Chondrogenic potency is lost but trophic repair is maintained by highly passaged mesenchymal stem/stromal cells

Abstract

Purpose: Bone marrow-derived mesenchymal stromal cells (MSCs) are defined as stem cells primarily because of their capacity for multipotent differentiation, however more recently they have been shown also to have trophic properties that may be important for promoting tissue repair through the release of regulatory molecules that can promote regeneration through the stimulation of endogenous cells, whilst also inhibiting immune/inflammatory processes. Each of these modes of action may be important when developing therapeutic strategies for the treatment of osteoarthritic joints using injectable MSCs. Since a viable MSC therapy is likely to require the use of highly expanded populations of allogenic cells, we have undertaken a detailed analysis of how the differentiation and trophic properties of MSCs change with in vitro ageing of the cells. Methods: MSCs were cultured from passage until senescence. Cells from multiple passages were tested for their potency when used either for chondrogenesis (cartilage tissue engineering assay) or trophic repair. Two methods were used as determinants of trophic repair: (i) a meniscal cartilage integration assay in which undifferentiated human MSCs on a collagen scaffold were used to induce integration of two pieces of sheep meniscal cartilage and (ii) an immunoregulation assay in which undifferentiated MSCs were used to suppress the proliferation of stimulated T-cells. Genomic and proteomic screening was used to identify potential markers of MSCS with capacity for differentiation and trophic repair. Results: MSCs from 4 different donors were found to proliferate for a minimum of 16 passages before growth arrest with cells from one donor continuing to divide even at passage 30 (Figure 1). The chondrogenic potency of these MSCs fell significantly for all four donors, when measured as either the dry weight of cartilage tissue engineering or the Glycosaminoglycans content of the engineered cartilage (Figure 2). However, there was no reduction in trophic repair whether measured as integration of meniscal cartilage or suppression of T-cell proliferation (Figure 3). Combined genomic and proteomic analysis demonstrated that earlier passage cells with both chondrogenic and trophic potency are positive for both the MMP13 gene and TIMP-1 protein, whereas later passage cells with poor chondrogenic potency but strong trophic potency are negative for the MMP13 gene but positive for the TIMP-1 protein (Figure 4). Conclusions: We have demonstrated clear differences in the regulation of chondrogenic potency and trophic properties of MSCs. The measurement of MMP13 gene and TIMP-1 protein markers will facilitate the design of future therapeutic strategies that exploit either of one these modes of action.View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)View Large Image Figure ViewerDownload Hi-res image Download (PPT)