Modulation of endochondral development of the distal femoral condyle by mechanical loading

Abstract

Although previous theoretical modeling studies have predicted that various mechanical stresses accelerate or inhibit the ossification process of the neonatal chondroepiphysis, there is a paucity of experimental data to verify these models. The present study was designed to provide experimental evidence on whether the ossification of the chondroepiphysis is modulated by mechanical loading on the distal femoral condyle explant of the neonatal (5-day-old) rabbit in organ culture. Upon aseptic dissection, the right condyle explant was immersed in and fixated to an organ culture system, and received cyclic forces at 200 mN and 1 Hz for 12 h (N=8) directly on its slightly convex articular surface, whereas the contralateral, left condyle explant was immersed separately in organ culture (N=8). Subsequently, both loaded and control explants were placed in a bioreactor rotating at 20 rpm for 72 h. In each mechanically loaded specimen, a structure reminiscent of the secondary ossification center (SOC) appeared with an average area of 1.17+/-0.13 mm(2), or 15.2+/-8.2% of the total epiphysis area. In contrast, no SOC was detected in any of the unloaded contralateral control specimens. The SOC in mechanically loaded specimens was stained intensively with fast green, whereas either the rest of the loaded epiphysis or the entire control epiphysis was stained intensely to safranin-O but lacked fast green staining. Immunolocalization revealed that the SOC of the mechanically loaded specimens expressed Run x 2 and osteopontin, both of which were absent in the unloaded control specimens. Type X collagen was expressed surrounding hypertrophic chondrocytes adjacent to the SOC, but was absent in the control specimen. Type II collagen and decorin were absent in the SOC of the loaded specimen, but were expressed throughout the rest of the loaded epiphysis and the unloaded control epiphysis. The intensity of type II collagen and decorin expression was significantly stronger among hypertrophic chondrocytes surrounding the SOC than the control. The numbers of hypertrophic chondrocytes surrounding the SOC and superior to metaphyseal bone were significantly higher in the loaded specimens than the unloaded controls. Taken together, mechanical stresses accelerate the formation of the secondary ossification center, and therefore modulate endochondral ossification.