Fig. 2

Evalution of cartilage-specific components in rAAV-transduced human bone marrow aspirates in static versus dynamic culture conditions. Aspirates (150 μl) were prepared and transduced with rAAV-luc or rAAV-FLAG-hsox9 as described in Fig. 1 or let untransduced for maintenance in dynamic or static culture conditions for up to 28 days (n = 4 independent samples tested in triplicate per vector treatment and culture condition in three independent experiments) as described in the Methods. The samples were processed (a) for safranin O staining and to detect the expression of type-II collagen after 7, 21, and 28 days as described in the Methods (magnification x10, scale bars: 200 μm; insets: complete section, magnification x4; all representative data) and (b) to evaluate the gene expression profiles (SOX9 and COL2A1, with GAPDH serving as a housekeeping gene and internal control) after 21 days by real-time RT-PCR amplification as described in the Methods. Ct values were obtained for each target and GAPDH as a control for normalization, and fold inductions (relative to untreated aggregates) were measured by using the 2^-ΔΔCt method. Statistically significant relative to ^ano vector treatment, ^brAAV-luc application, and ^cstatic culture