AIM: In our pilot study, we used plasma samples as liquid biopsy to search for miRNA signatures in patients with acute myeloid leukemia (AML) at diagnosis and in remission achieved after standard chemotherapy before planned transplantation. MATERIAL AND METHODS: We examined 10 plasma samples from healthy volunteers and 8 paired samples from patients with AML at diagnosis and in remission using TaqMan MicroRNA Arrays. The results were validated using single-target qPCR reactions run in triplicates. RESULTS: We selected 6 miRNAs with expressions significantly sensitive to therapy: miR-199b-5p, miR-301b, miR-326, miR-361-5p, miR-625 and miR-655. All selected miRNAs were not or very weakly expressed in healthy individuals. They were abundant in plasma in patients at diagnosis but their levels decreased after chemotherapy. CONCLUSION: We detected a therapy sensitive miRNA signature in plasma of patients with AML.

• MeSH
• Leukemia, Myeloid, Acute blood   drug therapy   genetics   MeSH
• Cytarabine administration & dosage   MeSH
• Adult MeSH
• Idarubicin administration & dosage   MeSH
• Remission Induction MeSH
• Consolidation Chemotherapy MeSH
• Real-Time Polymerase Chain Reaction MeSH
• Middle Aged MeSH
• Humans MeSH
• MicroRNAs biosynthesis   blood   genetics   MeSH
• Mitoxantrone administration & dosage   MeSH
• Pilot Projects MeSH
• Antineoplastic Combined Chemotherapy Protocols pharmacology   therapeutic use   MeSH
• Gene Expression Regulation, Leukemic drug effects   MeSH
• RNA, Neoplasm biosynthesis   blood   genetics   MeSH
• Transcriptome * MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Validation Study MeSH

The KRAS signalling pathway is pivotal for pancreatic ductal adenocarcinoma (PDAC) development. After the failure of most conventional cytotoxic and targeted therapeutics tested so far, the combination of taxane nab-paclitaxel (Abraxane) with gemcitabine recently demonstrated promising improvements in the survival of PDAC patients. This study aimed to explore interactions of conventional paclitaxel and experimental taxane SB-T-1216 with the KRAS signalling pathway expression in in vivo and in vitro PDAC models in order to decipher potential predictive biomarkers or targets for future individualised therapy. Mouse PDAC PaCa-44 xenograft model was used for evaluation of changes in transcript and protein levels of the KRAS signalling pathway caused by administration of experimental taxane SB-T-1216 in vivo. Subsequently, KRAS wild-type (BxPc-3) and mutated (MiaPaCa-2 and PaCa-44) cell line models were treated with paclitaxel to verify dysregulation of the KRAS signalling pathway gene expression profile in vitro and investigate the role of KRAS mutation status. By comparing the gene expression profiles, this study observed for the first time that in vitro cell models differ in the basal transcriptional profile of the KRAS signalling pathway, but there were no differences between KRAS mutated and wild-type cells in sensitivity to taxanes. Generally, the taxane administration caused a downregulation of the KRAS signalling pathway both in vitro and in vivo, but this effect was not dependent on the KRAS mutation status. In conclusion, putative biomarkers for prediction of taxane activity or targets for stimulation of taxane anticancer effects were not discovered by the KRAS signalling pathway profiling in various PDAC models.

• MeSH
• Albumins pharmacology   MeSH
• Deoxycytidine analogs & derivatives   pharmacology   MeSH
• Carcinoma, Pancreatic Ductal drug therapy   genetics   MeSH
• Humans MeSH
• Mice, Nude MeSH
• Mice MeSH
• Biomarkers, Tumor genetics   MeSH
• Cell Line, Tumor MeSH
• Pancreatic Neoplasms drug therapy   genetics   MeSH
• Paclitaxel pharmacology   MeSH
• Bridged-Ring Compounds pharmacology   MeSH
• Cell Proliferation drug effects   genetics   MeSH
• Antineoplastic Combined Chemotherapy Protocols pharmacology   MeSH
• Proto-Oncogene Proteins p21(ras) genetics   MeSH
• Signal Transduction drug effects   genetics   MeSH
• Taxoids pharmacology   MeSH
• Transcriptome drug effects   genetics   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND: Recently, several studies have demonstrated a negative prognostic impact of Ikaros (IKZF1) gene alterations in acute lymphoblastic leukemia (ALL). However, controversies still exist regarding the impact of IKZF1 in current treatment protocols. PROCEDURE: We simultaneously detected IKZF1 gene deletions by multiplex ligation-dependent probe amplification and gene expression of IKZF1 isoforms in 206 children with BCR/ABL-negative ALL treated with ALL IC-BFM 2002 protocol, in which risk stratification was not based on minimal residual disease (MRD), and validated the results on a cohort of 189 patients treated with MRD-directed ALL-BFM 2000 protocol. RESULTS: Deletion of IKZF1 was present in 14 of 206 (7%) ALL IC patients. Interestingly, gene expression did not completely correlate with the deletion status in either cohort. Deletions were not always reflected in the gene expression of dominant-negative isoforms, and conversely, 7 of 395 (2%) non-deleted cases overexpressed dominant-negative isoform Ik6. IKZF1 deletions significantly affected event-free survival (EFS) of the ALL IC cohort (41 ± 14% vs. 86 ± 3%, P < 0.0001). Regarding IKZF1 isoforms, only Ik6 overexpression had negative prognostic impact (EFS 50 ± 16% vs. 85 ± 3%, P = 0.003). In multivariate analysis, which included ALL IC risk criteria, flow-cytometric MRD and IKZF1 alterations, day 15 MRD and IKZF1 deletion status displayed an independent prognostic impact. CONCLUSIONS: We show that MRD-directed treatment diminishes prognostic impact of IKZF1 alterations. However, IKZF1 status alone or combined with day 15 flow cytometry can significantly improve risk stratification within BFM protocols at centers that do not perform antigen-receptor-based MRD monitoring.

• MeSH
• Precursor Cell Lymphoblastic Leukemia-Lymphoma genetics   mortality   pathology   MeSH
• Fusion Proteins, bcr-abl genetics   MeSH
• Child MeSH
• Humans MeSH
• Child, Preschool MeSH
• Disease-Free Survival MeSH
• Prognosis MeSH
• Flow Cytometry MeSH
• Neoplasm, Residual genetics   pathology   MeSH
• Risk Factors MeSH
• Ikaros Transcription Factor genetics   MeSH
• Transcriptome MeSH
• Check Tag
• Child MeSH
• Humans MeSH
• Male MeSH
• Child, Preschool MeSH
• Female MeSH
• Publication type
• Journal Article 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

Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) exhibit multilineage differentiation potential, adhere to plastic, and express a specific set of surface markers-CD105, CD73, CD90. Although there are relatively well-established differentiation protocols for WJ-MSCs, the exact molecular mechanisms involved in their in vitro long-term culture and differentiation remain to be elucidated. In this study, the cells were isolated from Wharton's jelly of umbilical cords obtained from healthy full-term deliveries, cultivated in vitro, and differentiated towards osteogenic, chondrogenic, adipogenic and neurogenic lineages. RNA samples were isolated after the differentiation regimen and analyzed using an RNA sequencing (RNAseq) assay, which led to the identification of differentially expressed genes belonging to apoptosis-related ontological groups. ZBTB16 and FOXO1 were upregulated in all differentiated groups as compared to controls, while TGFA was downregulated in all groups. In addition, several possible novel marker genes associated with the differentiation of WJ-MSCs were identified (e.g., SEPTIN4, ITPR1, CNR1, BEX2, CD14, EDNRB). The results of this study provide an insight into the molecular mechanisms involved in the long-term culture in vitro and four-lineage differentiation of WJ-MSCs, which is crucial to utilize WJ-MSCs in regenerative medicine.

• MeSH
• Apoptosis genetics   MeSH
• Cell Differentiation genetics   MeSH
• Chondrocytes MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Mesenchymal Stem Cells * MeSH
• Osteoblasts MeSH
• Nerve Tissue Proteins MeSH
• Transcriptome MeSH
• Adipocytes MeSH
• Wharton Jelly * MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Although improvements in culture system have enhanced in vitro embryo production, success rates are still not adequate. The reasons for developmental arrest of a part of in vitro produced embryos are unknown, but are connected in part with low cytoplasmic competence of oocytes. The immaturity of cytoplasm can negatively influence fertilization efficiency and subsequent progression through embryonic genome activation (EGA), which are necessary steps in further pre-implantation development. A large number of studies have compared mRNA abundance among oocytes with different developmental competence with the aim to find markers of the normal embryo development. The amount of mitochondrial DNA (mtDNA) and mRNA for mitochondrial transcriptional factors directing oxidative phosphorylation belongs to such promising markers. Nevertheless, recently published studies revealed that the mammalian embryo is able to compensate for a reduced level of mtDNA in oocyte during subsequent pre-implantation development. The search for other molecular markers is in progress. Characterization of oocyte and embryonic mRNA expression patterns during the pre-implantation period, and their relationship to the successful in vitro and in vivo development will be essential for defining the optimized culture conditions or the nuclear transfer protocols. Microarrays technology enables us to reveal the differentially expressed genes during EGA, and to compare the expression profile of in vivo and in vitro produced embryos. Recent evidence indicates that the depletion of the pool of stored maternal mRNAs is critical for subsequent embryo development. All these experiments gradually offer a list of possible candidates for quality and developmental competence markers for mammalian oocytes and pre-implantation embryos.

• MeSH
• Transcription, Genetic * MeSH
• RNA, Messenger genetics   metabolism   MeSH
• Oocytes metabolism   MeSH
• Cattle embryology   MeSH
• Transcriptome * MeSH
• Gene Expression Regulation, Developmental physiology   MeSH
• Animals MeSH
• Check Tag
• Cattle embryology   MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Adenosine triphosphate-binding cassette proteins constitute a large family of active transporters through extracellular and intracellular membranes. Increased drug efflux based on adenosine triphosphate-binding cassette protein activity is related to the development of cancer cell chemoresistance. Several articles have focused on adenosine triphosphate-binding cassette gene expression profiles (signatures), based on the expression of all 49 human adenosine triphosphate-binding cassette genes, in individual tumor types and reported connections to established clinicopathological features. The aim of this study was to test our theory about the existence of adenosine triphosphate-binding cassette gene expression profiles common to multiple types of tumors, which may modify tumor progression and provide clinically relevant information. Such general adenosine triphosphate-binding cassette profiles could constitute a new attribute of carcinogenesis. Our combined cohort consisted of tissues from 151 cancer patients-breast, colorectal, and pancreatic carcinomas. Standard protocols for RNA isolation and quantitative real-time polymerase chain reaction were followed. Gene expression data from individual tumor types as well as a merged tumor dataset were analyzed by bioinformatics tools. Several general adenosine triphosphate-binding cassette profiles, with differences in gene functions, were established and shown to have significant relations to clinicopathological features such as tumor size, histological grade, or clinical stage. Genes ABCC7, A3, A8, A12, and C8 prevailed among the most upregulated or downregulated ones. In conclusion, the results supported our theory about general adenosine triphosphate-binding cassette gene expression profiles and their importance for cancer on clinical as well as research levels. The presence of ABCC7 (official symbol CFTR) among the genes with key roles in the profiles supports the emerging evidence about its crucial role in various cancers. Graphical abstract.

• MeSH
• ATP-Binding Cassette Transporters biosynthesis   genetics   MeSH
• Carcinogenesis * MeSH
• Colorectal Neoplasms genetics   pathology   MeSH
• Humans MeSH
• Breast Neoplasms genetics   pathology   MeSH
• Pancreatic Neoplasms genetics   pathology   MeSH
• Cystic Fibrosis Transmembrane Conductance Regulator biosynthesis   genetics   MeSH
• Sulfonylurea Receptors biosynthesis   genetics   MeSH
• Gene Expression Regulation, Neoplastic MeSH
• Transcriptome genetics   MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Publication type
• Journal Article MeSH

Here we provide an overview of procedures for long-term cultivation, phenotyping, genotyping, and genetic transformation of cell cultures of tobacco cell lines BY-2 and VBI-0, and of A. thaliana, ecotype Landsberg erecta (LE) cell line. Notably, we present an improved protocol for BY-2 transformation and cloning and extend the available plant cell lines methodology toward high-throughput technologies like fluorescent-based cell sorting and transcriptomics.

• MeSH
• Arabidopsis cytology   genetics   MeSH
• Cell Culture Techniques methods   MeSH
• Cell Line MeSH
• Plants, Genetically Modified cytology   genetics   MeSH
• Genotyping Techniques methods   MeSH
• Cloning, Molecular methods   MeSH
• Flow Cytometry methods   MeSH
• Gene Expression Profiling methods   MeSH
• Nicotiana cytology   genetics   MeSH
• Transformation, Genetic MeSH
• Transcriptome MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Phylogenetic divergence in Asparagales plants is associated with switches in telomere sequences. The last switch occurred with divergence of the genus Allium (Amaryllidaceae) from the other Allioideae (formerly Alliaceae) genera, resulting in uncharacterized telomeres maintained by an unknown mechanism. To characterize the unknown Allium telomeres, we applied a combination of bioinformatic processing of transcriptomic and genomic data with standard approaches in telomere biology such as BAL31 sensitivity tests, terminal restriction fragment analysis, the telomere repeat amplification protocol (TRAP), and fluorescence in situ hybridization (FISH). Using these methods, we characterize the unusual telomeric sequence (CTCGGTTATGGG)n present in Allium species, demonstrate its synthesis by telomerase, and characterize the telomerase reverse transcriptase (TERT) subunit of Allium cepa. Our findings open up the possibility of studying the molecular details of the evolutionary genetic change in Allium telomeres and its possible role in speciation. Experimental studies addressing the implications of this change in terms of the interplay of telomere components may now be designed to shed more light on telomere functions and evolution in general.

• MeSH
• Allium enzymology   genetics   MeSH
• Chromosomes, Plant genetics   MeSH
• Phylogeny MeSH
• Genomics MeSH
• In Situ Hybridization, Fluorescence MeSH
• Evolution, Molecular * MeSH
• Base Sequence MeSH
• Sequence Analysis, DNA MeSH
• Telomerase genetics   metabolism   MeSH
• Telomere genetics   MeSH
• Transcriptome MeSH
• Computational Biology MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH