Standardized xeno- and serum-free culture platform enables large-scale expansion of high-quality mesenchymal stem/stromal cells from perinatal and adult tissue sources

Abstract

Background aims: Mesenchymal stem/stromal cells (MSCs) are of interest for the treatment of graft-versushost disease, autoimmune diseases, osteoarthritis and neurological and cardiovascular diseases. Increasing numbers of clinical trials emphasize the need for standardized manufacturing of these cells. However, many challenges related to diverse isolation and expansion protocols and differences in cell tissue sources exist. As a result, the cell products used in numerous trials vary greatly in characteristics and potency. Methods: The authors have established a standardized culture platform using xeno-and serum-free commercial media for expansion of MSCs derived from umbilical cord (UC), bone marrow and adipose-derived (AD) and examined their functional characteristics. Results: MSCs from the tested sources stably expanded in vitro and retained their biomarker expression and normal karyotype at early and later passages and after cryopreservation. MSCs were capable of colony formation and successfully differentiated into osteogenic, adipogenic and chondrogenic lineages. Pilot expansion of UC-MSCs and AD-MSCs to clinical scale revealed that the cells met the required quality standard for therapeutic applications. Conclusions: The authors' data suggest that xeno-and serum-free culture conditions are suitable for largescale expansion and enable comparative study of MSCs of different origins. This is of importance for therapeutic purposes, especially because of the numerous variations in pre-clinical and clinical protocols for MSCbased products.

Allogeneic human umbilical cord-derived mesenchymal stem/stromal cells for chronic obstructive pulmonary disease (COPD): study protocol for a matched case–control, phase I/II trial

“…In our previous study, we demonstrated that MSCs from perinatal and adult sources behaved differently even when they were cultured under a standardise culture platform (xeno-free and serum-free). 23 Therefore, although it seems that MSCs derived from different sources might show similar effects on COPD-induced animal models, we believe that the source of MSCs might play a role in the level of the therapy effectiveness and their mechanism of action might also different, especially when they are exposed to COPD-related microenvironment.…”

“…Thirty UC samples were obtained from healthy women with an uncomplicated, at term pregnancy who underwent serological testing, including tests for HIV, cytomegalovirus, Epstein-Barr virus, hepatitis A virus, hepatitis B virus, hepatitis C virus, syphilis, and chlamydia, at 38 weeks of pregnancy, as shown in previous study. 23 The UC tissues were collected at delivery and transferred to the Stem Cell Core Facility at the Vinmec Research Institute of Stem Cell and Gene Technology under ISO 14 644–1 (certification number: CR61119-1). To generate a UC-MSC line for the current study, a single eligible UC tissue will be processed, isolated and cultured under xeno-free and serum-free conditions as previously described.…”

“…To generate a UC-MSC line for the current study, a single eligible UC tissue will be processed, isolated and cultured under xeno-free and serum-free conditions as previously described. 23 UC-MSCs will be expanded under these conditions to passage 5 (P5) and cryopreserved in the serum- and xeno-free defined reagent CryoStore CS10 (Stem Cell Technology, Canada) in liquid nitrogen (gas phase) in an automated Brooks System (Brooks Life Science, USA) for long-term storage. The releasing criteria for UC-MSC products are shown in table 3 .…”

Human Adipose-derived Mesenchymal Stromal Cells Exhibit High HLA-DR Levels and Altered Cellular Characteristics under a Xeno-free and Serum-free Condition

“…To determine the proliferative capability of ADMSCs, cells were seeded with the density of 5,000 cells/cm 2 in T25 asks (Nunc) and cultured up to passage 7 as previously described [35]. Experiments were performed in triplicate for each donor.…”

“…Cells were seeded in triplicate in a six-well dish at the concentration of 20 cells/cm 2 and cultured in the respective media (PS or SM) for 10 to 14 days as previously described [35]. Cells were xed in Methanol (Merck, Germany) and stained with Giemsa solution (Merck, Germany), both for 5 minutes at room temperature.…”