Early loading-related responses of bone cells

The work in this thesis investigates the influence of applied intermittent mechanical loading on prostaglandin (PG) release from resident bone cells in vitro, early responses of resident bone cells to exogenous PGs, and whether functional ion channels and modulators of phospholipase A₂ (PLA₂) effect the loading-related PG release.

Loading cancellous bone explants for fifteen minutes causes significant, but transient, increases in PGE and prostacyclin (PGI₂) concentrations in the perfusate. Immuno-histochemistry localises PGE₂ to osteoblasts and 6-keto-PGF_{1<IMG WIDTH=7 HEIGHT=7 ALIGN=BOTTOM SRC="/maths/alpha.gif">} (the stable hydrolytic product of PGI₂) to osteoblasts and osteocytes. Exogenous PGE₂ has no effect on levels of insulin-like growth factor (IGF)-I or II release, whereas PGI₂ increases IGF-II release by six hours, and increases [³H]uridine incorporation, indicating a stimulation of RNA production, possibly mRNA coding for IGF-II.

Resident bone cells in rat ulnar explants also respond to intermittent mechanical loads with increases in PGE₂ and PGI₂ release, and G6PD activity, whereas, those in calvaria do not. G6PD activity increases linearly in ulnar bone cells in response to increasing PGI₂ concentrations. In calvarial bone cells, this response is bi-phasic. Loading-related responses of calvarial bone cells appears to be different from that of limb bone cells.

Loading-related PGE₂, but not PGI₂, release is blocked with either an inhibitor of cytosolic PLA₂ activity or pertussis toxin. Inhibition of secretory PLA₂ blocks loading-related increases of both PGE₂ and PGI₂. This implies different mechanisms by which arachidonic acid is liberated for loading-related PGE₂ and PGI₂ release. Immunohistochemistry localises secretory PLA₂ to osteocytes and osteoblasts, and cytosolic PLA₂ to osteoblasts only.

The L-type voltage-dependent calcium channel blocker, nifedipine, blocks increases in loading-related PGE₂ release and osteoblast G6PD activity. Gadolinium chloride, an inhibitor of stretch/shear activated cation channels, only reduces these responses. Increases in osteocyte G6PD activity and PGI₂ release are blocked by gadolinium chloride, and unaffected by nifedipine. This implies activation of osteoblasts by L-type voltage-dependent calcium channels and stretch/shear activated cation channels, and osteocytic activation by stretch/shear activated cation channels alone.