gms | German Medical Science

Objectives: Recombinant adeno-associated virus gene vectors (rAAV) are promising vehicles to treat focal cartilage defects when delivered via biomaterials [1]. The objective was to test whether rAAV carrying the reparative sox9 and TGF-β genes in an alginate hydrogel can stimulate the reparative activities in human chondral defects as treatments for cartilage repair.

Methods: rAAV-lacZ carrying the E. coli reporter β-galactosidase gene, rAAV-FLAG-hsox9 a human FLAG-tagged sox9 sequence, and rAAV-hTGF-β the human transforming growth factor beta 1 cDNA (all under the CMV-IE promoter) were routinely prepared [2], [3] and mixed in alginate (10 μl rAAV/0.2% alginate AlgPH155 per defect) as described [2]. Human articular cartilage biopsies (n = 9; 6-mm diameter) were used to create chondral defects with a 2-mm biopsy punch and placed in DMEM, 10% fetal bovine serum, 100 U/ml penicillin, and 100 μl/ml streptomycin as described [3] before application of the rAAV/alginate hydrogels for 21 days (duplicates in three experiments) [3]. The deposition of matrix proteoglycans and type-II collagen was monitored on tissue sections by safranin O staining and immunodetection, respectively, and scored using a modified Bern score grading system [2], [3]. Two-way ANOVA and multivariate analysis were employed with P ≤ 0.05 considered statistically significant.

Results and conclusion: Alginate hydrogel-guided rAAV sox9 and TGF-β delivery significantly increased the reparative activities in the non-calcified cartilage and calcified cartilage of the defects over time (21 days) versus control conditions (lacZ or absence of vector) as seen by an increased deposition of matrix proteoglycans and type-II collagen (always P ≤ 0.05). Significant differences were noted between the rAAV-FLAG-hsox9/alginate and rAAV-hTGF-β/alginate hydrogels and the control conditions (lacZ or absence of vector) and between the non-calcified cartilage, calcified cartilage, and subchondral bone (always P ≤ 0.05).

Delivering rAAV sox9 and TGF-β gene vectors via an alginate hydrogel can durably stimulate the reparative (anabolic) activities in human chondral defects as a novel off-the-shelf therapeutic strategy for cartilage repair.

Figure 1 [Fig. 1]