Project description:Vesper bats (family Vespertilionidae) experienced a rapid adaptive radiation beginning around 36 mya that resulted in the second most species rich mammalian family. Coincident with that radiation was an initial burst of DNA transposon activity that has continued into the present. Deep sequencing of small RNAs from the vespertilionid, Eptesicus fuscus, as well as dog and horse revealed that substantial numbers of novel bat miRNAs are derived from DNA transposons unique to vespertilionids. In fact, 35.9% of Eptesicus-specific miRNAs derive from DNA transposons compared to 2.2 and 5.9% of dog- and horse-specific miRNAs, respectively and targets of several miRNAs are identifiable. Timing of the DNA transposon expansion and the introduction of novel miRNAs coincides remarkably well with the rapid diversification of the family Vespertilionidae. We suggest that the rapid and repeated perturbation of regulatory networks by the introduction of many novel miRNA loci was a factor in the rapid radiation.
Project description:We report the application of next-generation sequencing for high-throughput profiling of microRNA expression characterictic for horse sarcoid, which is a skin tumor. With the use of HiScanSQ system (Illumina), we obtained over 124 million sequence reads from microRNA libraries. As a result, we identified over 200 known, as well as around 500 potentially novel, miRNA sequences. The analysis of differential expression of the identified miRNAs revealed that over 100 miRNAs were up- or downregulated in the sarcoid tissue in comparison to the control. The analysis of pathways showed e.g. pathways in cancer, viral carcinogenesis or transcriptional misregulation in cancer. Furthermore, microRNAs associated with carcinogenesis in humans were identified, such as: miR-10a, miR-21, miR-200 family. Concluding, our results suggest that microRNAs are largely involved in the neoplastic transformation of horse sarcoids.
Project description:Vesper bats (family Vespertilionidae) experienced a rapid adaptive radiation beginning around 36 mya that resulted in the second most species rich mammalian family. Coincident with that radiation was an initial burst of DNA transposon activity that has continued into the present. Deep sequencing of small RNAs from the vespertilionid, Eptesicus fuscus, as well as dog and horse revealed that substantial numbers of novel bat miRNAs are derived from DNA transposons unique to vespertilionids. In fact, 35.9% of Eptesicus-specific miRNAs derive from DNA transposons compared to 2.2 and 5.9% of dog- and horse-specific miRNAs, respectively and targets of several miRNAs are identifiable. Timing of the DNA transposon expansion and the introduction of novel miRNAs coincides remarkably well with the rapid diversification of the family Vespertilionidae. We suggest that the rapid and repeated perturbation of regulatory networks by the introduction of many novel miRNA loci was a factor in the rapid radiation. A testicular tissue sample from dog, horse, and two different Eptesicus fuscus individuals. Four samples total.
Project description:Growing evidence supports the importance of extracellular vesicle (EV) as mediators of communication in pathological processes, including those underlying respiratory disease. However, establishing methods for isolating and characterizing EVs remains challenging, particularly for respiratory samples. This study set out to address this challenge by comparing different EV isolation methods and evaluating their impacts on EV yield, markers of purity, and proteomic signatures, utilizing equine/horse bronchoalveolar lavage samples. Horses are an excellent translational animal model for respiratory studies due to similarities with human immune responses, shared environmental exposures, and naturally occurring respiratory diseases including asthma. Further, horses are long-lived large animals that allow for longitudinal sample collection, and provide large sample volume and cell yield, which are particularly useful since EV research is commonly limited by low sample yields. Here, EVs were isolated from horse bronchoalveolar lavage fluid (BALF) using four different methods (ultracentrifugation, microcentrifugation, and two sizes of size exclusion chromatography columns) and characterized by measuring particle counts, EV purity, total protein yield, and proteomic cargo, with a specific focus on vesicle surface marker expression potentially informing cell type of origin. We found that size exclusion chromatography yielded the highest particle counts, greatest EV purity markers and elevated vesicle surface marker expression. Overall proteomic profiles differed across isolation methods, with size exclusion chromatography clustering separately from centrifugation. Taken together, our results demonstrate that different isolation methods impact characteristics of EVs, notably that size exclusion chromatography, compared to centrifugation methods, resulted in higher EV purity and better characterized proteomic diversity, including information on EV cell of origin. This is the first study to characterize proteomic profiles of EVs following different isolation methods using equine BALF. The results of this study will pave the way for future studies using equine samples as a model to characterize human respiratory tract EVs.
Project description:Osteoblast is one of the bone marrow cells and not only play a central role in bone turnover, but also play a role as supporting cell for hematopoietic stem/progenitor cells. In the field of radiotherapy, internal radiotherapy using radioactive isotopes that emit alpha particles is performed, and antitumor effect is expected by radioisotopes (such as 223-Ra) when the primary cancer cells metastasize to bone marrow. In general, higher dose rates of ionising radiation can induce cell death through DNA damage. However, the extent of DNA damage and repair (radiosensitivity) varies between tissue cell types. In the bone marrow environment, there are many unknowns regarding the radiosensitivity and the its related gene expression in osteoblast, especially the expression status of micro RNAs (miRNAs). The purpose of this study is to reveal the expression pattern of miRNAs in osteoblasts exposed to alpha particle radiation and to identify miRNAs associated with radiosensitivity. Osteoblastic cell line, MC3T3-E1 cell was exposed to 0.223 Gy/min alpha particles (total dose: 0.5 Gy and 1 Gy) and extracted total RNAs after the incubation of 24 h. A significantly up- or down-regulated 21 miRNAs were observed in comparison to non-irradiated control. Using omicsnet data analysis system, we focused on 5 miRNAs (miR-467f, miR-362-3p, miR-5119, miR-292a-5p, miR-466h-3p) and validated them using RT-qPCR. As a result, A higher expression of miR-362-3p after exposure of alpha particle irradiation was reproducibly observed. These results suggested that osteoblastic cell irradiated to alpha particle radiation has a unique miRNA expression pattern.
Project description:miRNA-sequencing of grapefruit-derived extracellular vesicles and fusion nanovesicles derived from grapefruit-derived extracellular vesicles and gingival mesenchymal stem cell-derived vesicles. We then performed gene expression profiling analysis to explore the miRNAs derived from grapefruit-derived extracellular vesicles, and the retention rate of miRNAs after membrane fusion
Project description:We use the illumina high-throughput sequencing technology to identify miRNAs between the exosomes of H9c2 cells with or without alcohol-induced.The H9c2 cells were cultured in serum-free medium and stimulated with ethanol (100 mmol/L) or PBS for 24 h, then collected the exosomes samples from serum-free medium. Exosomes were isolated and extracted by differential centrifugation and detected by electron microscopy, particle size and related marker proteins.In total, 123 differentially expressed miRNAs (12 upregulated and 111 downregulated) were screened by miRNA sequence.