Strategies for non-invasive biomarker discovery in early detection of cancer are an urgent need. Extracellular vesicles (EVs) have generated increasing attention from the scientific community and are under intensive investigations due to their unique biological profiles and their non-invasive nature. EVs are membrane-enclosed vesicles with variable sizes and function. Such vesicles are actively secreted from multiple cell types and are considered as key vehicles for inter-cellular communications and signalling. The stability and potential to easily cross biological barriers enable EVs for exerting durable effects on target cells. These along with easy access to such vesicles, the consistent secretion from tumour during all stages of tumorigenesis and their content providing a reservoir of molecules as well as mirroring the identity of the cell of origin are virtues that have made EVs appealing to be assessed in liquid biopsy approaches and for using as a promising resource of biomarkers in cancer diagnosis and therapy and monitoring targeted cancer therapy. Early detection of EVs will guide time-scheduled personalised therapy. Surveying reliable and sensitive methods for rapid isolation of EVs from biofluids, the purity of isolated vesicles and their molecular profiling and marker specification for clinical translation in patients with cancer are issues in the area and the hot topics of many recent studies. Here, the focus is over methods for EV isolation and stratification for digging more information about liquid biopsy-based diagnosis. Extending knowledge regarding EV-based strategies is a key to validate independent patient follow-up for cancer diagnosis at early stages and inspecting the efficacy of therapeutics.

Recent advances on milk exosomes (EXO), cargoes in cell−cell communication, explored their role within and between individuals, including in dairy species. The potential use of EXO as biomarkers of disease and metabolic conditions adds significant interest to the study of EXO in milk. Although several researches have been carried out on circulating miRNA in the milk, less information is available about milk-derived exosomal miRNAs, which are stable over time and resistant to digestion and milk processing. EXO are taken up by recipient cells through specific mechanisms, which enable the selective delivery of cargoes. This suggests that EXO cargoes can be used as biomarkers of health. Nevertheless, methodological limitations and potential applications of milk EXO in dairy ruminants must be considered. The paucity of studies that associate the EXO cargo to specific challenges deserves further investigations to unravel the variation of miRNA and proteins cargo in relation to metabolic imbalance and infectious disease of the mammary gland.

Exosomes derived from hepatocellular carcinoma (HCC) cells are nanovesicles and are involved in the occurrence and development of HCC, they also serve as important carriers and drug targets of nanodrug delivery systems. The external shape and internal structure of exosomes are important indexes of identification, and isolated intact morphology is crucial to biological function integrity. However, given their susceptibility to various influencing factors, the external shape and internal structure of exosomes derived from HCC cells remain incompletely studied. In this study, exosomes purified from HCC cells were isolated at different centrifugation speeds and examined via multiple electron microscopy (EM) techniques. The results demonstrate that exosomes possess a nearly spherical shape and bilipid membranous vesicle with a concave cavity structure containing electron-dense and coated vesicles, suggesting the possible existence of subpopulations of exosomes with specific functions. The exosomes isolated at ultracentrifugation (UC) speed (≥110,000×g) presented irregular and diverse external morphologies, indicating the effect on the integrity of the exosomes. Transforming growth factor signaling bioactive substances (TGF-β1, S100A8, and S100A9) can be found in exosomes by performing Western blotting, showing that the internal content is associated with metastasis of HCC. These findings show that EMelectron microscopy and UC speed can affect exosome characteristics, including external shape, internal structure, and content of bioactive substances. The electron-dense and coated vesicles that had been discovered in exosomes might become new additional morphological features, which could help to improve the interpretation of experimental results and widen our understanding of exosome morphology.

The α-globin mRNA contains a C-rich stability element (CRE) in its 3′ untranslated region (3′ UTR) which is critical for the stability of this long-lived mRNA. A protein complex, termed the α-complex, forms on the CRE and has been shown to contribute to stabilization of the mRNA by at least two mechanisms, first by interacting with the poly(A)-binding protein (PABP) to prevent deadenylation, and second by protecting the mRNA from attack by an erythroid endoribonuclease. In this report, we demonstrate that the α-globin 3′ UTR can confer stability on a heterologous mRNA in cells, and this stability is dependent on the α-complex. Moreover, the stability was exclusively detected with cytoplasmic mRNA, suggesting that the regulation of α-globin mRNA stability is a cytoplasmic event. An additional mechanism by which the α-complex can confer stability on an RNA in vitro was also identified and shown to involve inhibition of 3′ to 5′ exonucleolytic degradation. Furthermore, using an in vitro mRNA decay system, we were able to follow the demise of the α-globin RNA and demonstrate that the decay was initiated by deadenylation followed by 3′-to-5′ decay carried out by the exosome and ultimately hydrolysis of the residual cap structure.

The majority of mRNA turnover is mediated either by mRNA decapping/5′-to-3′ decay or exosome-mediated 3′-to-5′ exonucleolytic decay. Current assays to assess mRNA decapping in vitro using cap-labeled RNA substrates rely on one-dimensional thin layer chromatography. This approach does not, however, resolve free phosphate from 7meGDP, the product of Dcp1p-mediated mRNA decapping. This can result in misinterpretation of the levels of mRNA decapping due to the generation of free phosphate following the action of the unrelated scavenger decapping activity on the products of exosome-mediated decay. In this report, we describe a simple denaturing acrylamide gel-based assay that faithfully resolves all of the possible products that can be generated from cap-labeled RNA substrates by turnover enzymes present in cell extracts. This approach allows a one-step assay to quantitatively assess the contributions of the exosome and DCP-1-type decapping on turnover of an RNA substrate in vitro. We have applied this assay to recalculate the effect of competition of cap-binding proteins on decapping in yeast. In addition, we have used the assay to confirm observations made on regulated mRNA decapping in mammalian extracts that contain much higher levels of exosome activity than yeast extracts.