Point-of-care treatment of focal cartilage defects with selected chondrogenic mesenchymal stromal cells-An in vitro proof-of-concept study.

Due to the poor self-healing capacities of cartilage, innovative approaches are a major clinical need. The use of in vitro expanded mesenchymal stromal cells (MSCs) in a 2-stage approach is accompanied by cost-, time-, and personnel-intensive good manufacturing practice production. A 1-stage intraoperative procedure could overcome these drawbacks. The aim was to prove the feasibility of a point-of-care concept for the treatment of cartilage lesions using defined MSC subpopulations in a collagen hydrogel without prior MSC monolayer expansion. We tested 4 single marker candidates (MSCA-1, W4A5, CD146, CD271) for their effectiveness of separating colony-forming units of ovine MSCs via magnetic cell separation. The most promising surface marker with regard to the highest enrichment of colony-forming cells was subsequently used to isolate a MSC subpopulation for the direct generation of a cartilage graft composed of a collagen type I hydrogel without the propagation of MSCs in monolayer. We observed that separation with CD271 sustained the highest enrichment of colony-forming units. We then demonstrated the feasibility of generating a cartilage graft with an unsorted bone marrow mononuclear cell fraction and with a characterized CD271 positive MSC subpopulation without the need for a prior cell expansion. A reduced volume of 6.25% of the CD271 positive MSCs was needed to achieve the same results regarding chondrogenesis compared with the unseparated bone marrow mononuclear cell fraction, drastically reducing the number of nonrelevant cells. This study provides a proof-of-concept and reflects the potential of an intraoperative procedure for direct seeding of cartilage grafts with selected CD271 positive cells from bone marrow.