Towards an integrated bioprocess for scalable production and isolation of MSC- derived extracellular vesicles for cardiac repair

Conference Dates

February 6 – 10, 2022


Mesenchymal stem/stromal cells (MSC) and their associated extracellular vesicles (EV) hold great promise in the treatment of heart diseases, such as myocardial infarction. Particularly, hypoxic preconditioned MSC have been shown to promote growth of blood vessels and stimulate endogenous repair pathways in the myocardium and could therefore constitute a relevant cell source for an augmented production of EV with cardiac regenerative properties. Since the clinical translation of MSC-derived EV is currently limited by their scalability, we propose the use of a bioreactor-based platform to maximize the yields and potency of the generated EV while implementing scalable and standardized downstream processing protocols that could be integrated in a continuous bioprocess.

To establish scalable manufacturing of MSC-derived EV, we have developed a baseline process using microcarrier technology combined with stirred-tank bioreactor for MSC expansion and EV production, and downstream processing for EV isolation and concentration by Tangential Flow Filtration followed by Size Exclusion Chromatography (TFF-SEC). The potential of the applied strategy is highlighted by direct comparison with cell culture in static systems and isolation of MSC-derived EV by the gold standard, although less scalable, density gradient ultracentrifugation-based protocols (DG-UC).

We propose the use of a chemically-defined medium to support continuous manufacturing of both MSC and derived EV under cGMP-compatible and reproducible conditions. Key process parameters have been optimized to improve EV production and purification yields without compromising their quality attributes, with the TFF-SEC protocol yielding 6.5 times higher number of EV comparatively to the conditioned medium processed by DG-UC (n=1). EV were characterized in terms of yield as well as particle size distribution, and surface marker analysis by bead-based flow cytometry. Functional assays were implemented to assess EV potency in cardiac regeneration settings.


This work was performed under the scope of the CardioPatch project (SOE4/P1/E1063) and iNOVA4Health (UIDB/04462/2020 and UIDP/04462/2020), a program financially supported by FCT/Ministério da Educação e Ciência, through national funds.

Poster 22.pdf (30 kB)

This document is currently not available here.