MICROENVIRONMENTAL INFLUENCES ON ARTICULAR CHONDROCYTE DIFFERENTIATION

Abstract

Background:Immature articular cartilage (a.k.a. articular epiphyseal cartilage complex, AECC), serves dual functions as an articulating layer and surface growth plate. The AECC originates from an uninterrupted cartilaginous template during embryogenesis, presumably stemming from the same pool of progenitors, but it is unclear how the articulating layer is maintained throughout life, while the surface growth plate undergoes endochondral ossification.Hypothesis:The function and fate of chondrocytes in the AECC are not predetermined in the embryo, but instead can be influenced during development by biochemical and/or biomechanical stimuli in the cellular microenvironment.Methods:Microenvironments of the articulating layer and surface growth plate were switched in an in vivo goat model. Twelve 3-week-old goats were used. Osteochondral autografts were harvested from medial and lateral condyles and trochlea of the distal femur. Subchondral bone was removed (Figures 1A and 1B). Grafts were inverted and press-fit into select sites, which were full (condyles) or low (trochlea) weightbearing (Figures 1C through 1E). Animals were allowed ambulation and range of motion as tolerated. Two goats each were euthanized at 1, 2, 3, 6, 12, and 24 weeks postoperatively. Distal femurs were harvested and grafts were analyzed with trichrome staining and in situ hybridization for Col10, Col2, and Prg4.Results:All animals survived surgery, ambulated within 24 hours, and none exhibited signs of surgical site infection. Macroscopically, grafts at the trochlea and lateral condyle maintained a more cartilaginous appearance than those at the medial condyle, but all eventually developed an osseous appearance. Histologically, there was healing over time, with early grafts often containing areas of hemorrhage and occasionally necrosis, followed by formation of granulation tissue and subjacent myelofibrosis, with eventual invasion and ossification. At later timepoints, there was better graft incorporation at the articular surface without replacement by fibrosis at the medial condyle (Figure 2). In situ hybridization demonstrated some Prg4 positive cells and a decrease in Col10 expression at the articular surface (Figure 3).Conclusions:The synovial microenvironment may play a protective role against endochondral ossification. Some incorporation of epiphyseal cartilage into articular cartilage occurred at sites subjected to both compressive and shear forces. Data suggest the existence of local cytokines and/or matrix effects on chondrocytes that stimulate different cell functions and fates. A limitation of this study is its observational nature. Further understanding of articular chondrocyte differentiation and identification of key biochemical and/or biomechanical stimuli hold therapeutic promise for articular cartilage regeneration, repair, and replacement.Figure 1A.through 1E. Osteochondral autografts measuring 4.75 x 7 mm harvested from donor (lateral condyle and distal trochlea) and recipient (medial condyle and proximal trochlea) sites at the distal femur (Figure 1A). Separation of subchondral bone from the AECC (Figure 1B). Grafts inverted and inserted at select sites (Figures 1C through 1E). Experimental control with 13 gauge bone biopsies harvested and inserted in original orientation (Figures 1C through 1E).Figure 2.Histology with trichrome staining showing the articular surface of osteochondral autografts transplanted to the medial femoral condyle at 1, 2, 3, 6, 12, and 24 weeks postoperatively.Figure 3. In situ hybridization of Col2, Col10, and Prg4 gene expression of an osteochondral autograft transplanted to the femoral trochlea at 3 weeks postoperatively.