Background Bone has a capacity for self-repair without scar formation. Most fractures heal spontaneously or with the help of surgical procedures. However, despite the inherent ability of the bone to regenerate itself, there are a number of clinical situations in which complete bone healing fails to occur and requires cell therapy.
Objectives The purpose of this study was to determine the synergistic effect of ASCs added to BMSCs for osteogenic differentiation and angiogenesis, and to find the optimal ratio of ASCs to BMSCs to promote these desired functions.
Methods In this study, we used transwell and mixed cocultures for the in vitro model and the subcutaneous ectopic ossification in nude mice for the in vivo model. In vitro segregated cocultures using transwell were carried out for 14 days in 4 ways using BMSCs and ASCs in passage 3 : #1, 1 x 105 BMSCs and 0.25 x 105 ASCs; #2, 1 x 105 BMSCs and 0.5 x 105 ASCs; #3, 1 x 105 BMSCs and 0.75 x 105 ASCs; #4, 1 x 105 BMSCs and 1 x 105 ASCs. In vitro mixed cocultures were also performed in the same proportion. For in vivo analysis, cells were seeded in PLGA scaffold and implanted on the subcutaneous tissue of 20 nude mice in 4 ways and analyzed after 5 weeks: 1) without seeded cells; 2) seeded with 1 x 105 BMSCs; 3) seeded with 1 x 105 BMSCs and 0.5 x 105 ASCs; and 4) seeded with 1 x 105 ASCs.
Results From the transwell culture, 1 x 105 BMSCs cultured with 5 x 105 ASCs showed significantly greater osteogenic differentiation and mineralization as shown by alkaline phosphatase (ALP) activity and calcium deposition than BMSCs alone. In the mixed coculture model, ASC/BMSC coculture at a ratio of 0.5/1 showed a significantly greater level of ALP activity and calcium deposition as well as greater gene and protein expression of osteogenic markers including COL1A, osteocalcin, and bone sialoprotein compared with BMSCs alone. The mixed ASC/BMSC coculture at a ratio of 0.5/1 showed the highest level of vascular endothelial growth factor which was significantly greater than BMSCs alone, comparable to ASCs alone. The HUVEC tube formation assay also demonstrated that the mixed ASC/BMSC coculture of 0.5/1 enhanced tube formation to a level similar to ASCs alone. In vivo implantation studies demonstrated that PLGA-ASCs-BMSCs showed a greater amount of CD31-positive microvessel formation than PLGA-BMSCs, comparable to that of PLGA-ASCs. PLGA-BMSCs and PLGA-ASCs-BMSCs had similar degrees of calcification, but were greater than that of PLGA-ASCs.
Conclusions ASCs added to BMSCs promoted osteogenesis and angiogenesis with the optimal ASCs/BMSCs ratio of 0.5/1.
Einhorn, TA.(2005) “The science of fracture healing.” J Orthop Trauma 19: S4-6.
Disclosure of Interest None Declared