Concentration of Connective-Tissue Progenitors Useful in Bone Marrow Grafting
Dr. Muschler is an orthopaedic surgeon specializing in adult reconstruction and treatment of fracture nonunions. He also heads the Orthopaedic Research Center's Bone Biology Laboratory in the Department of Biomedical Engineering.
Classic bone grafting techniques are used in over 1,000,000 procedures each year in the United States. Autogenous cancellous bone graft remains the gold standard for graft material. However, the harvest of autogenous bone is associated with significant cost in terms of postoperative pain, blood loss, infection, the need for a separate incision over the iliac crest, and possible fracture. Also, the amount and availability of autogenous bone may be insufficient for many clinical problems.
As a result, strategies to replace autogenous cancellous bone grafts are being sought. In the Cleveland Clinic's Department of Biomedical Engineering and Orthopaedic Research Center, we are researching methods to optimize the biology of stem cells for clinical orthopaedic applications. Using advanced tissue-engineering technology, we hope to improve the efficacy and reduce the morbidity of clinical bone grafting.
Bone marrow contains at least two populations of stem cells. The hematopoietic stem cell produces progeny that can reconstruct all blood-cell lineages, including immune cells, red blood cells and platelets.
A less known, but increasingly well-characterized, stem-cell population is the connective-tissue progenitor cell population (CTPs), which can give rise to bone, cartilage, muscle, fat, tendon, ligament, and even cardiac muscle and neurons. As precursors of bone-forming cells, CTPs are key contributors to bone formation and repair.
(Above) Microscopic view of a colony of osteoblastic cells shows bone formation in the center (dark area) and cells continuing to grow around the periphery (pink). The cells are expressing a protein marker for a primitive bone cell, endoglin (CD105).
(Above) These connective-tissue progenitor cells (CTPs) stain green, showing that they express CD105. This cell-surface protein serves as a marker for many primitive cells contributing to skeletal and soft-tissue healing.
A fundamental biological principle in the formation, repair or regeneration of any tissue is that the cells do all the work. No prosthetic surface, growth factor, medication or nutrient supplement can influence bone formation except through its specific effects on the activation, proliferation, migration, differentiation or survival of CTPs or their progeny.
Osteoconductive materials can provide a matrix or scaffold that facilitates the attachment, migration and proliferation of bone-forming cells. Biological stimuli from selected growth factors can be used to induce proliferation, migration and differentiation of CTPs through osteoinduction. However, optimal bone healing requires a sufficient number of CTPs, a component lacking in many complex wound sites.
Optimizing the concentration and delivery of bone-marrow-derived cells and CTPs can reduce the morbidity and increase the efficacy of clinical bone grafting. One alternative to autogenous cancellous bone grafts is to supplement allograft matrix from bone banks with aspirated bone marrow to augment the number of connective-tissue progenitors. Bone marrow can be aspirated from the iliac crest with minimal morbidity.
Work done in our laboratory has provided surgeons with guidance on optimizing the concentration of CTPs by limiting aspiration volume to 2 mL from each individual needle site. This significantly reduces dilution of these cells by peripheral blood.
However, our review of 120 consecutive bone grafting procedures found that bone marrow grafts combining simple bone marrow aspiration with allograft matrix were not as effective as autogenous cancellous bone grafts. This was particularly true in the treatment of fracture nonunions or segmental long-bone defects. Therefore, while bone marrow aspiration has value, further improvement is needed to make this strategy useful in complex grafting procedures.
Building on these observations, Cleveland Clinic orthopaedic surgeons and biomedical engineers have pioneered methods to increase the concentration and selection of CTPs from bone marrow to enhance bone graft performance. Clinically applicable techniques for rapid concentration and selection of CTPs from bone marrow involve using the surface of an implantable porous matrix as an affinity column to which CTPs selectively attach. Composite cellular grafts enriched 4- to 16-fold in CTPs can be prepared in only 20 minutes, and have improved bone formation and spinal fusion in animal models.
Clinical trials using enriched cellular grafts to replace autogenous cancellous bone grafts are now under way in our Department of Orthopaedic Surgery. Members of the Section of Spine Surgery have evaluated 15 patients undergoing anterior lumbar interbody fusion; ongoing clinical and radiographic evaluation of this approach has been satisfactory, with no suspected failures. Other methods to further improve the concentration and selection of CTPs for use in regeneration of bone and other tissues are being investigated. One possibility being studied is that bone marrow aspiration from the vertebral body during pedicle screw placement may serve as an alternative source for enriched composite bone graft preparations.
Success Rate of Bone Grafting Procedures by Clinical Setting and Graft Technique (ACB = Autogenous cancellous bone; BM = bone marrow; OP-1 = Growth Factor OP-1)
