Introduction: Until recently adult stem cells were presumed to be committed to differentiation of specific tissues. Adult hematopoietic stem cells (HSCs, CD34+) for example, originally believed to be limited to hematopoiesis are capable of transdifferentiation to generate cells of different lineages. This capability is referred to as stem cell plasticity. Studies in cardiac and peripheral vascular disease and nonunions and osteonecrosis in orthopedics have demonstrated that concentrated bone marrow is an effective and safe method of treatment. The present study evaluated a methodology to concentrate a small sample of bone marrow at point of care to compare with described techniques.
Methods: Sixty or 120 mL of bone marrow was withdrawn from the posterior iliac crest. The concentration process utilized the standard SmartPReP-2/DePuy Symphony Centrifuge (Harvest Technologies, Plymouth, MA). The shape and density of the floating shelf was modified to enhance collection of nuclear cells. The bone marrow was analyzed for cell counts, morphology, and flow cytometry. Hematopoietic stem cells (CD34+) were used as a marker for stem cell concentration. Bone marrow stem cells were cultured using specialized media supplements. The systems were also compared using the hind limb ischemia (HLI) model.
Results: Using the Harvest BMC System™ system, the results for the Colony Forming Unit (CFU’s) Analysis were as follows (Mean ± S.D.): the aspirate volume: 120 mL, CFU’s/cm3: 3040±1251, BMC volume delivered: 20mL, and Progenitor cells delivered: 60,800±29,200.
The cultures demonstrated viable hMSCs that were identical to a commercially available cell line. The cultures were transferred into osteogenic media; after 10 days the bone marrow derived cells and the commercial cell lines were stained with Von Kossa silver stain and for alkaline phosphatase demonstrating osteoblastic differentiation.
Hind limb ischemia studies have demonstrated that laser doppler blood flow was significantly better following BMAC infusion as compared to cells concentrated with Ficoll. These results were confirmed by a Boyden chamber migratory assay.
Discussion: A bone marrow concentrate can be prepared at point of care within 15 minutes of collection. The Harvest BMAC system is capable of producing a concentration of stem cells equivalent to or greater than those used in successful clinical studies. Successful clinical results can be obtained using one-third (1/3) of the aspirate volume required by other methods. Ongoing clinical and animal studies are confirming its clinical application.
