A Chick Embryo in-Vitro Model of Knee Morphogenesis

   Background: In this feasibility study, a mechanically loaded in-vitro tissue culture model of joint morphogenesis using the isolated lower extremity of the ۸ day old chick embryo was developed to assess the effects of mechanical loading on joint morphogenesis. Methods: The developed in-vitro system allows controlled flexion and extension of the chick embryonic knee with a range of motion of ۲۰ degrees from a resting position of ۹۰-۱۰۰ degrees of flexion. Joint morphogenesis at ۲, ۳, ۴ and ۷ days of culture was assessed by histology and micro MRI in ۴ specimen types: undisturbed in-ovo control embryos, in-ovo paralyzed embryos, in-vitro unloaded limb cultures, and in-vitro loaded limb cultures. Relative glycosaminoglycan (GAG) concentration across the joint was assessed with an MRI technique referred to as dGEMRIC (delayed gadolinium enhanced MRI of cartilage) where T۱ is proportional to glycosaminoglycan concentration. Results: Average T۱ over the entire tissue image for the normal control (IC) knee was ۴۸۰ msec; for the ۴ day loaded specimen average T۱ was ۳۵۴ msec; and for the ۷ day loaded specimens T۱ was ۳۹۳ msec. The ۴ day unloaded specimen had an average T۱ of ۲۷۹ msec while the ۷ day unloaded specimen had an average T۱ of ۲۲۴ msec. The higher T۱ values in loaded than unloaded specimens suggest that more glycosaminoglycan is produced in the loaded culture than in the unloaded preparation. Conclusion: Isolated limb tissue cultures under flexion-extension load can be viable and exhibit more progression of joint differentiation and glycosaminoglycan production than similarly cultured but unloaded specimens. However, when compared with controls consisting of intact undisturbed embryos in-ovo , the isolated loaded limbs in culture do not demonstrate equivalent amounts of absolute growth or joint differentiation.

Keywords:

chick embryo , chick embryo culture , Glycosaminoglycan , in-vitro loaded limb culture , joint morphogenesis , paralyzed chick embryo