Introduction

Mesenchymal stem cells (MSCs) have long been proposed as a potential cell source for musculoskeletal regeneration. Within the cartilage field there have been several efforts to develop novel therapeutic strategies using these cells (Johnstone et al., 2013). Broadly speaking MSC therapies can be split into two categories; those that use freshly isolated cells, and those that use monolayer expanded cells. This distinction brings significant ramifications, both in terms of regulatory requirements and in the mechanism of action. From a scientific perspective, there have been two main challenges that have delayed the translation of MSCs into the clinic. Insufficient tools to adequately characterize the cell populations obtained and a lack of a detailed understanding of the mechanism of action of the implanted cells.

Content

Naïve cells are already commonly utilized in the clinic in the form of marrow stimulation techniques or intra-operative cell transfer of bone marrow aspirate concentrates (BMAC). The ease of application has been a major driver although it is widely accepted that the repair tissue produced often has inferior properties due to its fibrocartilage nature (Farr et al., 2011). This in itself is not an impediment to their use, as delaying the need for a prosthetic joint is still a great advantage as years of fitness and life expectancy increase.

Monolayer expansion of adherent MSCs is an attractive alternative strategy due to the increase in numbers than can then be implanted. This process is fraught with difficulties as it is known the expansion process can lead to major changes in cellular function (Bara et al., 2014). Whether this is due to changes in cell behavior during expansion (phenotypic drift), or whether it is due to the expansion conditions preferentially selecting for a subpopulation is unclear with data suggesting both may be possible. Multicolor barcode lentivirus labelling has shown that a smaller number of clones become dominant during expansion (Selich et al., 2016). While adapting monolayer expansion conditions during the last few days of culture can also prime cells to be more receptive to differentiation. This particular issue has been highlighted in an article by Arnold Caplan that postulates monolayer expanded cells do not behave as naïve MSCs and raises the question as to whether tripotency, a commonly used MSC assessment, is actually an in vitro artifact (Caplan, 2017).

The use of accurate markers to identify the cells being studied is the central issue around most other topics revolve. With no marker being truly MSC specific, consensus and reproducibility is almost impossible to achieve. This compounds the problem that phenotype and function of monolayer expanded cells will vary depending on several factors, such as serum batch, making comparisons between laboratories difficult. Furthermore, markers predictive of MSC function are lacking. Commonly used CD markers do not provide an indication of function which limits their use (Sacchetti et al., 2016). Work in this area is increasing from our laboratory and others (Dickinson et al., 2017; Loebel et al., 2015) and this will hopefully lead to new assays being developed that can predict potency of individual donors.

The use of monolayer expanded cells is hindered by a number of still open questions. Are undifferentiated or predifferentiated cells preferable in a clinical setting? Does monolayer expansion under 21% oxygen select for a population of cells that are more susceptible to cell death when placed in the hypoxic injured joint? Is the main mechanism of action cell differentiation or paracrine activity? The answers to these questions are likely to be interrelated and more work needs to be done.

References

Bara JJ, Richards RG, Alini M, Stoddart MJ (2014) Concise review: Bone marrow-derived mesenchymal stem cells change phenotype following in vitro culture: implications for basic research and the clinic. Stem Cells 32: 1713-1723.

Caplan AI (2017) Mesenchymal Stem Cells: Time to Change the Name! Stem Cells Transl Med 6: 1445-1451.

Dickinson SC, Sutton CA, Brady K, Salerno A, Katopodi T, Williams RL, West CC, Evseenko D, Wu L, Pang S, Ferro de Godoy R, Goodship AE, Peault B, Blom AW, Kafienah W, Hollander AP (2017) The Wnt5a Receptor, Receptor Tyrosine Kinase-Like Orphan Receptor 2, Is a Predictive Cell Surface Marker of Human Mesenchymal Stem Cells with an Enhanced Capacity for Chondrogenic Differentiation. Stem Cells.

Farr J, Cole B, Dhawan A, Kercher J, Sherman S (2011) Clinical cartilage restoration: evolution and overview. ClinOrthopRelat Res 469: 2696-2705.

Johnstone B, Alini M, Cucchiarini M, Dodge GR, Eglin D, Guilak F, Madry H, Mata A, Mauck RL, Semino CE, Stoddart MJ (2013) Tissue engineering for articular cartilage repair--the state of the art. Eur Cell Mater 25: 248-267.

Loebel C, Czekanska EM, Bruderer M, Salzmann G, Alini M, Stoddart MJ (2015) In vitro osteogenic potential of human mesenchymal stem cells is predicted by Runx2/Sox9 ratio. Tissue Eng Part A 21: 115-123.

Sacchetti B, Funari A, Remoli C, Giannicola G, Kogler G, Liedtke S, Cossu G, Serafini M, Sampaolesi M, Tagliafico E, Tenedini E, Saggio I, Robey PG, Riminucci M, Bianco P (2016) No Identical "Mesenchymal Stem Cells" at Different Times and Sites: Human Committed Progenitors of Distinct Origin and Differentiation Potential Are Incorporated as Adventitial Cells in Microvessels. Stem Cell Reports 6: 897-913.

Selich A, Daudert J, Hass R, Philipp F, von Kaisenberg C, Paul G, Cornils K, Fehse B, Rittinghausen S, Schambach A, Rothe M (2016) Massive Clonal Selection and Transiently Contributing Clones During Expansion of Mesenchymal Stem Cell Cultures Revealed by Lentiviral RGB-Barcode Technology. Stem Cells Transl Med 5: 591-601.

Acknowledgments

The presented work has been supported by the AO Foundation and the Swiss National Science Foundation Grants 31003A_179438, 31003a_146375/1., 320000-116846/