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FRI0001 All stages of synovial mesenchymal stem cell activity including adhesion, proliferation, migration and chondrogenic differentiation are supported by human platelet lysate- implications for novel one stage joint regenerative procedures
  1. A Altaie,
  2. TG Baboolal,
  3. E Jones,
  4. O Wall,
  5. D McGonagle
  1. Leeds Institute of Rheumatic and Musculoskeletal Medicine, University of Leeds, Leeds, United Kingdom

Abstract

Background Both synovial and related synovial fluid-derived mesenchymal stem cells (SF-MSC) are highly proliferative and chondrogenic and have direct access to superficial cartilage injuries [1]. Human platelet lysate (PL) is being increasingly used for joint regenerative approaches [2,3] but the biological basis for in vivo effect, if any is lacking.

Objectives The study evaluated the effect of PL on adhesion, proliferation, migration and trilineage differentiation of synovial cavity derived MSCs in comparison to “gold standard” foetal calf serum (FCS) based stem cell media.

Methods MSCs were derived from joint cavity washout in patients undergoing anterior cruciate ligament reconstruction or meniscal repairs. Cells were plated in StemMACS (FCS media) or with DMEM Supplemented with 10% Stemulate (PL media, Cook Regentec). Standard MSC proliferation, trilineage differentiation assays, and transwell migration assays were performed for both conditions.

Results PL enhanced MSC colonies sizes (CFU-F) by 2.5 folds compared to FCS (p=0.0023, n=5). PL shortened MSC doubling times (p=0.0411, n=6). However, PL had no impact on CFU-F numbers (p=0.5973, n=5). In transwell migration assays, MSC migration was significantly increased toward 10%PL compared to 10%FCS (p=0.0286, n=6). Under standard chondrogenic induction conditions, PL-expanded MSC showed 30% more sulphated GAG (sGAG) production compared to FCS-expanded cells (n=4). In osteogenic and adipogenic induction, Alizarin red staining Ca++ assay and Nile Red showed no significant difference compared to FCS expanded cells (p=0.3429, n=4). Interestingly, replacing TGFbβ with 20% PL was as good as standard chondrogenic media and with increasing PL percentage sGAG production increased (n=4). In same time using PL with high glucose media in 1:1 ratio was efficient to produced sGAG as compared to negative control (media only) (p=0.0286, n=4, figure 1). Moreover, substituting FCS with PL in the osteogenic media resulted in increased Ca++ deposition in both PL and FCS expanded cells by 90% and 100% compared standard osteogenic media.

Conclusions This is the first study that assessed joint cavity MSC responses to PL. All aspects of joint cavity derived MSC biology towards cartilage repair were augmented in vitro with PL based media over “gold standard” MSC media. This work provides biological plausibility for the combination of PL with joint resident MSC towards endogenous joint repair strategies.

References

  1. Jones, E.A., et al., Synovial fluid mesenchymal stem cells in health and early osteoarthritis: detection and functional evaluation at the single-cell level. Arthritis Rheum, 2008. 58(6): p. 1731–40.

  2. Morito, T., et al., Synovial fluid-derived mesenchymal stem cells increase after intra-articular ligament injury in humans. Rheumatology (Oxford), 2008. 47(8): p. 1137–43.

  3. Ben Azouna, N., et al., Phenotypical and functional characteristics of mesenchymal stem cells from bone marrow: comparison of culture using different media supplemented with human platelet lysate or fetal bovine serum. Stem Cell Res Ther, 2012. 3(1): p. 6.

References

Disclosure of Interest None declared

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