovars DNA-targets.

Aims: This study was conducted to determine the occurrence of staphylococci from cows with subclinical mastitis from independent herds in Algeria, and to characterize Staphylococcus aureus isolates. Methods and Results: Quarter milk samples were collected separately, somatic cells were counted and samples with more than 200 000 somatic cells per ml were cultured on blood agar. Staphylococci isolates were identified by routine diagnostics, and S. aureus isolates were tested for antibiotic susceptibility by disk diffusion and microdilution. Congo red agar was used to detect biofilm formation and capsule synthesis was detected on serum soft agar (SSA). The S. aureus isolates were characterized by spa typing. DNA microarray analysis was performed to detect resistance and virulence genes. Overall, 40·0% (167/418) of the cows suffered from mastitis. In 63·5% (106/167) of the cows staphylococci were identified. Nine of the 106 Staphylococcus isolates (8·5%) were S. aureus. The coagulase‐negative staphylococci belonged to 14 species. All S. aureus isolates were multiresistant and biofilm forming, with 66·67% of them showing diffuse colonies on SSA and belonged to CC97‐agrI‐cap5. Biofilm genes (icaA/C/D), 13 genes encoding for adhesion, six genes encoding proteases, 11 genes encoding superantigen like toxins were found. Genes conferring resistance to tetracycline (tet(K)), penicillin (blaZ/I/R) and macrolide–lincosamide–streptogramin B (erm(B), erm(A)) were also detected in the S. aureus from this study. Conclusions: The current investigation provides a detailed molecular and biofilm formation ability of S. aureus involved in subclinical mastitis in Algeria and shows the wide distribution of adhesion and enterotoxin(‐like) genes among S. aureus responsible for causing subclinical bovine mastitis. Significance and Impact of the Study: These findings are valuable in tracking the evolution and genomic variation of S. aureus from bovine origin. [ABSTRACT FROM AUTHOR]

Gene selection (GS) is an important research area in the analysis of DNA-microarray data, since it involves gene discovery meaningful for a particular target annotation or able to discriminate expression profiles of samples coming from different populations. In this context, a wide number of filter methods have been proposed in the literature to identify subsets of relevant genes in accordance with prefixed targets. Despite the fact that there is a wide number of proposals, the complexity imposed by this problem (GS) remains a challenge. Hence, this paper proposes a novel approach for gene selection by using cluster techniques and filter methods on the found groupings to achieve informative gene subsets. As a result of applying our methodology to Colon cancer data, we have identified the best informative gene subset between several one subsets. According to the above, the reached results have proven the reliability of the approach given in this paper.

Abstract To understand the prevalence and characteristics of Staphylococci causing goat mastitis in Algeria, mastitis milk samples were collected and cultured on both blood agar and mannitol salt agar medium. Staphylococci isolates were identified by API-Staph gallery and tested for antibiotic susceptibility by disk diffusion method. DNA microarrays analysis was performed on S. aureus strains in order to detect virulence factors, including toxins, and to assign the isolates to theirs MLST clonal complexes. Overall, 86/266 (32.3%) goats suffered from mastitis, whose 32.5% (28/86) infected with Staphylococci. Most of strains were susceptible to many antibiotics, except for penicillin, tetracycline and fusidic acid, and the resistance incidence of which were 21.4%, 25%, and 10.7% respectively. Only twelve (42.8%) isolates were resistant to utmost two antibiotics and bi-drug resistance was associated to four Coagulase negative Staphylococci (CNS) strains. Nine different species were identified in CNS isolates while S. aureus strains were clustered within CC130- agr 3-[ lukF-P 83, lukM +] (four isolates; 50%), CC6- agr 1 (three isolates; 37.5%) and one agr 4 non-typables cluster. Enterotoxins genes were found in 62.5% of the S. aureus strains. Of which sec, sel were the most prevalent (62.5%: n = 5) followed by sea (12.5%: n = 1). CC130 showed edinB , tst and tetK (2/4) genes and CC6 were positive to cna and fosB genes. The current investigation provide the data for prevalence of Staphylococci in goats in Tizi-Ouzou region and S. aureus characterization which will help in tracking evolution of epidemic strains and their control methods. [ABSTRACT FROM AUTHOR]

Non-steroidal anti-inflammatory drugs (NSAIDs) represent a growing concern for marine ecosystems due to their ubiquitous occurrence and documented adverse effects on non-target organisms. Despite the remarkable efforts to elucidate bioaccumulation and ecotoxicological potential under short-term conditions, limited and fragmentary information is available for chronic exposures. In this study bioavailability, molecular and cellular effects of diclofenac (DIC), ibuprofen (IBU) and ketoprofen (KET) were investigated in mussels Mytilus galloprovincialis exposed to the realistic environmental concentration of 2.5 μg/L for up to 60 days. Results indicated a significant accumulation of DIC and IBU but without a clear time-dependent trend; on the other hand, KET concentrations were always below the detection limit. Analyses of a large panel of molecular, biochemical and cellular biomarkers highlighted that all investigated NSAIDs caused alterations of immunological parameters, genotoxic effects, modulation of lipid metabolism and changes in cellular turn-over. This study provided the evidence of long-term ecotoxicological potential of NSAIDs, further unraveling the possible hazard for wild marine organisms. [ABSTRACT FROM AUTHOR]

Gene selection (or feature selection) from DNA-microarray data can be focused on different techniques, which generally involve statistical tests, data mining and machine learning. In recent years there has been an increasing interest in using hybrid-technique sets to face the problem of meaningful gene selection; nevertheless, this issue remains a challenge. In an effort to address the situation, this paper proposes a novel hybrid framework based on data mining techniques and tuned to select gene subsets, which are meaningfully related to the target disease conducted in DNA-microarray experiments. For this purpose, the framework above deals with approaches such as statistical significance tests, cluster analysis, evolutionary computation, visual analytics and boundary points. The latter is the core technique of our proposal, allowing the framework to define two methods of gene selection. Another novelty of this work is the inclusion of the age of patients as an additional factor in our analysis, which can leading to gaining more insight into the disease. In fact, the results reached in this research have been very promising and have shown their biological validity. Hence, our proposal has resulted in a methodology that can be followed in the gene selection process from DNA-microarray data. [ABSTRACT FROM AUTHOR]

In 2013, an adult red squirrel ( Sciurus vulgaris) was found dead showing, during necropsy, acute diffuse catarrhal enteritis. Using a virus screening pipeline based on a pan-viral DNA microarray with downstream next-generation sequencing a novel squirrel-associated virus was identified with mastadenovirus-typical sequence elements. Phylogenetic analysis of hexon protein amino acid sequences demonstrated the highest similarity to Equine adenovirus (AdV) 2, but a strong divergence to a squirrel AdV from Korea and other rodent AdVs. Shorter hexon gene segment investigations confirmed a close relationship with other squirrel-derived AdVs from Europe. The novel virus species was tentatively designated as Squirrel Adenovirus-1. [ABSTRACT FROM AUTHOR]

Gene selection is a major research area in microarray analysis, which seeks to discover differentially expressed genes for a particular target annotation. Such genes also often called informative genes are able to differentiate tissue samples belonging to different classes of the studied disease. Despite the fact that there is a wide number of proposals, the complexity imposed by this problem remains a challenge today. This research proposes a gene selection approach by means of a clustering-based multi-agent system. This proposal manages different filter methods and gene clustering through coordinated agents to discover informative gene subsets. To assess the reliability of our approach, we have used four important and public gene expression datasets, two Lung cancer datasets, Colon and Leukemia cancer dataset. The achieved results have been validated through cluster validity measures, visual analytics, a classifier and compared with other gene selection methods, proving the reliability of our proposal. [ABSTRACT FROM AUTHOR]

The aim of the present investigation was to provide new insights on accumulation and possible adverse effects of various non-steroidal anti-inflammatory drugs (NSAIDs) in mussels, Mytilus galloprovincialis , exposed to an environmentally realistic concentration (0.5 μg/L) of individual compounds, Acetaminophen (AMP), Diclofenac (DIC), Ibuprofen (IBU), Ketoprofen (KET) or Nimesulide (NIM). The measurement of drugs in mussel tissues was integrated with both functional alterations at cellular level and transcriptomic responses. Results indicated the capability of mussels to accumulate DIC and NIM, while AMP, IBU and KET were always below detection limit. A large panel of ecotoxicological biomarkers revealed the early onset of alterations induced by tested NSAIDs on immunological responses, lipid metabolism and DNA integrity. The gene transcription analysis through DNA microarrays, supported cellular biomarker results, with clear modulation of a large number of genes involved in the arachidonic acid and lipid metabolism, immune responses, cell cycle and DNA repair. The overall results indicated an ecotoxicological concern for pharmaceuticals in M. galloprovincialis , with transcriptional responses appearing as sensitive exposure biomarkers at low levels of exposure: such changes, however, are not always paralleled by corresponding functional effects, suggesting caution when interpreting observed effects in terms of perturbed cellular pathways. Fascinating similarities can also be proposed in the mode of action of NSAIDs between bivalves and vertebrate species. [ABSTRACT FROM AUTHOR]

In this paper, a hybrid algorithm is presented based on five data filtering methods to make a first selection of genes of DNA- microarray. A genetic algorithm combined with a simulated annealing and a SVM-classifier, then a second selection and classification is performed from a genes subset, with the combination of these techniques, we seek a better classification rate obtained with a small genes subset and the results are compared with other methods reported in the literature. [ABSTRACT FROM PUBLISHER]

The increasing incidence of invasive fungal diseases ( IFD), most of all invasive aspergillosis ( IA) in immunocompromised patients emphasises the need to improve the diagnostic tools for detection of fungal pathogens. We investigated the diagnostic performance of a multifungal DNA-microarray detecting 15 different fungi [ Aspergillus, Candida, Fusarium, Mucor, Rhizopus, Scedosporium and Trichosporon species (spp.)] in addition to an Aspergillus specific polymerase chain reaction ( PCR) assay. Biopsies, bronchoalveolar lavage and peripheral blood samples of 133 immunocompromised patients (pts) were investigated by a multifungal DNA-microarray as well as a nested Aspergillus specific PCR assay. Patients had proven ( n = 18), probable ( n = 29), possible ( n = 48) and no IFD ( n = 38) and were mostly under antifungal therapy at the time of sampling. The results were compared to culture, histopathology, imaging and serology, respectively. For the non- Aspergillus IFD the microarray analysis yielded in all samples a sensitivity of 64% and a specificity of 80%. Best results for the detection of all IFD were achieved by combining DNA-microarray and Aspergillus specific PCR in biopsy samples (sensitivity 79%; specificity 71%). The molecular assays in combination identify genomic DNA of fungal pathogens and may improve identification of causative pathogens of IFD and help overcoming the diagnostic uncertainty of culture and/or histopathology findings, even during antifungal therapy. [ABSTRACT FROM AUTHOR]

Background: Clostridium difficile can cause antibiotic-associated diarrhea and a possibility of outbreaks in hospital settings warrants molecular typing. A microarray was designed that included toxin genes (tcdA/B, cdtA/B), genes related to antimicrobial resistance, the slpA gene and additional variable genes. Results: DNA of six reference strains and 234 clinical isolates from South-Western and Eastern Germany was subjected to linear amplification and labeling with dUTP-linked biotin. Amplicons were hybridized to microarrays providing information on the presence of target genes and on their alleles. Tested isolates were assigned to 37 distinct profiles that clustered mainly according to MLST-defined clades. Three additional profiles were predicted from published genome sequences, although they were not found experimentally. Conclusions: The microarray based assay allows rapid and high-throughput genotyping of clinical C. difficile isolates including toxin gene detection and strain assignment. Overall hybridization profiles correlated with MLST-derived clades. [ABSTRACT FROM AUTHOR]

Voltage-dependent sodium channels are membrane proteins essential for cell excitability. They are composed by a pore-forming α-subunit, encoded in mammals by up to nine different genes, and four different ancillary β-subunits. The expression pattern of the α subunit isoforms confers the distinctive functional and pharmacological properties to different excitable tissues. β-Subunits are important modulators of channel function and expression. Mutation C121W of the β1-subunit causes an autosomal dominant epileptic syndrome without cardiac symptoms. In neuroectoderm GH3 and cardiac H9C2 cells, the over-expression of β1 subunit augments α subunit mRNA and protein levels as well as sodium current density. Interestingly, the introduction of the epileptogenic C121W-β1 subunit produces additional changes in the α-subunit expression pattern of H9C2 cells, leaving unaltered the sodium channel isoform composition of GH3 cells. The challenge of the present work was to identify those genes that were differentially expressed in response to WT- or C121W-β1 subunit over-expression in the two rat cell lines under analysis. Hence, we analyzed the total mRNA extracted from control-untransfected and from WT- and C121W-β1-transfected GH3 and H9C2 cells by DNA-microarray. We found that, in agreement with their different embryonal origin, the over-expression of WT- and C121W-β1 subunits modifies the expression of different gene sets in GH3 and H9C2 cells. Focusing on the effects of the C121W mutation, we found that it causes the modification of 214 genes, most of them were down-regulated (202) in GH3 cells; on the contrary, it determined the up-regulation of only five genes in H9C2 cells. Interestingly, most genes modified by the C121W β1 subunit are involved in pivotal processes of the cell such as cellular communication and protein expression. Our results confirm the important role of the sodium channel β1 subunit in the control of NaCh gene expression, and highlight once more the tissue-specific effect of the C121W mutation. [ABSTRACT FROM AUTHOR]

Death receptors (DRs) and the participants of DR-mediated signaling are characterized by a large number of mRNA isoforms generated by alternative splicing. Due to their high labor intensity and high cost, conventional methods (RT-PCR and RT-PCR in real time) are ineffective when the simultaneous detection of a plurality of mRNA isoforms is needed. In this regard, the use of DNA biochips is has prospective applications in analyzing the expression of many genes simultaneously. In this paper, we suggest an optimal strategy of probes selection aimed at detecting the maximum number of mRNA splice variants generated by major participants of DR-signaling. The objects of the study were 185 genes that form 1134 mRNA isoforms. As a result, a biochip design was developed that enables the detection of 499 mRNA isoforms (44% of total mRNA splice variants). The proposed strategy combines a high degree of modularity, the use of modern high-performance computers, and broad opportunities for setting up the selection criteria in accordance with the objectives of the study. [ABSTRACT FROM AUTHOR]

In the present work, an integrated analysis was performed on DNA-microarray data of bovine muscle samples belonging to controls, animals treated with various growth promoters (GPs) and unknown commercial samples. The aim was identify a robust gene expression signature of corticosteroid treatment for the classification of commercial samples, despite the effects of biological variation and other confounding factors. DNA-Microarray data from 5 different batches of bovine skeletal muscle samples were analyzed (146 samples). After preprocessing, expression data from animals treated with corticosteroids and controls from the different batches (89 samples) were used to train a Support Vector Machines (SVMs) classifier. The optimal number of gene probes chosen by our classification framework was 73. The SVMs with linear kernel built on these 73 biomarker genes was predicted to perform on novel samples with a high classification accuracy (Matthew's correlation coefficient equal to 0.77) and an average percentage of false positive and false negative equal to 5% and 6%, respectively. Concluding, a relatively small set of genes was able to discriminate between controls and corticosteroid-treated animals, despite different breeds, animal ages, and combination of GPs. The results are extremely promising, suggesting that integrated analysis provides robust transcriptomic signatures for GP abuse. [ABSTRACT FROM AUTHOR]

Cyanobacterial blooms in aquatic environments are frequently characterized by elevated levels of microcystins, a potent hepatotoxin. Here we exposed the nematode Caenorhabditis elegans with environmentally realistic concentrations of MC-LR to explore its non-hepatic toxicity. Lifespan, reproduction and growth assays confirmed the toxic potential of 100 μg/L MC-LR even in this liver-lacking invertebrate. Whole-genome microarray analysis revealed that a neuromodulating action was the dominant response in nematodes challenged with 100 μg/L MC-LR. Indeed, most of the 201 differentially expressed genes were associated with neurobehavior, neurogenesis, and signaling associated pathways. In addition, a whole-genome miRNA-microarray highlighted that, in particular, members of the let-7 family were differentially regulated. These miRNAs are involved in the developmental timing of cell fates, including neurons, and are probably also part of the stress response system. To conclude, neurological modulation is the main transcriptional stress response in C. elegans exposed to MC-LR. [ABSTRACT FROM AUTHOR]

Summary Objective Human-papillomaviruses (HPV) type 16 is a causative agent in an increasing subset of oropharyngeal squamous cell carcinomas (OPSCCs). These tumors have distinct oncogenic mechanisms and a more favorable prognosis than tobacco-induced OPSCCs. Although these differences emphasize the need for a specific therapeutic approach, HPV status is still not used to guide treatment. A better characterization of the molecular profile related to HPV16-induced OPSCC might help to develop personalized treatments. Patients and methods Using a human whole-genome DNA-microarray, we have examined the gene expression profiles in 15 HPV-negative and 15 transcriptionally-active HPV-positive OPSCCs. The study was conducted in two steps. Firstly, a learning/training-set consisting of 8 HPV16-positive and 8 HPV16-negative OPSCCs was analyzed to identify a specific signature. Potentially confounding factors (stage, sex and tobacco) were equally distributed in both groups. Subsequently the robustness of this signature was confirmed by blind case-by-case classification of a validation-set composed of the 14 remaining tumors. Results We have identified a signature composed of 224 genes, which discriminates HPV16-induced OPSCC from their HPV-negative counterparts. After the blind classification of the 14 tumours, the viral status was revealed: 13 out of 14 tumors were correctly classified according to tumor etiology, 1/14 was not determined and none were misclassified. Several of the differentially expressed genes were involved in cell-cycle regulation, DNA replication and repair, transcription regulation, immune response and apoptosis. Conclusion Our study contributes to a better understanding of pathogenic mechanisms involved in the development of HPV-positive OPSCCs and in the identification of potential therapeutic targets. [ABSTRACT FROM AUTHOR]

The effects of steroid hormone implants containing trenbolone alone (Finaplix-H), combined with 17β- oestradiol (17β-E; Revalor-H), or with 17β-E and dexamethasone (Revalor-H plus dexamethasone per os) on the bovine muscle transcriptome were examined by DNA-microarray. Overall, large sets of genes were shown to be modulated by the different growth promoters (GPs) and the regulated pathways and biological processes were mostly shared among the treatment groups. Using the Prediction Analysis of Microarray program, GP-treated animals were accurately identified by a small number of predictive genes. A meta-analysis approach was also carried out for the Revalor group to potentially increase the robustness of class prediction analysis. After data pre-processing, a high level of accuracy (90%) was obtained in the classification of samples, using 105 predictive gene markers. Transcriptomics could thus help in the identification of indirect biomarkers for anabolic treatment in beef cattle to be applied for the screening of muscle samples collected after slaughtering. [ABSTRACT FROM AUTHOR]

In Drosophila melanogaster differentiation of the male germ cells is accompanied by chromatin rearrangement and activation of the specific genes. These processes are regulated by few transcription factors that belong to two classes, can and aly that form distinct functional complexes. Mechanisms of action of aly and can class transcription factors on gene expression and chromatin state remain unclear. To investigate this question we have built the whole genome binding profile of transcription factor Comr belonging to aly class using the tissue-specific DamID method. Resulting data were correlated with gene expression in comr ( aly class) and can ( can class) mutant testes. It was shown that Comr is a direct activator for about 300 testis-specific genes. Furthermore a set of genes revealed decreased expression in comr mutants but did not bind Comr protein, suggesting the existence of secondary regulation. Indeed, among the Comr gene targets we found a gene coding an uncharacterized transcription factor that could be a secondary participant in the genetic pathway in spermatocytes. These date allowed us to advance a model of gene activation needed for male gametes differentiation in D. melanogaster. [ABSTRACT FROM AUTHOR]

Abstract: Several studies regarding the transcriptome of Mycobacterium tuberculosis following the exposure to various in vitro simulated phagosomal stressors, have already tried to elucidate the bacterium behavior during the intracellular infection. An in vitro acid-nitrosative multi-stress was carried out for M. tuberculosis H37Rv and Mycobacterium smegmatis MC 2 155 in order to analyze by DNA-microarray the gene expression changes associated respectively to pathogenic and non-pathogenic mycobacterial species. During acid-nitrosative multi-stress both mycobacteria shift their transcriptome to allow the anaerobic respiratory state and energy pathways characteristic of starvation. M. tuberculosis counteracts the combined acid-nitrosative stress more efficiently than M. smegmatis as also shown by the up-regulation of glbN and hmp genes, that are specifically directed to NO detoxification. Moreover, the down-regulation of some virulence factors involved in phthiocerol dimycocerosates synthesis strengthens the hypothesis that these major virulence determinants may be attenuated by M. tuberculosis in the presence of reactive nitrogen species. In fact, it down-regulates other genes implicated in the synthesis of membrane structural lipids but in contrast to M. smegmatis, M. tuberculosis up-regulates many genes annotated for the synthesis of peptidoglycan. Results suggest a gene regulation of M. tuberculosis which reveals a distinctive expression pattern under stressful environment. [Copyright &y& Elsevier]

We performed transcriptome analysis of some human induced pluripotent stem cells, embryonic stem cells, and human somatic cells using DNA microarrays. PluriTest bioinformatic system was used for evaluation of cell pluripotency. Changes in the genome structure and status of X-chromosome gene expression was analyzed using microarray technology. [ABSTRACT FROM AUTHOR]

Abstract: It is known that a combined glycine/lysozyme treatment is able to induce in vitro the mycobacterial conversion from the bacillary to the cell wall defective forms. These forms also naturally occur in vivo as a response to various antimicrobial factors such as lysozyme released by phagocytic cells. Although they have been successfully isolated from patients with several chronic diseases, their role in pathogenesis is still unknown, mainly due to the difficulties in handling the in vivo isolated variants. Moreover, nothing is known about the transcriptional peculiarities that may exist in comparison to the vegetative phase. Hence, in this study, we simulated in vitro the induction of the mycobacterial cell wall defective state by using a glycine and lysozyme-based treatment in order to identify the gene expression profiles of both pathogenic and non-pathogenic mycobacteria. DNA-microarray results showed that in contrast to the non-pathogenic Mycobacterium smegmatis species, glycine and lysozyme treated forms of Mycobacterium tuberculosis and Mycobacterium avium subspecies paratuberculosis regulated a repertoire of genes usually expressed in vivo during adaptation and persistence within host environments. Results suggest that the cell wall defective state may represent an important stage in the life-cycle of pathogenic mycobacteria that potentially coordinates persistence. [Copyright &y& Elsevier]

Background: Recent studies have identified in Mycobacterium avium subsp. paratuberculosis (MAP), already known as a pathogen in ruminants, a potential zoonotic agent of some autoimmune diseases in humans. Therefore, considering the possible risk for public health, it is necessary a thorough understanding of MAP's gene expression during infection of human host as well as the identification of its immunogenic and/or virulence factors for the development of appropriate diagnostic and therapeutic tools. Results: In order to characterize MAP's transcriptome during macrophage infection, we analyzed for the first time the whole gene expression of a human derived strain of MAP in simulated intraphagosomal conditions and after intracellular infection of the human macrophage cell line THP-1 by using the DNA-microarray technology. Results showed that MAP shifts its transcriptome to an adaptive metabolism for an anoxic environment and nutrient starvation. It up-regulates several response factors to oxidative stress or intracellular conditions and allows, in terms of transcription, a passive surface peptidoglycan spoliation within the macrophage along with an intensification of the anabolic activity for lipidic membrane structures. Conclusions: These results indicate a possible interactive system between MAP and its host cell based on the internal mimicry unlike other intracellular pathogens, bringing new hypothesis in the virulence and pathogenicity of MAP and its importance in human health. [ABSTRACT FROM AUTHOR]

Background: The use of growth-promoters in beef cattle, despite the EU ban, remains a frequent practice. The use of transcriptomic markers has already proposed to identify indirect evidence of anabolic hormone treatment. So far, such approach has been tested in experimentally treated animals. Here, for the first time commercial samples were analyzed. Results: Quantitative determination of Dexamethasone (DEX) residues in the urine collected at the slaughterhouse was performed by Liquid Chromatography-Mass Spectrometry (LC-MS). DNA-microarray technology was used to obtain transcriptomic profiles of skeletal muscle in commercial samples and negative controls. LC-MS confirmed the presence of low level of DEX residues in the urine of the commercial samples suspect for histological classification. Principal Component Analysis (PCA) on microarray data identified two clusters of samples. One cluster included negative controls and a subset of commercial samples, while a second cluster included part of the specimens collected at the slaughterhouse together with positives for corticosteroid treatment based on thymus histology and LC-MS. Functional analysis of the differentially expressed genes (3961) between the two groups provided further evidence that animals clustering with positive samples might have been treated with corticosteroids. These suspect samples could be reliably classified with a specific classification tool (Prediction Analysis of Microarray) using just two genes. Conclusions: Despite broad variation observed in gene expression profiles, the present study showed that DNA-microarrays can be used to find transcriptomic signatures of putative anabolic treatments and that gene expression markers could represent a useful screening tool. [ABSTRACT FROM AUTHOR]

Background: Renal cell carcinoma (RCC) is characterized by a number of diverse molecular aberrations that differ among individuals. Recent approaches to molecularly classify RCC were based on clinical, pathological as well as on single molecular parameters. As a consequence, gene expression patterns reflecting the sum of genetic aberrations in individual tumors may not have been recognized. In an attempt to uncover such molecular features in RCC, we used a novel, unbiased and integrative approach.Methods: We integrated gene expression data from 97 primary RCC of different pathologic parameters, 15 RCC metastases as well as 34 cancer cell lines for two-way non-supervised hierarchical clustering using gene groups suggested by the PANTHER Classification System. We depicted the genomic landscape of the resulted tumor groups by means of Single Nuclear Polymorphism (SNP) technology. Finally, the achieved results wereimmunohistochemically analyzed using a tissue microarray (TMA) composed of 254 RCC.Results: We found robust, genome wide expression signatures, which split RCC into three distinct molecular sub-groups. These groups remained stable even if randomly selected gene sets were clustered. Notably, the pattern obtained from RCC cell lines was clearly distinguishable from that of primary tumors. SNP array analysis demonstrated differing frequencies of chromosomal copy number alterations among RCC sub-groups. TMA analysis with group-specific markers showed a prognostic significance of the different groups.Conclusion: We propose the existence of characteristic and histologically independent genome-wide expression outputs in RCC with potential biological and clinical relevance. [ABSTRACT FROM AUTHOR]

Abstract: Staphylococcus aureus is a common cause of mastitis and other diseases in camels. In order to obtain data on population structure as well as on the carriage of toxin genes and resistance markers, a collection of 45 isolates from dromedaries of Dubai, United Arab Emirates, were genotyped. These isolates belonged to clonal complexes CC6 (twenty isolates; 44.44%), CC30 (sixteen isolates; 35.56%), CC188 (five isolates; 11.11%), CC152 (1 isolate, 2.2%) and to a previously un-described sequence type (ST1755: arcc-18, aroe-115, glpf-6, gmk-2 pta-109, tpi-50 and yqil-2; three isolates; 6.67%). Resistance genes proved to be rare. Only three out of 45 isolates (6.67%) carried the beta-lactamase operon. The tetracycline resistance gene tetK was also detected in three isolates (6.67%). Neither the mecA gene, defining MRSA, nor other resistance genes were found. Common virulence markers included leukocidin genes lukD + lukE (in twenty-five isolates; 55.56%), the staphylokinase gene sak (twenty-two isolates; 48.89%), the enterotoxin gene cluster egc (fifteen isolates; 33.33%), and a distinct variant of the enterotoxin A gene (sea-320E, GenBank AY196686.1; thirteen isolates; 28.89%). One CC152 isolate was positive for genes encoding the Panton-Valentine leukocidin (lukF-PV + lukS-PV). This study provides first genotyping data on the population structure and the presence of toxin genes and resistance markers of S. aureus strains in Middle Eastern camels. [Copyright &y& Elsevier]

During several decades microscopy and culture based methods have been the most important techniques for the detection of fungal infections. Culture, though often slow, sometimes insensitive and sometimes confusing with respect to contamination or colonization, may yield the specific aetiological agent, and may allow susceptibility testing to be performed. However, molecular detection and identification using PCR for the amplification of fungal DNA from tissue is being applied more and more frequently for the early diagnosis and identification of fungal pathogens. Other tools such as fluorescence in situ hybridization (FISH) or DNA microarrays have also been developed and their performance is currently being evaluated. Since standardization and validation for most of these newer techniques are still lacking the combination of various diagnostic tools is still mandatory to allow earlier diagnosis of systemic fungal infections. [ABSTRACT FROM AUTHOR]

Functions of glycosphingolipids on the eukaryotic cell membranes during the onset of oncogenic processes and cell death are not well understood. Inhibitors of glycosphingolipid biosynthesis were recently found to trigger apoptosis in many carcinoma including breast cancer SKBR-3, MCF-7, and MDA-468 cells through either intrinsic or extrinsic apoptotic pathways as we previously reported. These anti-cancer inhibitors could increase ceramide concentration by blocking functions of glycolipid glycosyltransferases (GLTs). In this study, using a novel fluorescent dye (ASK-0) revealed the damage of cell organelle membranes by an inhibitor of glucosylceramide biosynthesis (L-PPMP). A highly drug- and cell-dependent regulation of MAPKs was also found by cis-platin and L-PPMP when inducing apoptosis in SKBR-3, MCF-7, and MDA-468 cells. A dose and time-dependent regulation of GLTs were investigated by enzymatic assay and DNA microarray analyses. These GLTs are involved in biosynthesis of LeX and sialosyl-LeX (neolactosyl-ceramide series) such as GalT-4 (UDP-Gal: LcOse3cer β−galactosyltransferase, GalT-5 (UDP-Gal: nLcOse4Cer α1, 3galactosyltransferase, FucT-3 (GDP-Fucose: LM1 α1, 4fucosyltransferase). A similar effect was observed with the GLTs involved in the biosyntheses of Gg-series gangliosides, such as SAT-4 (CMP-NeuAc: GgOse4Cer α2, 3sialyltransferase, and SAT-2 (CMP-NeuAc: GM3 α2, 8sialyltransferase). The glycol-related gene DNA-microarrays also suggested the transcriptional regulation of several GLTs involved in the biosynthesis of neolactosylceramide containing cell-surface antigens in these apoptotic breast carcinoma cells. In the early apoptotic stages (2 to 6 h after L-PPMP treatment) in addition to GlcT-1 gene, several genes (βGalTs and βGlcNAcTs) in the SA-Lea pathway were stimulated. [ABSTRACT FROM AUTHOR]

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Abstract: 65–80% of the patients with breast cancer might not benefit from the adjuvant therapy they receive based on ‘classical’ markers used for the selection for adjuvant therapy. Therefore it is necessary to develop new markers that are able to tailor treatment for an individual patient. A number of microarray methods have been developed in recent years to accommodate this search for new factors that determine breast cancer progression. We give an overview of the most commonly used microarray methods to identify tumour progression markers (oligo- or cDNA arrays, CGH arrays, PCR arrays, and tissue microarrays). Their applications will be illustrated using the most influential examples from literature. The potentials, limitations and the related statistical analyses of each method are discussed. We conclude that microarray studies have led to an increase in the understanding of the complexity and diversity of breast carcinoma and have provided clinical relevant subgroups of breast cancer that may benefit from patient tailored treatment. Still, more extensive external validation and long-term follow-up will be necessary before such assays can be implemented into routine clinical practice. Most likely, these novel prognostic indicators will be complementary to the already available classical prognostic factors. [Copyright &y& Elsevier]

DNA-microarray technology is a powerful tool to explore the potentials of functional genomics. Almost a decade back, it was not so easy to trace the molecular pathway process in the pathogenesis of diseases. Today DNA-microarray technology allows identifying candidate genes that are implicated in the development and progression of diseases. Because this technology is new, it remains a challenge to perform DNA-microarray experiments in the laboratory. All the technical factors associated with DNA-microarray experiments have a strong impact on the results and therefore, all parts/steps of the protocol, in particular sample preparation (i.e., RNA isolation, RNA preamplification, and cDNA labeling), hybridization, washing, and scanning settings need to be checked and optimized to obtain reliable results from DNA-microarray experiments. [ABSTRACT FROM AUTHOR]

Kallikreins play an important role in tumour microenvironment and as cancer biomarkers in different cancer entities. Previous studies suggested an upregulation of KLK10 and KLK6 in pancreatic ductal adenocarcinoma (PDAC). Therefore, we evaluated the clinicopathological role of these kallikreins and their value as biomarkers in PDAC.Differential expression was validated by DNA-microarrays and immunohistochemistry in normal and malignant pancreatic tissues. Sera concentrations of both kallikreins were evaluated using ELISA. In silico analysis of possible protein interactions and gene silencing of KLK10 in vitro using siRNAs gave further insights in the pathomechanisms.Gene expression analysis and immunohistochemistry demonstrated a strong expression for KLK10 and KLK6 in PDAC. Statistical analysis showed that co-expression of these kallikreins correlated with an R1-resection status (P=0.017) and worse outcome for overall survival (P=0.031). Multivariate analysis proofed that co-expression is an independent prognostic factor for survival (P=0.043). Importantly, KLK10 knockdown in AsPC-1 cells significantly reduced cell migration, whereas computational analysis suggested interaction of KLK6 with angiogenetic factors as an important mechanism.Co-expression of KLK10 and KLK6 plays an unfavourable role in PDAC. Our results suggest that this effect is likely mediated by an interaction with the factors of the extracellular matrix and enhancement of cancer cell motility.British Journal of Cancer (2008) 99, 1484–1492. doi:10.1038/sj.bjc.6604717 www.bjcancer.com Published online 14 October 2008 [ABSTRACT FROM AUTHOR]

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Abstract: Objective: The problem of marker selection in DNA microarray analysis has been addressed so far by two basic types of approaches, the so-called filter and wrapper methods. Wrapper methods operate in a recursive fashion where feature (gene) weights are re-evaluated and dynamically changing from iteration to iteration, while in filter methods feature weights remain fixed. Our objective in this study is to show that the application of filter criteria in a recursive fashion, where weights are potentially adjusted from cycle to cycle, produces noticeable improvement on the generalization performance measured on independent test sets. Methods and materials: Toward this direction we explore the behavior of two well known and broadly accepted pattern recognition approaches namely the support vector machines (SVM) and a single linear neuron (LN), properly adapted to the problem of marker selection. Within this context we also show how the kernel ability of SVM could be employed in a practical manner to provide alternative ways to approach the problem of reliable marker selection. Results: We explore how the proposed approaches behave in two application domains (breast cancer and leukemia), achieving comparable or even better results than those reported in the related bibliography. An important advantage of these approaches is their ability to derive stable performance without deteriorating due to the complexity of the application domain. Validation is performed using internal leave one out (ILOO) and 10-fold cross validation as well as independent test set evaluation. Conclusions: Results show that the proposed methodologies achieve remarkable performance and indicate that applying filter criteria in a wrapper fashion (‘wrapper filtering criteria’) provides a useful tool for marker selection. The contribution of this study is threefold. First it provides a methodology to apply filter criteria in a wrapper way (which is a new approach), second it introduces a fundamental pattern recognition component namely the single neuron (which is a linear estimator) and explores its behavior on marker selection and third it demonstrates an approach to exploit the kernel ability of SVMs in a practical and effective manner. [Copyright &y& Elsevier]

Abstract: Staphylococcus aureus is a common pathogen which can colonise and infect not only man, but also domestic animals. Especially, infection of cattle is of high economic relevance as S. aureus is an important causal agent of bovine mastitis. In the present contribution, a DNA microarray was applied for the study of 144 different gene targets, including resistance genes and genes encoding exotoxins, in S. aureus isolated from cows. One hundred and twenty-eight isolates from Germany and Switzerland were tested. These isolates were assigned to 20 different strains and nine clonal complexes. The majority of isolates belonged either to apparently closely related clonal complexes 8, 25, and 97 (together 34.4%) or were related to the sequenced bovine strain RF122 (48.4%). Notable characteristics of S. aureus of bovine origin are the carriage of intact haemolysin beta (in 82% of isolates tested), the absence of staphylokinase (in 89.1%), the presence of allelic variants of several exotoxins such as toxic shock syndrome toxin and enterotoxin N, and the occurrence of the leukocidin lukF-P83/lukM (in 53.1%). Two isolates were methicillin-resistant S. aureus (MRSA). One of them was a clonal complex 8 MRSA related to the epidemic MRSA strain Irish 01. The other one belonged to ST398/spa-type 34 resembling a newly emerging MRSA strain which has been described to occur in humans as well as in domestic animals. The presence of these two strains highlights the possibility of transfers of S. aureus strains between different host species. [Copyright &y& Elsevier]

The molecular pathogenesis of pleomorphic xanthoastrocytoma (PXA), a rare astrocytic brain tumor with a relatively favorable prognosis, is still poorly understood. We characterized 50 PXAs by comparative genomic hybridization (CGH) and found the most common imbalance to be loss on chromosome 9 in 50% of tumors. Other recurrent losses affected chromosomes 17 (10%), 8, 18, 22 (4% each). Recurrent gains were identified on chromosomes X (16%), 7, 9q, 20 (8% each), 4, 5, 19 (4% each). Two tumors demonstrated amplifications mapping to 2p23–p25, 4p15, 12q13, 12q21, 21q21 and 21q22. Analysis of 10 PXAs with available high molecular weight DNA by high-resolution array-based CGH indicated homozygous 9p21.3 deletions involving the CDKN2A/p14ARF/CDKN2B loci in six tumors (60%). Interphase fluorescence in situ hybridization to tissue sections confirmed the presence of tumor cells with homozygous 9p21.3 deletions. Mutational analysis of candidate genes on 9q, PTCH and TSC1, revealed no mutations in PXAs with 9q loss and no evidence of TSC1 promoter methylation. However, PXAs consistently showed low TSC1 transcript levels. Taken together, our study identifies loss of chromosome 9 as the most common chromosomal imbalance in PXAs and suggests important roles for homozygous CDKN2A/p14ARF/CDKN2B deletion as well as low TSC1 mRNA expression in these tumors.Oncogene (2007) 26, 1088–1097. doi:10.1038/sj.onc.1209851; published online 7 August 2006 [ABSTRACT FROM AUTHOR]

Loss of intestinal mucosa integrity is an important factor in the pathogenesis of inflammatory bowel disease (IBD). The aim of this study was to characterize expression changes and allelic variants of genes related to intestinal epithelial barrier function in this disease. Therefore, ileal and colonic mucosal biopsies from nonaffected regions of patients with ulcerative colitis (UC) and Crohn’s disease (CD), as well as non-IBD probands, were subjected to Affymetrix DNA-microarray analysis. Real-time reverse transcription polymerase chain reaction was used for verification in larger IBD sample numbers. Disturbed mRNA expression was identified for several mucin genes in both disease groups and tissues. A significant downregulation in the colon was obtained for MUC2 in CD and MUC12 in CD and UC. Expression analysis of all dysregulated mucins in a broad human tissue panel revealed dominant epithelial tissue-specific transcription. In silico analysis of the regulatory regions of these mucins indicated nuclear factor κB (NFκB) binding sites in each promoter. Furthermore, NFκB was overrepresented in mucin promoters and a component of a specific combination of transcription factors (composite module). In vivo stimulation experiments in the adenocarcinoma cell line LS174T showed inducible mucin expression by the cytokines tumor necrosis factor-α and transforming growth factor-β, which could be blocked by NFκB signaling inhibitors. Allelic discrimination screening obtained statistically significant associations for the MUC2–V116M ( P = 0.003) polymorphism with CD and for MUC4–A585S ( P = 0.025), as well as MUC13–R502S ( P = 0.0003) with UC. These data suggest that the disturbed expression of mucin genes and the connection to the NFκB pathway may influence the integrity of the intestine and therefore contribute to the pathophysiology of IBD. [ABSTRACT FROM AUTHOR]

Abstract: The goal of this study was to identify glycosyltransferases that are specifically expressed in pancreatic cancer. To investigate the gene expression of glycosyltransferases between pancreatic cancer and normal pancreatic tissues, we performed DNA-microarray (involving about 1000 oligosaccharide-related genes) using RNA mixtures of pancreatic cancer cells and normal pancreatic tissues. Eighty-six genes were up-regulated and thirty-two were down-regulated in pancreatic cancer, compared to normal pancreatic tissue. Among these changes, it is noteworthy that the expression of GnT-IVa was decreased and the expression of GnT-IVb was increased in pancreatic cancer, compared to normal pancreatic tissues. Although GnT-IVa and -IVb are involved in the same reaction as a glycosyltransferase, their chromosomal localization is different. When 5 cases of pancreatic cancer tissues were examined using the real-time RT-PCR method, the expression of GnT-IVb was dominant in tumor tissues and the expression of GnT-IVa was dominant in the surrounding normal tissues. The expression of GnT-IVa was increased in all 3 cell lines that had been treated with 5-aza-C and butyrate. These results suggest that the down-regulation of GnT-IVa in pancreatic cancer cells is due to an epigenetic abnormality in the gene. [Copyright &y& Elsevier]

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Abstract: Intracoronary stenting has markedly improved the patency of native coronary arteries after percutaneous transluminal coronary angioplasty (PTCA). Advances in stent technology and design, including drug releasing stents, have contributed to reduce the long-term restenosis rate. However, stenosis caused by neointimal hyperplasia, vascular remodeling and thrombosis is still a major problem after endocoronary stent procedures. This study focuses on differential gene expression of circulating peripheral blood cells after 90min exposure to stents to search for initially activated cellular pathways, which may foster restenosis. Fresh human whole blood (1IUheparin/ml), taken from non-medicated healthy volunteers, was incubated under flow conditions in an in vitro closed-loop stent-testing model (modified Chandler-Loop). Differential gene expression compared to resting conditions and to the experimental controls was investigated by a DNA-microarray technique encoding for over 17,000 genes simultaneously. As expected, a large variety of genes showed differential gene expression. Interestingly, Thrombospondin 1 (TSP-1), which plays a key role in initial immune defense, was found to be the most markedly up-regulated gene. We propose TSP-1 expression as an early indicator for the activation of immune responses following intracoronary stenting. After clarifying the participation of TSP-1 in vivo, future studies will therefore focus on TSP-1 as a potential prognostic factor, which may also help to develop and control new surface materials with an improved biocompatibility. [Copyright &y& Elsevier]

Abstract: Background and aims.: Probiotics have been used for cure and prevention of several clinical conditions. However, further insights into the mechanism of action are needed to understand the rationale of their use. The aim of this study was to investigate the influence of Lactobacillus GG on the genetic expression patterns in the small bowel mucosa. Methods.: Six male patients (38±5 years) with endoscopically proven oesophagitis were enrolled. All patients were treated for 1 month with esomeprazole and randomised to receive Lactobacillus GG or placebo. After 1 month of treatment, upper endoscopy was repeated. Biopsies of the duodenal mucosa were taken prior to and after the treatment, and the genes expression patterns were assessed using GeneChip Human U133A array. Genes with significant expression changes were selected and analysed to identify specific cellular pathways modified by Lactobacillus GG. To support the array data, 10 target genes were studied using Syber-Green PCR. Results.: Microarray analysis showed that Lactobacillus GG administration determined the up- and down-regulation of 334 and 92 genes, respectively. Real-time PCR confirmed the reliability of the analysis. Lactobacillus GG mainly affected the expression of genes involved in immune response and inflammation (TGF-beta and TNF family members, cytokines, nitric oxide synthase 1, defensin alpha 1), apoptosis, cell growth and cell differentiation (cyclins and caspases, oncogenes), cell–cell signalling (ICAMs and integrins), cell adhesion (cadherins), signal transcription and transduction. Conclusions.: These data indicate that administration of Lactobacillus GG is associated with a complex genetic response of the duodenal mucosa, reflected by the up- and down-regulation of several genes involved in specific cellular pathways. [Copyright &y& Elsevier]