Tissue-Engineered Articular Cartilage Construct in Hand Surgery for Sub-mm Fractional Repair
Yingfang Fan, M.D., Ph.D., William A. Farinelli, BS, Samuel D. Zarfos, BS, Gem Runyan, BS, Ying Wang, M.D., Joshua Tam, Ph.D., R. Rox Anderson, M.D., Mark A. Randolph, MAS, Robert W. Redmond, Ph.D..
Massachusetts General Hospital, Boston, MA, USA.
PURPOSE: Articular cartilage lesions in large joints have been successfully treated with autologous chondrocyte implantation (ACI) with or without a matrix scaffold (e.g., MACI). However, lesions in small joints of the hand and wrist cannot be easily treated with current ACI techniques. Fractional treatment, either mechanically or by laser, to make perforations into skin has been shown promote regeneration. The purpose of this study was to develop a new strategy that combines fractional treatment with our method for generating new cartilage matrix using dynamic Self-Regenerating Cartilage (dSRC) to treat cartilage lesions in the joints of the hand and wrist.
METHODS: 10e7 freshly harvested autologous swine chondrocytes were cultured on a rocker for 14 days at 37 degree C to form dSRC. The dSRC was then mixed with collagen gel and placed into mechanically punched holes or laser (wavelength 10.6 Ám) drilled holes at the center of a swine articular cartilage disc and capped with collagen gel. The control group consisted of isolated chondrocytes (3x10e7 cells per mL) injected into punched or laser drilled holes. Four groups were tested - (1) dSRC + 0.3 mm laser hole, (2) dSRC + 0.3 mm punch hole, (3) chondrocytes + 0.3 mm laser hole, and (4) chondrocytes + 0.3 mm punch hole. The cartilage discs were cultured in medium at 37 degree C for 8 weeks (n=6/group) then evaluated. In a pilot, in vivo study, the dSRC were encapsulated in collagen hydrogels and placed into 0.3 mm diameter punched and laser drilled holes in swine articular cartilage discs and capped with hydrogel. These discs were implanted subcutaneously in female nude mice. Constructs were harvested at 8 weeks. All constructs were evaluated histologically and immunohistochemically for cartilage formation and integration with native cartilage.
RESULTS: After 8 weeks in vitro, dSRC-treated constructs generated contiguous new cartilage matrix, compared to isolated chondrocyte-filled constructs that showed only pericellular matrix formation. dSRC groups demonstrated intense staining with Safranin-O and Toluidine blue stains indicating high glycosaminoglycan (GAG) production when compared to faint staining of groups treated with isolated chondrocytes. After 8 weeks in vivo in mice, contiguous cartilage matrix was observed in both dSRC filled punched holes and CO2 laser-drilled holes. Immunohistochemical staining further confirmed that the matrix of dSRC group was typical of normal hyaline cartilage, rich in collagen type II and no collagen type I, similar to native cartilage. Results of this study demonstrate that dSRC capped with hydrogels can successfully engineer contiguous articular cartilage matrix in both mechanical and fractional laser created environments.
CONCLUSIONS: Fractional treatment combined with dSRC demonstrates successful hyaline cartilage formation and integration with native cartilage both in vitro and in vivo. Such a strategy could be employed to translate ACI techniques commonly used in the knee to treat cartilage defects or osteoarthritis in the smaller joints of the hand and wrist.
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