Bone Abstracts

We have previously demonstrated the presence of circulating mesenchymal stem cells (MSCs) in fracture patients. However, the exact time for their appearance in circulation remains unknown. Nine patients (five females, four males, age 70±12 years, range 55–89 years) with a fresh femoral neck fracture were recruited in the study, which was approved by the local ethical committee. Bone marrow (BM) sample drawn from the iliac crest of all patients served as the individual MSC reference. Peripheral blood (PB) sample was drawn prior to fracture surgery and subsequently on days 1, 2, 3, 7, 14, and 42, corresponding the median times of 31, 46, 77, 99, 194, 363, and 1031 h after fracture. Mononuclear cells were successfully isolated from 8/9 BM samples and from 56/57 PB samples by Ficoll gradient centrifugation. Small colonies of plastic-adherent, fibroblast-like cells were found in BM samples of all patients and in PB samples of 6 patients. In PB samples, the earliest time-point for positive cultures of plastic-adherent cells after fracture was 26 h and the latest time-point 434 h. Cultures of these cells were expanded and cells were characterized for proliferation, colony formation and tri-lineage differentiation, as well as surface marker expression. No significant differences were observed between BM and PB derived cells in proliferation, colony formation or osteogenic differentiation. Both were also positive for CD105, CD73 and CD90 and negative for CD14, CD45 and CD34. Majority of PB samples, which were positive for MSCs were obtained within 4 days after fracture. Two patients had cells in two or more samples and four only in a single sample. In conclusion, MSCs appear in peripheral blood mainly within the first four days after fracture but individual variation exists. Catching and identification of MSCs proved to be tedious, making this kind of studies challenging.