Visit our Expo - Redox and Inflammation signaling 2012

Session II : Receptor signaling and G proteins Poster II, 12
ATP-dependent mechanotransduction by chondrocytes seeded in agarose constructs and
subjected to dynamic compression
T T Chowdhury and M M Knight
Cell and Tissue Engineering, Department of Engineering, Queen Mary, University of
London, Mile End Road, London. E1 4NS. UK E-mail : t.t.chowdhury@qmul.ac.uk
Introduction: Mechanical stimulation is essential in maintaining the biochemical properties of
articular cartilage. Both .NO and calcium are well known intracellular signaling molecules
which play an important role in chondrocyte mechanotransduction. It has been shown that
chondrocytes respond to mechanical stimuli by releasing ATP. Extracellular ATP binds to the
P2Y2 purinoreceptors, which may activate intracellular calcium levels and release of .NO,
therefore influencing chondrocyte metabolism. The current study examines whether inhibition
of mechanosensitive ATP by receptor antagonists will influence .NO levels, cell proliferation
and proteoglycan synthesis by chondrocytes cultured in agarose constructs and subjected to
dynamic compression.
Methods: Bovine chondrocytes were seeded in agarose gel and equilibrated in culture for 24
hrs. Using a full characterized cell-straining system, constructs were subjected to 15%
dynamic compression at a frequency of 1Hz for 48 hours in 1 ml of unsupplemented culture
medium or in medium supplemented with 20 µM gadolinium chloride (putative
mechanosensitive ion channel blocker), 100 µM suramin (P2 receptor antagonist) and 10
units.ml-1 apyrase (catalyses extracellular ATP). Medium was additionally incubated with 1
µCi.ml-1 [3H]-thymidine and 10 µM 35SO4, for the assessment of cell proliferation and
proteoglycan synthesis, respectively. Nitrite, a stable end product of .NO was measured using
the Griess assay.
Results: The application of dynamic compression resulted in the strain-induced stimulation of
[3H]-thymidine incorporation in unsupplemented constructs (p