Background: MiRNAs are essential mediators of many biological processes. The aim of this study was to investigate the dynamics of miRNA-mRNA regulatory networks during exercise and the subsequent recovery period. Results: Here we monitored the transcriptome changes using microarray analysis of the whole blood of eight highly trained athletes before and after 30 min of moderate exercise followed by 30 min and 60 min of recovery period. We combined expression profiling and bioinformatics and analysed metabolic pathways enriched with differentially expressed mRNAs and mRNAs which are known to be validated targets of differentially expressed miRNAs. Finally we revealed four dynamically regulated networks comprising differentially expressed miRNAs and their known target mRNAs with anti-correlated expression profiles over time. The data suggest that hsa-miR-21-5p regulated TGFBR3, PDGFD and PPM1L mRNAs. Hsa-miR-24-2-5p was likely to be responsible for MYC and KCNJ2 genes and hsa-miR-27a-5p for ST3GAL6. The targets of hsa-miR-181a-5p included ROPN1L and SLC37A3. All these mRNAs are involved in processes highly relevant to exercise response, including immune function, apoptosis, membrane traffic of proteins and transcription regulation. Conclusions: We have identified metabolic pathways involved in response to exercise and revealed four miRNA-mRNA networks dynamically regulated following exercise. This work is the first study to monitor miRNAs and mRNAs in parallel into the recovery period. The results provide a novel insight into the regulatory role of miRNAs in stress adaptation.

Background Quantification and normalization of RT-qPCR data critically depends on the expression of so called reference genes. Our goal was to develop a strategy for the selection of reference genes that utilizes microarray data analysis and combines known approaches for gene stability evaluation and to select a set of appropriate reference genes for research and clinical analysis of breast samples with different receptor and cancer status using this strategy. Methods A preliminary search of reference genes was based on high-throughput analysis of microarray datasets. The final selection and validation of the candidate genes were based on the RT-qPCR data analysis using several known methods for expression stability evaluation: comparative ∆Ct method, geNorm, NormFinder and Haller equivalence test. Results A set of five reference genes was identified: ACTB, RPS23, HUWE1, EEF1A1 and SF3A1. The initial selection was based on the analysis of publically available well-annotated microarray datasets containing different breast cancers and normal breast epithelium from breast cancer patients and epithelium from cancer-free patients. The final selection and validation were performed using RT-qPCR data from 39 breast cancer biopsy samples. Three genes from the final set were identified by the means of microarray analysis and were novel in the context of breast cancer assay. We showed that the selected set of reference genes is more stable in comparison not only with individual genes, but also with a system of reference genes used in commercial OncotypeDX test. Conclusion A selection of reference genes for RT-qPCR can be efficiently performed by combining a preliminary search based on the high-throughput analysis of microarray datasets and final selection and validation based on the analysis of RT-qPCR data with a simultaneous examination of different expression stability measures. The identified set of reference genes proved to be less variable and thus potentially more efficient for research and clinical analysis of breast samples comparing to individual genes and the set of reference genes used in OncotypeDX assay.

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Significance: Post-traumatic sterile inflammation is the first necessary step of wound healing. In addition, sterile inflammation underlies the pathogenesis of a multitude of common diseases, such as allergies and autoimmune diseases. The molecular mechanisms underlying sterile inflammation are still not fully understood. Here, we show that the receptor Mincle (Clec4e), the expression of which is highly induced in the skin in response to damage, recognizes cholesterol sulfate, a molecule that is abundant in the epidermal layer of the skin, subsequently inducing a pro-inflammatory response. We also identify a role for Mincle as a driving component in the pathogenesis of allergic skin inflammation. The results demonstrate a previously unconsidered important role of Mincle in mediating sterile inflammation.

High and moderate intensity endurance exercise alters gene expression in human white blood cells (WBCs), but the understanding of how this effect occurs is limited. To increase our knowledge of the nature of this process, we investigated the effects of passing the anaerobic threshold (AnT) on the gene expression profile in WBCs of athletes. Nineteen highly trained skiers participated in a treadmill test with an incremental step protocol until exhaustion (ramp test to exhaustion, RTE). The average total time to exhaustion was 14:40 min and time after AnT was 4:50 min. Two weeks later, seven of these skiers participated in a moderate treadmill test (MT) at 80% peak O₂ uptake for 30 min, which was slightly below their AnTs. Blood samples were obtained before and immediately after both tests. RTE was associated with substantially greater leukocytosis and acidosis than MT. Gene expression in WBCs was measured using whole genome microarray expression analysis before and immediately after each test. A total of 310 upregulated genes were found after RTE, and 69 genes after MT of which 64 were identical to RTE. Both tests influenced a variety of known gene pathways related to inflammation, stress response, signal transduction and apoptosis. A large group of differentially expressed previously unknown small nucleolar RNA and small Cajal body RNA was found. In conclusion, a 15-min test to exhaustion was associated with substantially greater changes of gene expression than a 30-min test just below the AnT.