Article
Functionalized and via 2-photon-polymerization (2PP) nanostructured biphasic implants as a treatment for circumscript osteochondral lesions in an ovine model
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Published: | October 5, 2015 |
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Objectives: Treatment of large osteochondral lesions in weight bearing regions of a joint with osteochondral autograft transfer has the major disadvantage of donor site morbidity. An alternative and possible solution to this problem is the usage of synthetic biphasic implants. In this study functionalized biphasic implants with an osseous phase consisting of hydroxyapatite-granules and a via 2-photon-polymerization (2PP) technique nanostructured cartilaginous phase were prospectively tested to treat above-mentioned lesions in sheep.
Methods: Biopsies of articular cartilage were taken from 14 sheep and cultured in vitro. After 6 weeks osteochondral defects with a size of 5.5 mm in diameter and 8.5 mm in depth were surgically created in the weight bearing femoral condyle as well as in the non-weight bearing trochlear groove of the knee. Prior to press-fit insertion of the biphasic implants, calcium phosphate nanoparticles with Dickkopf-1 (DKK-1) siRNA were added into the defect. In-vitro cultured chondrospheres were added to the apical layer of the implant. The animals were euthanized after 6 and 12 weeks and the implants were collected for molecular and cell biological analysis. Histology, immunohistochemistry and transmission electron microscopy (TEM) were used for cell biological analysis. Molecular biological analysis was performed separately for the osseous and cartilaginous phase using real-time PCR. Statistical evaluation was executed with SPSS Statistics using Kruskal-Wallis and Mann-Whitney test.
Results and Conclusion: Molecular biological analysis of the cartilaginous phase revealed a significantly higher expression of SOX-9, a transcription factor in chondrogenesis, in those implants with added chondrospheres (p = 0.018). The expression of type II collagen, the basis for hyaline cartilage, was detectable. However, there was no significant difference between the implants. Regarding the osseous phase a significantly higher expression of eNOS (p = 0.011), a marker for angiogenesis, and type I collagen (p = 0.008), a marker for bone formation, was measured for weight bearing samples. These results correlate well with the histological findings. After 6 weeks hydroxyapatite granules in the osseous phase were surrounded by granulation tissue. New bone tissue was formed after 6 weeks in non-weight bearing and after 12 weeks in weight-bearing samples. Analysis of the cartilaginous phase revealed type II collagen immunopositive granulation tissue. Regeneration of cartilage tissue was found to be further advanced in non-weight bearing samples.
The presented findings support the concept of 2PP nanostructured biphasic implants as a suitable treatment for large osteochondral lesions. The densely packed hydroxyapatite granules built a well integrated osseous phase and provided an appropriate fixation for the cartilaginous phase. Furthermore, molecular and cell biological analysis indicate the tremendous effect of eNOS expression and angiogenesis on both ossification and cartilage regeneration.