Chondrogenesis of adipose stem cells in a porous polymer scaffold: influence of the pore size.

This study examined how the difference in pore size of porous scaffolds affected the in vitro chondrogenic differentiation of seeded adipose stem cells (ASCs) and the in vivo cartilage repair of ASC/scaffold construct. ASCs were isolated from 18 rabbits and seeded in a porous poly (ε-caprolactone) (PCL) scaffold with different pore sizes (100, 200, 400 µm). The ASCs underwent in vitro chondrogenic induction under TGF-ß2 and BMP-7 for 21 days before analysis. The ASC/scaffold construct was also implanted on the osteochondral defect created on the distal femur of the same rabbits, and the quality of cartilage regeneration was analyzed after 8 weeks. At day 21, the ASCs proliferated and spread on the surface of the scaffolds with a pore size 100 and 200 µm, whereas there were many lumps of conglomerated ASCs on those with a pore size of 400 µm. The DNA content was significantly lower in the scaffold with a pore size of 400 µm than in that with a pore size of 100 or 200 µm. Proteoglycan production was significantly greater in the scaffold with a pore size of 400 and 200 µm than in that with a pore size of 100 µm. The chondrogenic marker gene expression including SOX9 and COL2A1 was greatest in the scaffold with a pore size of 400 µm followed by 200 µm. Immunofluorescent imaging showed that, while SOX9 was localized to nucleus, type II collagen was observed on the cytoplasm and secreted matrix around the cells most abundantly in the scaffold with a pore size of 400 µm followed by 200 µm. The gross and histological findings from the osteochondral defects showed that the cartilage repair was better in the scaffold with a pore size of 400 and 200 µm than in that with a pore size of 100 µm.

Improved spinal fusion efficacy by long-term delivery of bone morphogenetic protein-2 in a rabbit model.

BACKGROUND AND PURPOSE: Various new delivery systems for recombinant human bone morphogenetic protein-2 (rhBMP-2) have been introduced to improve its efficacy in osteogenesis. Of these, we have previously developed heparin-conjugated PLGA nanospheres (HCPN) as a long-term delivery system for BMP-2. In vitro studies have shown that the BMP-2 long-term delivery system enhances the level of bone formation. However, the long-term effects of BMP-2 on spinal fusion have not been assessed. Therefore, we now tested the hypothesis that the long-term delivery of BMP-2 using HCPN improves spinal fusion compared to short-term delivery in a rabbit fusion model. METHODS: 24 adult New Zealand White rabbits underwent posterolateral fusion (6 animals in 4 groups). The autograft group received an autologous iliac chip bone graft as a positive control. The BMP-2-PN group received rhBMP-2 (20 µg per implant) and PLGA nanospheres (PN) suspended in fibrin gel, and served as a short-term release group. The HCPN group received HCPN suspended in fibrin gel without BMP-2 as a negative control. The BMP-2-HCPN group received rhBMP-2 (20 µg per implant)-bound HCPN suspended in fibrin gel and served as a long-term release group. All animals were killed 12 weeks after surgery. Manual palpation, axial tensile tests, radiography, and histological evaluations were then performed. RESULTS: The spinal fusion rate and Young's modulus of the fusion mass were better in the BMP-2 long-term delivery group than in the short-term delivery group at an equivalent dose. However, the outcome of the long-term delivery was inferior to that of the autograft group. INTERPRETATION: The HCPN system showed potential as an effective carrier that might improve the osteogenic efficacy of BMP-2 for spinal fusion.

Novel hyaluronate-atelocollagen/beta-TCP-hydroxyapatite biphasic scaffold for the repair of osteochondral defects in rabbits.

The authors devised a novel biphasic scaffold combining hyaluronic acid and atelocollagen for the chondral phase and combining hydroxyapatite (HA) and beta-tricalcium phosphate (beta-TCP) for the osseous phase. The biphasic scaffold was fabricated by placing the freeze-dried chondral phase over the HA/beta-TCP scaffold prewetted with hyaluronate/atelocollagen solution. Chondrocytes were isolated in 28 rabbits, expanded, injected inside the chondral phase of the biphasic scaffold, and then cultured in chondrogenic medium. After 2 weeks of in vitro culture, chondrocytes had evenly infiltrated inside the chondral phase and produced extracellular matrix. For in vivo study, a large osteochondral defect was made on the patellar groove of the right distal femur and managed using one of the following methods: filling with cell-biphasic scaffold composite (group I); implanting only biphasic scaffold (group II); placing the removed osteochondral fragments back into the defect (group III, positive control); leaving empty (group IV, negative control). Seven rabbits were allocated to each group. After 12 weeks, the International Cartilage Repair Society Macroscopic Score was highest in group III, followed by group I, group II, and lastly group IV. Depression of the defect was greatest in group IV. There were three rabbits (two in group I and one in group II) that were completely denuded of the chondral phase. The junction to adjacent native cartilage was distinct in rabbits of all groups. The International Cartilage Repair Society Visual Histological Score was highest in group III, followed by groups II and I, and lastly group IV. In conclusion, our results suggest that a biphasic osteochondral composite using a chondral phase consisting of hyaluronate and atelocollagen and an osseous phase consisting of HA and beta-TCP holds the promise for repair of osteochondral defects.