Tissue repair cells (TRCs) are CD90+ mesenchymal stem cell-enriched bone marrow-derived cells. TRC characteristics: at least 30 × 106 mononuclear cells, 5–26% CD90+/CD14+ cells, 9.6–48.5% CD14+ cells, 71.2–81.8% CD45+ cells and 0–33.3% CD105+/CD14+/CD45+ cells.
Isolation of bone marrow mononuclear cells (BMCs) is performed at room temperature for 75 min and 400 × g, followed by two washing cycles with human serum albumin-supplemented normal saline, resulting in a 50 ml suspension of BMCs. To increase the number of stem and early progenitor cells purified from bone marrow, the Aastrom Replicell® System, (ARS) can be used, incorporating single-pass perfusion in which fresh medium flows slowly over cells without retention of waste metabolites or differentiating cytokines. After cultivation for 12 days, with a ramped continuous medium perfusion schedule, the cell product, termed ‘tissue repair cells (TRCs)', including both non-adherent and mostly adherent cells, is harvested by trypsinisation and collected into a harvest bag. The culture medium consists of Iscove’s Modified Dulbecco’s Medium (IMDM), foetal bovine serum (10%), horse serum (10%), hydrocortisone (5 μM), gentamycin sulfate, and vancomycin. Cells are cultured at 37 °C and 5% CO2.