Abstract
Introduction
Iliac crest bone marrow aspirate (ICBMA) is frequently cited as the ‘gold-standard’ source of MSCs. MSCs have been shown to reside within the intramedullary (IM) cavities of long-bones [Nelea, 2005] however a comparative assessment with ICBMA has not yet been performed and the phenotype of the latter compartment MSCs remains undefined in their native environment.
Methods
Aspiration of the IM cavities of 6 patients' femurs with matched ICBMA was performed. The long-bone-fatty-bone-marrow (LBFBM) was filtered (70μm) to separate liquid and solid fractions and the solid fraction was briefly (60min, 37oC) digested with collagenase. MSC enumeration was performed using the colony-forming-unit-fibroblast (CFU-F) assay and quantification of cells with the CD45low CD271+ phenotype by flow-cytometry. [Jones 2002, Buhring 2007] MSCs were cultured and standard expansion media and passage 2 cells were differentiated towards osteogenic, adipogenic and chondrogenic lineages.
Results
ICBMA and LBFBM-liquid had similar median frequencies of MSCs/CFU-Fs per 200μl of sample (median 8, range 2-21, and 14, 0-53, respectively). LBFBM-solid fraction contained ∼10-fold more MSCs (116, 23-171). Correspondingly, LBFBM-solid fraction contained the highest proportions of CD45low CD271+ cells (median 0.315, 0.221-0.874) compared with (0.052, 0.023-0.083 and 0.152, 0.063-0.956) for ICBMA and LBFBM-liquid respectively (N =4) - thus sharing this phenotype of ICBMA cells. LBFBM MSCs were negative for the CD34 marker which has previously been reported on extra-bone marrow fat sources [Lin 2008]. MSCs isolated from the LBFBM phases were equivalent to ICBMA in terms of their osteogenic, chondrogenic or adipogenic potencies and their phenotypes following expansion was consistent with MSCs (CD73+ CD90+ CD105+CD33-CD34- CD45-).
Conclusions
Intra-medullary cavities of long-bones are frequently accessed by the orthopaedic/trauma surgeon. Aspiration of LBFBM prior to reaming insertion of prosthesis/nail is a ‘low-tech’ method of harvesting large numbers of MSCs that can be liberated by brief enzymatic digestion. LBFBM can be used as an alternative to ICBMA for autologous use in trauma surgery. Both the in vivo phenotypes and functionality of LBFBM and ICBMA are comparable.
