title: Calcium-channel activation and matrix protein upregulation in bone cells in response to mechanical strain.
abstract: Femur-derived osteoblasts cultured from rat femora were loaded with Fluo-3 using the AM ester. A quantifiable stretch was applied and [Ca(2+)]i levels monitored by analysis of fluorescent images obtained using an inverted microscope and laser scanning confocal imaging system. Application of a single pulse of tensile strain via an expandable membrane resulted in immediate increase in [Ca(2+)]i in a proportion of the cells, followed by a slow and steady decrease to prestimulation levels. Application of parathyroid hormone (10(-6) M) prior to mechanical stimulation potentiated the load-induced elevation of [Ca(2+)]i. Mechanically stimulating osteoblasts in Ca(2+)-free media or in the presence of either nifedipine (10 microM; L-type Ca(2+)-channel blocker) or thapsigargin (1 microM; depletes intracellular Ca(2+) stores) reduced strain-induced increases in [Ca(2+) ]i. Furthermore, strain-induced increases in [Ca(2+)]i were enhanced in the presence of Bayer K 8644 (500 nm), an agonist of L-type calcium channels. The effects of mechanical strain with and without inhibitors and agonists are described on the total cell population and on single cell responses. Application of strain and strain in the presence of the calcium-channel agonist Bay K 8644 to periosteal-derived osteoblasts increased levels of the extracellular matrix proteins osteopontin and osteocalcin within 24 h postload. This mechanically induced increase in osteopontin and osteocalcin was inhibited by the addition of the calcium-channel antagonist, nifedipine. Our results suggest an important role for L-type calcium channels and a thapsigargin-sensitive component in early mechanical strain transduction pathways in osteoblasts.
authors.affiliation: Centre for Science and Technology in Medicine, Keele University, Stoke on Trent, Birmingham, United Kingdom.
authors.forename: L M
authors.initials: LM
authors.lastname: Walker
authors.forename: S J
authors.initials: SJ
authors.lastname: Publicover
authors.forename: M R
authors.initials: MR
authors.lastname: Preston
authors.forename: M A
authors.initials: MA
authors.lastname: Said Ahmed
authors.forename: A J
authors.initials: AJ
authors.lastname: El Haj
mesh_terms: D000818:Animals; D001857:Bone Matrix / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism*; D002478:Cells, Cultured; D010006:Osteoblasts / Q000187:drug effects / Q000378:metabolism*; D015675:Osteocalcin / Q000096:biosynthesis; D053495:Osteopontin; D010281:Parathyroid Hormone / Q000494:pharmacology; D051381:Rats; D012795:Sialoglycoproteins / Q000096:biosynthesis; D013314:Stress, Mechanical; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D015220:Calcium Channels; D010281:Parathyroid Hormone; D012795:Sialoglycoproteins; C506897:Spp1 protein, rat; D015675:Osteocalcin; D053495:Osteopontin; D019284:Thapsigargin; D002118:Calcium