Abstract
Recently fluid flow has been shown to be a potent physical stimulus in the regulation of bone cell metabolism. However, most investigators have applied steady or pulsing flow profiles rather than oscillatory fluid flow, which occurs in vivo because of mechanical loading. Here oscillatory fluid flow was demonstrated to be a potentially important physical signal for loading-induced changes in bone cell metabolism. We selected three well known biological response variables including intracellular calcium (Ca(2+)i), mitogen-activated protein kinase (MAPK) activity, and osteopontin (OPN) mRNA levels to examine the response of MC3T3-E1 osteoblastic cells to oscillatory fluid flow with shear stresses ranging from 2 to -2 Newtons/m(2) at 1 Hz, which is in the range expected to occur during routine physical activities. Our results showed that within 1 min, oscillatory flow induced cell Ca(2+)i mobilization, whereas two MAPKs (ERK and p38) were activated over a 2-h time frame. However, there was no activation of JNK. Furthermore 2 h of oscillatory fluid flow increased steady-state OPN mRNA expression levels by approximately 4-fold, 24 h after exposure to fluid flow. The presence of both ERK and p38 inhibitors and thapsigargin completely abolished the effect of oscillatory flow on steady-state OPN mRNA levels. In addition, experiments using a variety of pharmacological agents suggest that oscillatory flow induces Ca(2+)i mobilization via the L-type voltage-operated calcium channel and the inositol 1,4,5-trisphosphate pathway.
Highlights
Fluid flow has been shown to be a potent physical stimulus in the regulation of bone cell metabolism
We selected three well known biological response variables including intracellular calcium (Ca2؉i), mitogen-activated protein kinase (MAPK) activity, and osteopontin (OPN) mRNA levels to examine the response of MC3T3–E1 osteoblastic cells to oscillatory fluid flow with shear stresses ranging from 2 to ؊2 Newtons/m2 at 1 Hz, which is in the range expected to occur during routine physical activities
In this study we examine a novel physical stimulus and loading-induced oscillatory fluid flow and demonstrate that when applied to cultured osteoblastic cells at levels expected to occur in vivo it regulates mRNA levels for an important bone matrix protein, osteopontin (OPN)
Summary
Fluid flow has been shown to be a potent physical stimulus in the regulation of bone cell metabolism. In this study we examine a novel physical stimulus and loading-induced oscillatory fluid flow and demonstrate that when applied to cultured osteoblastic cells at levels expected to occur in vivo it regulates mRNA levels for an important bone matrix protein, osteopontin (OPN).. In this study we examine a novel physical stimulus and loading-induced oscillatory fluid flow and demonstrate that when applied to cultured osteoblastic cells at levels expected to occur in vivo it regulates mRNA levels for an important bone matrix protein, osteopontin (OPN).1 This regulation occurs via an increase in intracellular calcium (Ca2ϩi) and mitogen-activated protein kinases (MAPKs). The sensitivity of bone tissue to mechanical loading has been proposed to involve a variety of cellular biophysical signals including loading-induced electric fields, matrix strain, and fluid flow The latter effect of loading, originally described by Piekarski et al [3], has recently been proposed to directly regulate bone cell metabolism in vivo [4, 5]. Oscillatory Flow Activated Ca2ϩi, MAP Kinases, and Osteopontin diate, and long term effects of oscillatory fluid flow on bone cells, as well as on their inter-relationships
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