La aplicación de Agroquímicos afecta negativamente a la Flora Nativa

There has been a huge surge in interest in the use of circulating exosome- encapsulated miRNAs as biomarkers for breast cancer in recent years. O’Brien et al., (2015) (129) recently reported on the role of exosomal miR- 134 in Triple-negative breast cancer (91). In this study the investigators isolated extracellular vesicles from the cell-conditioned media of a triple- negative cell line, Hs578T, and an aggressive clonal variant, Hs578Ts(i)8,

which they previously demonstrated to be capable of influencing the phenotype of recipient cells (90). MiRNA expression profiling was then carried out on the parent cells and their secreted exosomes using TaqMan Low-Density Assays (TLDA). Differing miRNA profiles were detected in the Hs578Ts(i)8 cells and exosomes when compared to the less aggressive

counterparts (Hs578T), with a greater proportion of miRNAs noted to be down-regulated in the former variant. MiR-134 demonstrated the greatest degree of down-regulation and was therefore further evaluated using the

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publically-accessible GSE40525 and GSE 26659 datasets, where it was confirmed to be significantly down-regulated in breast cancer (n=138) compared to normal breast tissue (n=73). In vitro studies demonstrated reduced proliferation, migration and invasion in Hs578Ts(i)8 cells following

transfection with miR-134, and so the functional impact of exosomes derived from these cells was investigated using Hs578Ts(i)8 parentcells. In

this setting, migration and invasion were significantly reduced, with no impact on proliferation and an accompanying decreased in STAT5B and Hsp90 protein levels. Increased sensitivity to Hsp90 inhibitors which have been investigated as a potentially valuable therapy in Triple-negative breast cancer was also demonstrated. The results of this study highlighted a number of key points with respect to exosomes in this cancer subtype. Firstly, the detection of a decrease in miR-134 levels in cancer cells may be indicative of a more aggressive cancer phenotype, which may assist in cancer diagnosis and more specifically, molecular subtype identification. Furthermore, the uptake of miR-134 enriched exosomes by recipient cancer cells may represent a valuable therapeutic modality in the management of Triple-negative breast cancer.

The impact of MSC-secreted exosomes has also been investigated, with one study noting a reduction in bone marrow-metastatic breast cancer cell line proliferation in response to the addition of these exosomes (130). A similar result has also been demonstrated by Del Fattore et al., (2015) (131) where glioblastoma cell proliferation reduced following a period of 48 hours incubation in the presence of MSC exosomes.

Our understanding of exosomes is continuously evolving as result of robust laboratory based research. The elegant work performed by Melo et al., (2014) (59) underscored the fact that cancer cell-derived exosomes differ greatly in content and functional effect to exosomes derived from non- tumourigenic cell lines. The authors first demonstrated how metastatic breast cancer cell-derived (MDA-MB-231 and 4T1) exosomes were enriched in miRNAs when compared to non-metastatic breast cancer exosomes (MCF7). In addition, profiling of purified cancer cell exosomes

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after 72 hours culture revealed enrichment of miRNAs compared to 24 hours incubation, an effect that was not observed in the case of control cell (MCF10A and NMuMG) exosomes (normosomes). Down-regulation of pre- miRNAs was detected in cancer exosomes over time with levels being inversely proportional to the corresponding miRNAs. No variation in normosome pre-miRNA levels were observed under the same conditions. Exosomes derived from cancer cells and from the serum of patients with breast cancer were found to contain the RNA-induced silencing complex (RISC) –loading complex (RLC) proteins, Dicer, TRBP, and AGO2, which are involved in miRNA biogenesis. The accumulation of Dicer specifically in cancer exosomes is reportedly mediated through CD43. These data demonstrated the ability of cancer cell exosomes to convert pre-miRNAs to mature miRNAs. Subsequent examination of the effect of transfer of cancer cell exosomes revealed increased survival and proliferation in a non- tumourigenic population of cells (59). This effect was examined further by injecting non-tumour cells with exosomes derived from a metastatic breast cancer cell line in a murine model, which resulted in tumour formation. A key finding in this study was that Dicer blockade inhibited tumour formation, suggesting the pivotal downstream effect that it plays in the transformation of non-tumourigenic cells to malignant cells. Similarly, serum exosomes from patients with breast cancer coinjected with MCF-10A cells resulted in tumour formation in mice while no tumours were formed when cancer exosomes were replaced with exosomes from healthy volunteers (59). Serum from patients with breast cancer was also noted to contain higher levels of exosomes than that which was isolated from healthy donors (59).

Work carried out by Tosar et al., (2015) (86) involved sequencing of the intracellular small regulatory RNAs (sRNA) content of breast epithelial cell lines (MCF-7 and MCF-10A) and comparison with extracellular fractions enriched in microvesicles, exosomes and ribonucleoprotein complexes. The results demonstrated a non-selective secretion model for the majority of microRNAs, with a few showing preferential secretion patterns. The authors

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proposed that this model might explain, at least in part, some of the conflicting published data regarding miRNA secretion. In contrast, 5′ tRNA halves and 5′ RNA Y4-derived fragments of 31–33 nt showed preferential secretion patterns. These data demonstrate that different sRNA families have characteristic secretion patterns and highlight their potential role in the extracellular space.