Abstract
Top of pageAbstract The transfer of therapeutic genes may have potential value in enhancing the reparative activities of human articular cartilage. We tested the hypothesis that overexpression of sox9 via rAAV leads to the production of a transcription factor capable of influencing the tissue architecture and cell phenotype in normal human and osteoarthritic articular cartilage explants in situ. A human sox9 cDNA tagged by a FLAG sequence was cloned in rAAV (rAAV-FLAG-hsox9). Explants from normal human (tumor surgery) and osteoarthritic articular cartilage (total knee arthroplasty; Mankin score 7-9) were transduced by rAAV-FLAG-hsox9 or rAAV-lacZ and processed after 10 days to monitor transgene expression, for histological and morphometric analyses (safranin O, hematoxylin eosin, type-II collagen), and to measure the proteoglycan, type-II collagen, and DNA contents. Morphometric measurements were performed along the explant surface (AnalySIS and Scion Image programs). Safranin O staining intensity is the ratio positively stained surface to the total surface examined. Type-II collagen staining intensity is in pixels per standardized area. Dose-dependent transgene expression (b-gal, FLAG, sox9) was seen in the explants. Treatment by rAAV-FLAG-hsox9 did not modify the cell densities in the explants nor the DNA contents, at any vector dose employed. In contrast, a dose-dependent increase in safranin O and type-II collagen staining intensities was noted when applying rAAV-FLAG-hsox9, as well as an augmentation of the proteoglycan and type-II collagen contents. The data show that sox9 overexpressed in normal human and osteoarthritic articular cartilage explants via rAAV promotes type-II collagen and matrix synthesis in situ, suggesting that rAAV may have value in enhancing the reparative properties of injured articular cartilage.
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