Autologous bone marrow stromal cells loaded onto porous hydroxyapatite ceramic accelerate bone repair in critical-size defects of sheep long bones E. Kon, 1 A. Muraglia, 2 A. Corsi, 3 P. Bianco, 3 M. Marcacci, 1 I. Martin, 2 A. Boyde, 4 I. Ruspantini, 5 P. Chistolini, 5 M. Rocca, 1 R. Giardino, 1 R. Cancedda, 2,6 R. Quarto 2 1 Istituti Ortopedici Rizzoli, Bologna, Italy 2 Istituto Nazionale per la Ricerca sul Cancro, Centro di Biotecnologie Avanzate, Largo Rosanna Benzi 10, Genova, Italy 3 Dipartimento di Medicina Sperimentale, Universita’ dell’ Aquila, and Universita’ La Sapienza, Roma, Italy 4 Department of Anatomy and Developmental Biology, University College London, London, United Kingdom 5 Laboratorio di Ingegneria Biomedica, Istituto Superiore di Sanita `, Roma, Italy 6 Dipartimento di Oncologia, Biologia e Genetica, Universita’ di Genova, Italy Received 16 February 1999; revised 8 July 1999; accepted 6 August 1999 Abstract: The ability of marrow-derived osteoprogenitor cells to promote repair of critical-size tibial gaps upon au- tologous transplantation on a hydroxyapatite ceramic (HAC) carrier was tested in a sheep model. Conditions for in vitro expansion of sheep bone marrow stromal cells (BMSC) were established and the osteogenic potential of the ex- panded cells was validated. Ectopic implantation of sheep BMSC in immunocompromised mice led to extensive bone formation. When used to repair tibial gaps in sheep, cell- loaded implants (n = 2) conducted a far more extensive bone formation than did cell-free HAC cylinders (n = 2) over a 2-month period. In cell-loaded implants, bone formation was found to occur both within the internal macropore space and around the HAC cylinder while in control cell- free implants, bone formation was limited mostly to the outer surface and was not observed in most of the inner pores. As tested in an indentation assay, the stiffness of the complex HAC–bone material was found to be higher in cell- loaded implants compared to controls. Our pilot study on a limited number of large-sized animals suggests that the use of autologous BMSC in conjunction with HAC-based carri- ers results in faster bone repair compared to HAC alone. Potentially this combination could be used clinically in the treatment of extensive long bone defects. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res, 49, 328–337, 2000. Key words: bone marrow stromal cells; hydroxyapatite; bone reconstruction; orthopedic surgery; tissue engineering INTRODUCTION Multipotent precursor cells able to differentiate along the osteogenic and chondrogenic lineage and to give rise to skeletal tissues are found among the stro- mal cells of the postnatal marrow of mammals. 1 Upon isolation in culture, these cells give rise to discrete colonies of fibroblast-like cells (hence their designa- tion as colony-forming unit-fibroblastic, CFU-f). These cells can be thoroughly expanded and assayed for their osteogenic and chondrogenic potential either in vitro, using appropriate sets of phenotypic markers, or in vivo by means of defined transplantation assays. 2–4 The physiological significance of this unique cell population rests with the multiple pathways of tissue turnover and repair in which they are postulated to be involved, as critical effectors, in the postnatal organ- ism. Hence, their discovery has prompted increasing attention over the past few years as important poten- tial tools for the development of novel therapeutic ap- proaches in a variety of skeletal disorders, including somatic cell and gene therapy. 5–8 In orthopedic sur- gery, a cell-based therapeutic approach could be de- sirable in all instances calling for extensive bone re- construction. Although a number of biomaterial-based approaches have been developed and used with vari- able success to date, 1,9 none has proven ideal so far. Among options attractive in terms of quality of re- sults, cost effectiveness, and risk-benefit balances, hy- Correspondence to: R. Quarto; e-mail: quarto@ermes. cba.unige.it Contract grant sponsor: Associazione Italiana Ricerca sul Cancro Contract grant sponsor: Agenzia Spaziale Italiana Contract grant sponsor: Istituto Superiore di Sanita `, Roma, Italy © 2000 John Wiley & Sons, Inc. CCC 0021-9304/00/030328-10