Mechanical loading induces primary cilia disassembly in tendon cells via TGFβ and HDAC6

Abstract

This study used isolated human tenocytes to test the hypothesis that cyclic mechanical strain directly stimulates primary cilia disassembly, and to elucidate the mechanisms involved. Cells were seeded onto flexible membranes and strained at 0–3%; 1 Hz, for up to 24 hours. Cilia length and prevalence progressively reduced with increasing strain duration but showed full recovery within 2 hours of strain removal. The response to loading was not influenced by actin organisation as seen in other cell types. However, the loading response could be recreated by treatment with TGFβ. Furthermore, treatment with the HDAC6 inhibitor Tubacin, or a TGFβ receptor inhibitor both prevented strain induced cilia disassembly. These data are the first to describe primary cilia expression in isolated tenocytes, showing that mechanical strain regulates cilia expression independent of changes in tendon extracellular matrix. Furthermore, we show that cilia disassembly is mediated by the activation of TGFβ receptors leading to activation of HDAC6. Previous studies have shown that cilia are required for TGFβ signalling and that tendon mechanosignalling is mediated by TGFβ. The present study therefore suggests a novel feedback mechanism whereby cilia disassembly inhibits prolonged TGFβ activation in response to continuous cyclic loading.

Primary cilia as the nexus of biophysical and hedgehog signaling at the tendon enthesis

“…Consistent with the current in vivo results, an inverse relationship has been demonstrated previously between ciliogenesis and mechanical loading in vitro (17,23,24). Mechanical stimulation above a threshold leads to cilia shortening and disassembly, and removal of mechanical loading leads to cilia elongation and assembly (24)(25)(26).…”

Regulation of the Extracellular Matrix by Ciliary Machinery

“…Primary cilia are aligned with parallel collagen fibrils in tendon [139], but this orientation is not affected by stress deprivation in rat tail tendon [140]. Also, isolated human tenocytes have random orientation of cilia, possibly due to lack of matrix in 2D culture, and thus this orientation could be due simply to constraints imposed by the collagen matrix [141]. In contrast, not much is known about cilia positioning in tendon.…”

The factory, the antenna and the scaffold: the three-way interplay between the Golgi, cilium and extracellular matrix underlying tissue function

“…The mechanical environment can also impact ciliogenesis itself. Cyclic and static strain induce cilium shortening in chondroctyes and tendons respectively, courtesy of HDAC6-mediated resorption ( Thompson et al, 2014 ; Rowson et al, 2018 ). Longer periods of strain also reduce cilium number ( McGlashan et al, 2010 ).…”

“…The relationship between the ECM and cilium orientation has also been challenging to dissect. In tendon, which is mechanically loaded in a single direction, collagen fibres and the ciliary axoneme align parallel to the direction of stretch ( Donnelly et al, 2010 ) whereas cilium orientation is random in isolated tenocytes ( Rowson et al, 2018 ). It remains unclear if this alignment in tissue is determined passively by the structural constraints imposed by an organised ECM, or more actively directed by mechanotransduction.…”