It is widely accepted that connective tissue cells respond to biomechanical forces transmitted through a complex extracellular matrix (ExMx), and produce a functional ExMx commensurate with the load applied. Functional tissue engineering aims to replicate this process using mechanically active bioreactor systems to produce tissues in vitro with mechanical properties similar to native tissues. However, the fundamental mechanotransductory pathways involved in signalling biomechanical change and polarizing the cellular secretory response, still remain unclear.
We have used precise tomographic modeling to illustrate the complex structural continuum that exists between the ExMx, the primary cilium and the Golgi apparatus in chondrocytes. Uniquely located at an intermediate position between the mechanically functional ExMx, and the synthetic organelles secreting ExMx, the primary cilium plays an integrating role between different signalling mechanisms including the calcium, integrin, hedgehog, Wnt, purinergic, PDGFaa and TGF-b signalling pathways. Recent chondrocyte compression studies show that primary cilia mediate mechanotransduction through regulation of polycystin-1-dependent ATP-induced Ca2+ signalling to modulate matrix synthesis in response to physiological strain(1).
We are currently assessing the role of primary cilia in cartilage tissue engineering using chondrocytes derived from an ovine model of the human ciliopathy, Meckel-Gruber Syndrome, where the matrix-cilium-Golgi continuum is disrupted. Mutant and wild-type chondrocytes were prepared for neocartilage production by a three-phase process of proliferation, pellet formation and bioreactor cultivation, with or without mechanical loading. Mutant neocartilage beads had the longest cilia and greatest volume, but were the least dense beads, displayed obvious vesiculation of cells and matrix, had poor pericellular matrix differentiation, and showed abnormal sequestration of collagens II, VI, and aggrecan when compared to controls.
While we do not understand the putative receptor function of meckelin, the protein mutated in our model, the perturbation of this transition zone protein is sufficient to compromise the matrix-cilium-Golgi continuum during cartilage tissue engineering.