Fibroblast Growth Factor-1 Is a Mesenchymal Stromal Cell-Secreted Factor Stimulating Proliferation of Osteoarthritic Chondrocytes in Co-Culture

Abstract

Previously, we showed that mesenchymal stromal cells (MSCs) in co-culture with primary chondrocytes secrete soluble factors that increase chondrocyte proliferation. The objective of this study is to identify these factors. Human primary chondrocytes (hPCs) isolated from late-stage osteoarthritis patients were co-cultured with human bone marrow-derived MSCs (hMSCs) in pellets. Genome-wide mRNA expression analysis and quantitative polymerase chain reactions (qPCR) were used to identify soluble factors that were specifically induced in co-cultures. Immunofluorescent staining combined with cell tracking and enzyme-linked immunosorbent assay (ELISA) were performed to validate up-regulation at the protein level and to identify the cellular origin of the increased proteins. Chemical blockers and neutralizing antibodies were used to elucidate the role of the identified candidate genes in co-cultures. A number of candidate factors were differentially regulated in co-cultures at the mRNA level. Of these, fibroblast growth factor-1 (FGF-1) mRNA and protein expression were markedly increased in co-cultures predominantly due to up-regulated expression in MSCs. Blocking of FGF signaling in co-culture pellets by specific FGF receptor inhibitors or FGF-1 neutralizing antibodies completely blocked hPCs proliferation. We demonstrate that MSCs increase FGF-1 secretion on co-culture with hPCs, which, in turn, is responsible for increased hPCs proliferation in pellet co-cultures. C artilage repair is among the key targets of regenerative medicine. Several stem cell-based therapies have been proposed to treat cartilage defects, rheumatic arthritis, osteoarthiris (OA), and other joint diseases. These therapeutic strategies include implantation or injection of mesenchymal stromal cells (MSCs) in the diseased joint or bone marrow stimulation techniques such as micro fracture. These procedures rely on cellular interactions between MSCs and native cells in the joint [1][2][3][4]. Recent studies showed that intra-articularly injected MSCs can reduce incidence and severity of arthritis in animal model by inducing immune tolerance [5]. Further investigations showed that MSCs are capable of inducing hyporesponsiveness of T-lymphocytes and of expressing antiinflammatory mediators in macrophages [6,7]. Besides immuno-suppressive effects, intra-articularly injected MSCs were also reported to inhibit thickening of the synovium and to protect against cartilage destruction in a mouse OA model [8]. However, with the cellular interactions between MSCs and native chondrocytes receiving less attention [9], the molecular mechanism of interactions between MSCs and native chondrocytes is poorly understood [10]. In order to maximize the benefits of stem cell therapy, these cellular interactions need to be clarified.Previous reports showed that cartilage matrix formation was increased in co-cultures of human primary chondrocytes (hPCs) with MSCs compared with monocultures [11,12]. Further investigations revealed that the inductive effects of MSCs on cartila...

Tissue engineering scaled-up, anatomically shaped osteochondral constructs for joint resurfacing

“…In agreement with previous studies, a mixed co-culture of CCs and MSCs was found to enhance cartilage specific extracellular matrix (ECM) deposition in vitro (Acharya et al, 2012;Bian et al, 2011;Meretoja et al, 2012;Tsuchiya et al, 2004;Wu et al, 2011). Previous studies have demonstrated that this is due to MSCs secreting factors that drive proliferation of the CC population (Acharya et al, 2012;Wu et al, 2013;Wu et al, 2011). Co-culture led to the development of thicker, more homogeneous and more morphologically stable cartilaginous constructs in vivo (compared to corresponding stem cell only groups) that better integrated with the underlying osseous layer.…”

“…It has previously been speculated that suppression of hypertrophy is mediated, at least in part, by CCs secreting parathyroid hormone-related protein (PTHrP) (Fischer et al, 2010). In addition, a relative increase in the ratio of CCs to MSCs due to the latter cell type releasing factors that increase CC proliferation (Wu et al, 2013) would also be expected to reduce the over hypertrophic potential of the engineered tissue by increasing the ratio of phenotypically stable CCs to hypertrophic BMSCs. In addition, it has been reported that when BMSCs and CCs are co-cultured, the former cells die off over time (Meretoja et al, 2012;Wu et al, 2011), further increasing the ratio of CCs to BMSCs in the engineered tissue.…”

Metabolic programming of mesenchymal stromal cells by oxygen tension directs chondrogenic cell fate

“…This observation underlines the importance of the progenitor cell's trophic role in tissue regeneration. Trophic factors can mediate tissue regeneration in both direct and indirect manners (23)(24)(25). Progressive insight indicates that current culture and treatment protocols allow progenitor cells to contribute mainly to regenerative effects via trophic roles rather than direct tissue formation (26).…”

Engineering Cartilage Tissue by Pellet Coculture of Chondrocytes and Mesenchymal Stromal Cells

“…We also demonstrated that these observations present in coculture pellets of chondrocytes and MSCs are independent of donor variation and culture conditions [ 8 ]. Subsequent experiments indicated that increased secretion of fi broblast growth factor 1 (FGF1) in coculture of MSCs and chondrocytes is responsible for increased chondrocyte proliferation in pellet cocultures [ 9 ]. Thrombospondin-2 has also been reported to be secreted by MSCs to promote chondrogenic differentiation both in vitro and in vivo [ 10 ].…”