Integrated culture platform based on a human platelet lysate supplement for the isolation and scalable manufacturing of umbilical cord matrix-derived mesenchymal stem/stromal cells

Abstract

Umbilical cord matrix (UCM)-derived mesenchymal stem/stromal cells (MSCs) are promising therapeutic candidates for regenerative medicine settings. UCM MSCs have advantages over adult cells as these can be obtained through a non-invasive harvesting procedure and display a higher proliferative capacity. However, the high cell doses required in the clinical setting make large-scale manufacturing of UCM MSCs mandatory. A commercially available human platelet lysate-based culture supplement (UltraGRO , AventaCell BioMedical) (5%(v/v)) was tested to effectively isolate UCM MSCs and to expand these cells under (1) static conditions, using planar culture systems and (2) stirred culture using plastic microcarriers in a spinner flask. The MSC-like cells were isolated from UCM explant cultures after 11 ± 2 days. After five passages in static culture, UCM MSCs retained their immunophenotype and multilineage differentiation potential. The UCM MSCs cultured under static conditions using UltraGRO -supplemented medium expanded more rapidly compared with UCM MSCs expanded using a previously established protocol. Importantly, UCM MSCs were successfully expanded under dynamic conditions on plastic microcarriers using UltraGRO -supplemented medium in spinner flasks. Upon an initial 54% cell adhesion to the beads, UCM MSCs expanded by >13-fold after 5-6 days, maintaining their immunophenotype and multilineage differentiation ability. The present paper reports the establishment of an easily scalable integrated culture platform based on a human platelet lysate supplement for the effective isolation and expansion of UCM MSCs in a xenogeneic-free microcarrier-based system. This platform represents an important advance in obtaining safer and clinically meaningful MSC numbers for clinical translation. Copyright © 2016 John Wiley & Sons, Ltd.

Scalable Manufacturing of Human Mesenchymal Stromal Cells in the Vertical‐Wheel Bioreactor System: An Experimental and Economic Approach

“…Importantly, the process did not compromise UCM identity, since they maintained the typical MSC immunophenotype and multilineage differentiation potential . To our best knowledge, the expansion results obtained are comparable, or even superior, to those described in the literature for MSC obtained from UCM …”

“…To establish the expansion platform, we first adapted the culture conditions previously reported by our group for the expansion of UCM MSC in spinner flasks, using plastic microcarriers combined with hPL‐supplemented medium . The same culture conditions were translated to the VW‐PBS system, including the initial cell density, microcarrier type and concentration, coating solution, agitation protocol during cell adhesion and feeding regime.…”

“…Conversely, agitation rate was maintained at 25 rpm throughout the culture period. Cell growth, viability, and metabolite analysis were assessed every day, as described in de Soure et al The specific growth rate during exponential growth phase was calculated as described in Hanga et al Cell visualization on microcarriers was performed from day 1 onwards, by staining the cells with 4′,6‐diamidino‐2‐phenylindole (DAPI, 1.5 µg mL −1 in phosphate‐buffered saline [PBS]), as previously described in de Soure et al…”

“…Before and after PBS‐0.1 MAG culture, cells were characterized in terms of immunophenotypic profile, while cells after PBS‐0.1 MAG culture were also tested for multilineage differentiation capacity …”

Multifunctional laminarin microparticles for cell adhesion and expansion

“…Recently, Soure and co-workers studied the effect of PL on the expansion of mesenchymal stem cells (MSCs) derived from umbilical cord using plastic microcarriers under dynamic conditions. (de Soure et al, 2017) Their results demonstrated the advantages of the use of PL for the effective expansion of MSCs in a xenogeneic-free microcarrier-based system. In the present study, it is proposed the fabrication of MeLam microparticles with encapsulated PL to be used as microplatforms for cell adhesion and expansion.…”