Autologous Platelet Lysate Does Not Enhance Chondrogenic Differentiation of Equine Bone Marrow-Derived Mesenchymal Stromal Cells Despite Increased TGF-β1 Concentration.

Abstract

Bone marrow-derived mesenchymal stromal cells (BM-MSCs) are being investigated for their potential in the treatment of musculoskeletal injuries, including tendon and ligament lesions, and cartilage lesions. Culture expansion of cells has traditionally been performed in medium supplemented with fetal bovine serum (FBS), however, concerns regarding the antigenicity and potential viral or prion contamination of FBS have prompted interest in alternative medium supplements. Platelet lysate (PL) contains elevated concentrations of growth factors, including transforming growth factor-β (TGF-β), platelet-derived growth factors, and fibroblast growth factor, released from the α-granules of platelets; therefore, PL could be an ideal medium supplement. The effect of PL on mesenchymal stromal cell (MSC) growth and differentiation has not been fully elucidated. We hypothesized that PL medium would contain significantly higher amounts of TGF-β1 than FBS medium and would be associated with enhanced osteogenic and chondrogenic differentiation. MSCs were isolated from bone marrow collected from five adult horses. Cells were cultured in traditional medium supplemented with FBS or in medium supplemented with fibrinogen depleted-PL (FD-PL). Immunophenotyping was performed using flow cytometry. Trilineage differentiation was assessed through histology and gene expression analysis using quantitative reverse transcription-polymerase chain reaction. TGF-β1 was quantified in both medium types. The immunophenotypes of BM-MSCs cultured in FBS and FD-PL medium were similar with both culture types containing cells positive for stromal cell markers [cluster of differentiation 29 (CD29), CD44, CD90, CD105, and major histocompatibility complex I (MHCI)] and negative for exclusion markers (CD45, CD79α, and MHCII). Despite significantly higher TGF-β1 concentration in FD-PL medium, chondrogenic and osteogenic differentiation were not significantly different between FBS and FD-PL supplemented cultures. PL is an appropriate alternative medium supplement for the culture of equine BM-MSCs up to passage 3. However, despite increased TGF-β1 concentration in FD-PL medium, significant changes in chondrogenic differentiation compared with FBS medium should not be expected.