Dynamin inhibitor dynasore modulates longitudinal bone growth in a hormetic manner

Longitudinal bone growth occurs via endochondral ossification, involving a complex interplay of chondrocyte proliferation, differentiation, and matrix remodeling. As with all mammalian cells, chondrocytes require dynamin for mitochondrial fission, to shuttle vesicles from the Golgi apparatus, and for both clathrin- and caveolin-mediated endocytosis. Here, we aimed to test the functions of dynamin on bone growth. To do so, we applied dynasore-a small molecule that is a reversible dynamin inhibitor-to mouse metatarsal bones cultured ex vivo. We assessed gross changes using bone length measurements combined with EdU detection, immunostaining, super-resolution microscopy and transmission electron microscopy.

Dynasore induced a dose-dependent hormetic effect on bone elongation: while high concentrations (220 µM) impaired growth and abolished chondrocyte proliferation, low-dose treatment (40 µM) significantly increased longitudinal bone growth. Histological analysis demonstrated that low dose dynasore augmented epiphyseal cartilage expansion and matrix accumulation, while reducing chondrocyte proliferation. Immunostaining indicated that 40 µM dynasore preserved collagen type X synthesis, activated mTORC1 signaling, and blocked autophagy, based on SQSTM1 accumulation. Low dose dynasore treatment expanded the thickness of the filamentous actin layer at the plasma membrane and deepened endocytic pits containing collagen fibril-like electron-dense extracellular structures, indicating that impaired cartilage remodeling was associated with growth-associated matrix accumulation. Finally, the use of a structurally unrelated dynamin inhibitor, dynole, indicated that the effects of dynasore were partially mediated by its actions on dynamin.

Dynasore exerts hormetic effects on epiphyseal chondrocytes, wherein low doses stimulate bone elongation, and high doses impair chondrocyte function.