Comparative proteome profiling in exosomes derived from porcine colostrum versus mature milk reveals distinct functional proteomes

Extracellular Vesicles

Ferreira, Rafaela Furioso, Thomas Blees, Farhad Shakeri, Andreas Buness, Marc Sylvester, Giovanni Savoini, Alessandro Agazzi, Vladimir Mrljak, and Helga Sauerwein. "Comparative proteome profiling in exosomes derived from porcine colostrum versus mature milk reveals distinct functional proteomes." Journal of Proteomics (2021): 104338.

Exosomes are membranous vesicles of endocytic origin, recently been considered as major players in cell-cell communication. Milk is highly complex, and diverse biocomponents provide adequate nutrition, transfer immunity, and promote adequate neonate development. Milk exosomes are suggested to have a key role in these processes, yet to be further explored, and the alteration of the exosomes' cargo in different stages of lactation stages is important for understanding the factors relevant in nursing and also for improving milk replacer products both for humans and animals. We isolated exosomes from porcine milk in different lactation stages and analyzed their content using a TMT-based high-resolution quantitative proteomic approach. Exosomes were isolated using ultracentrifugation coupled with size exclusion chromatography to enrich milk-derived exosomes in samples obtained at day 0, 7, and 14 after parturition, and characterized by nanoparticle tracking analysis, transmission electron microscopy, and Western blotting. Quantitative proteomics analysis revealed different proteome profiles for colostrum exosomes and milk exosomes. The functional analysis highlighted pathways related to the regulation of homeostasis to be upregulated in colostrum exosomes, and pathways such as endothelial cell development and lipid metabolism to be upregulated in mature milk exosomes. This study endorses the importance of exosomes as active biocomponents of milk and provides knowledge for future studies exploring their role in the regulation of immunity and growth of the newborn.

View full article

Recent Publications

Cigarette smoke (CS) represents one of the most relevant environmental risk factors for several chronic pathologies. Tissue damage caused by CS exposure is mediated, at least in part, by oxidative stress induced by its toxic and pro-oxidant components. Evidence demonstrates that extracellular vesicles (EVs) released by various cell types exposed to CS extract (CSE) are characterized by altered biochemical cargo and gained pathological properties. In the present study, we evaluated the content of oxidized proteins and phospholipid fatty acid profiles of EVs released by human bronchial epithelial BEAS-2B cells treated with CSE. This specific molecular characterization has hitherto not been performed. After confirmation that CSE reduces viability of BEAS-2B cells and elevates intracellular ROS levels, in a dose-dependent manner, we demonstrated that 24 h exposure at 1% CSE, a concentration that only slight modifies cell viability but increases ROS levels, was able to increase carbonylated protein levels in cells and released EVs. The release of oxidatively modified proteins via EVs might represent a mechanism used by cells to remove toxic proteins in order to avoid their intracellular overloading. Moreover, 1% CSE induced only few changes in the fatty acid asset in BEAS-2B cell membrane phospholipids, whereas several rearrangements were observed in EVs released by CSE-treated cells. The impact of changes in acyl chain composition of CSE-EVs accounted for the increased saturation levels of phospholipids, a membrane parameter that might influence EV stability, uptake and, at least in part, EV-mediated biological effects. The present in vitro study adds new information concerning the biochemical composition of CSE-related EVs, useful to predict their biological effects on target cells. Furthermore, the information regarding the presence of oxidized proteins and the specific membrane features of CSE-related EVs can be useful to define the utilization of circulating EVs as marker for diagnosing of CS-induced lung damage and/or CS-related diseases.