Attachment, Growth, and Detachment of Human Mesenchymal Stem Cells in a Chemically Defined Medium

Abstract

The manufacture of human mesenchymal stem cells (hMSCs) for clinical applications requires an appropriate growth surface and an optimized, preferably chemically defined medium (CDM) for expansion. We investigated a new protein/peptide-free CDM that supports the adhesion, growth, and detachment of an immortalized hMSC line (hMSC-TERT) as well as primary cells derived from bone marrow (bm-hMSCs) and adipose tissue (ad-hMSCs). We observed the rapid attachment and spreading of hMSC-TERT cells and ad-hMSCs in CDM concomitant with the expression of integrin and actin fibers. Cell spreading was promoted by coating the growth surface with collagen type IV and fibronectin. The growth of hMSC-TERT cells was similar in CDM and serum-containing medium whereas the lag phase of bm-hMSCs was prolonged in CDM. FGF-2 or surface coating with collagen type IV promoted the growth of bm-hMSCs, but laminin had no effect. All three cell types retained their trilineage differentiation capability in CDM and were detached by several enzymes (but not collagenase in the case of hMSC-TERT cells). The medium and coating did not affect detachment efficiency but influenced cell survival after detachment. CDM combined with cell-specific surface coatings and/or FGF-2 supplements is therefore as effective as serum-containing medium for the manufacture of different hMSC types.

RETRACTED: A Bio Polymeric Adhesive Produced by Photo Cross-Linkable Technique

“…In addition, they can be used as delivery frameworks that would be further engineered for delayed, localized release of medications, such as pain treatment drugs, antibiotics or chemotherapy. We can use them as mediums for growth factors [1][2][3][4][5] and actual cell lines for…”

The Challenge of Human Mesenchymal Stromal Cell Expansion: Current and Prospective Answers

“…-compatibility with downstream processing Some commercially available microcarriers have been developed to improve cell attachment using new synthetic or natural materials, whereas others are optimized for the specific harvest requirements of hMSCs [35]. Nonporous microcarriers are most suitable for hMSC expansion and harvest because successful detachment with high viability is difficult to achieve using porous microcarriers [36].…”

“…XFM containing a mixture of proteins including human serum albumin and recombinant growth factors was used to expand hMSCs derived from bone marrow (bm-hMSCs) and adipose tissue on microcarriers [77] and a similar approach with a XFM containing components from human plasma has also been described [78]. A new CDM for hMSC expansion has been described in which each component has a Chemical Abstracts Service (CAS) registration number and none of the components frequently used in XFM formulations are present, such as serum albumin, insulin, transferrin, progesterone, hydrocortisone or estradiol [35]. The absence of attachment-promoting factors limits the attachment behavior of the hMSCs and the growth surface must therefore be coated with attachment-promoting substances.…”

“…To our knowledge, this stem cell 1 medium (Cell Culture Technologies, Switzerland) is the only protein/peptide-free CDM for hMSC expansion, mainly comprising defined concentrations of low-molecular weight compounds (50-250 Da, with only one component larger than 1000 Da). The addition of recombinant growth factors in combination with surface coatings makes this medium suitable for the attachment, spreading, growth and detachment of hMSCs derived from different tissues [35].…”

“…The optimal harvest yield was achieved by incubating the cells in PsP for 20 min. Studies using static cultures have shown that the choice of the enzyme must be considered in combination with the growth surface coating and the type of medium for each cell type on a case-by-case basis [35]. The authors compared the detachment of hMSCs and hMSC-TERT cells with trypsin, Accutase, collagenase and PsP when the cells were grown on untreated surfaces, or surfaces coated with collagen or fibronectin, combined with CDM or serum-containing media (SCM).…”