Maintenance of “stem cell” features of cartilage cell sub-populations during in vitro propagation

Abstract

BackgroundThe discovery of mesenchymal stem cells (MSCs) or MSC-like cells in cartilage tissue does not tie in well with the established view that MSCs derive from a perivascular niche. The presence of MSCs may raise concerns about specificity and application safety, particularly in terms of the regulatory site. The aim of the present study was to investigate the benefits or possible risks of the MSC-like properties of cells isolated from cartilage in the context of autologous chondrocyte implantation.MethodsChondrocytic cells were isolated from cartilage or intervertebral disc tissue. Flow cytometry was used to analyze the expression of cell surface antigens. MSC-like cells were either enriched or depleted by means of magnetic cell sorting (MACS) involving the monoclonal antibodies W5C5/SUSD2 and W8B2/MSCA-1. We addressed the issues of prolonged expansion of such cells as well as the influence of culture medium as a trigger for selecting a single cell type. Established protocols were used to study in vitro differentiation. In addition to histological and biochemical assessment, the acquired phenotypes were also evaluated on the mRNA transcript level.ResultsIn the studied cells, we found strongly analogous expression of antigens typically expressed on MSCs, including CD49e, CD73, CD90, CD105, CD140b and CD166. The expression of W5C5 and W8B2 antigens in cartilage cell sub-populations did not correlate with multi-potency. We demonstrated that a chondroid precursor, but not a bona fide multipotent mesenchymal, cell type can be obtained under established in vitro culture conditions. The culture media used for expansion influenced the cell phenotype.ConclusionsThe risk of adverse adipose or osseous differentiation is not posed by expanded chondrocyte cultures, even after enrichment of putative MSC-like cell populations by MACS. It is possible that this limited “stemness” in chondrocytes, expanded for use in ACI, may instead be beneficial as it allows re-differentiation under appropriate conditions despite prolonged times in culture.

Immortalisation with hTERT Impacts on Sulphated Glycosaminoglycan Secretion and Immunophenotype in a Variable and Cell Specific Manner

“…Up-regulation of HLA-DR expression has been observed previously on MSCs, more so when cells were isolated from whole bone marrow aspirate rather than separated mononuclear cells, and particularly in response to exposure to bFGF[ 71 ][ 72 ] . Interestingly we found that OK3 chondrocytes expressed CD73, CD90 and CD105, expression of MSC markers on chondrocytes has been reported previously[ 8 ][ 73 ] with a greatly increased expression of CD90 in response to monolayer culture[ 8 ]. Alternatively it has been proposed that these cells may be representative of a chondroprogenitor or MSC like cell type present within cartilage[ 73 ].…”

In vitro characterization of human articular chondrocytes and chondroprogenitors derived from normal and osteoarthritic knee joints

“…A very important observation was that all chondrocyte groups including freshly isolated cells expressed high levels of CD49e and CD29, matching those expressed in CPs. These results indicate that CD49e is not a cell specific marker when differentiation between chondrocytes and CPs is required as has already been evidenced by earlier reports(15,39). CD166 in addition to being considered a marker of strong chondrogenic potential has also been reported to be an identifier for mesenchymal progenitor cells within human cartilage(31,40).…”

Human adult stem cells from diverse origins: An overview from multiparametric immunophenotyping to clinical applications

“…The recent generation of Nanog‐GFP knock‐in human ESCs demonstrates that Nanog exhibits heterogeneous expression in human ESCs as well, but it remains to be analyzed whether similar functional differences exist between NANOG low and NANOG high human ESCs . Besides some authors consider MSC plasticity as a consequence of cell culture conditions rather than an intrinsic MSC in vivo differentiation potential it is reasonable that Nanog heterogeneity expression and maybe other transcription factor on downstream Nanog's cascade could control MSC plasticity. It was demonstrated that germ layer marker expression pattern would support the concept that MSCs possess plasticity, as it was shown by three‐germ line marker co‐expression and MSC capacity to differentiate into 3 germ lines (see Table for differentiation protocols of MSCs).…”