Oscillatory fluid flow regulates glycosaminoglycan production via an intracellular calcium pathway in meniscal cells.
Mechanical loading in the form of oscillatory fluid flow-induced shear stress was applied to meniscal cells while the biochemical response [intracellular calcium mobilization and sulfated glycosaminoglycan (GAG) production] was studied. Isolated rabbit meniscal cells were cultured onto microscope slides and placed in a parallel plate flow chamber. Cells were exposed to oscillating fluid flow-induced shear stresses of 4 Pa for sulfated GAG studies and 0-6.5 Pa for calcium studies. The calcium response was monitored using a fluorescent probe and imaging techniques, sulfated GAG production was measured using the modified 1,9-dimethylmethylene blue method, and thapsigargin was used to block intracellular calcium ([Ca2+]i) mobilization. A significant dose-dependent relationship was found for the percentage of cells responding to oscillating fluid flow with an increase in [Ca2+]i versus shear stress level. The percentage of cells responding decreased linearly from 72% +/- 17% at 6.5 Pa to 28% +/- 7% at 2.0 Pa to 2% +/- 1% for baseline no-flow (0 Pa). No differences were found in the amplitude of the calcium response of responding cells for any shear stress level. Oscillating fluid flow-induced shear stresses of 4 Pa produced a significantly greater amount of sulfated GAGs (253 +/- 95 ng GAG/microg cell protein) compared to the no-flow control (158 +/- 86 ng/microg). The addition of thapsigargin to the media inhibited both the intracellular calcium response to oscillating fluid flow (less than 1.5% of the cells responded) and the increase in GAG production following oscillating fluid flow, which was returned to control levels (170 +/- 72 ng/microg). These findings suggest that oscillatory fluid flow-induced shear stress increases intracellular calcium levels and sulfated GAG production. Furthermore, they suggests that calcium may modulate the biochemical pathway that leads to sulfated GAG production.
Eifler
Robert L
RL
Department of Mechanical Engineering, Michigan Technological University, Houghton, Michigan 49931, USA.
Blough
Eric R
ER
Dehlin
Jennifer M
JM
Haut Donahue
Tammy L
TL
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000378:metabolism*; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000187:drug effects / Q000502:physiology*; D008592:Menisci, Tibial / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011817:Rabbits; D013314:Stress, Mechanical; D019284:Thapsigargin / Q000494:pharmacology
D002121:Calcium Channel Blockers; D006025:Glycosaminoglycans; C013786:A73025; D019284:Thapsigargin; D002118:Calcium