MAP kinase and calcium signaling mediate fluid flow-induced human mesenchymal stem cell proliferation

Abstract

Although the mechanisms by which osteoblasts and osteocytes respond to fluid flow are being elucidated, little is known about the mechanisms by which bone marrow-derived mesenchymal stem cells respond to such stimuli. Here we show that the intracellular signaling cascades activated in human mesenchymal stem cells by fluid flow are similar to those activated in osteoblastic cells. Oscillatory fluid flow inducing shear stresses of 5, 10, and 20 dyn/cm 2 triggered rapid, flow rate-dependent increases in intracellular calcium that pharmacological studies suggest are inositol trisphosphate mediated. The application of fluid flow also induced the phosphorylation of extracellular signal-regulated kinase-1 and -2 as well as the activation of the calcium-sensitive protein phosphatase calcineurin in mesenchymal stem cells. Activation of these signaling pathways combined to induce a robust increase in cellular proliferation. These data suggest that mechanically induced fluid flow regulates not only osteoblastic behavior but also that of mesenchymal precursors, implying that the observed osteogenic response to mechanical loading may be mediated by alterations in the cellular behavior of multiple members of the osteoblast lineage, perhaps by a common signaling pathway. mechanotransduction; bone; marrow MAINTENANCE OF APPROPRIATE bone mass requires the coordination of bone resorption by osteoclasts and bone deposition by osteoblasts, and it is well established that mechanical stimuli can regulate the balance between bone formation and resorption. The addition of exogenous mechanical load is believed to stimulate new bone formation through increases in osteoblastic activity and concomitant decreases in osteoclastic activity (20). Conversely, removal of mechanical load, as is the case during spaceflight and disuse, leads to decreased osteoblastic activity and loss of bone mass (42,56).Accumulating evidence suggests that individual bone cells, including osteocytes and osteoblasts, are responsible for perceiving and responding to mechanical signals. Such signals, which may include streaming potentials, mechanical strain, and fluid shear stress, elicit a host of biochemical responses including mobilization of second messengers such as calcium (22,72), nitric oxide (26, 30), prostaglandins, and inositol trisphosphate (IP 3 ) (49); activation of kinase cascades including the MAP kinase and PKC pathways (44, 50); and modulation of gene expression (5, 46, 73). Strain-induced oscillatory fluid flow has been shown by our laboratory (73) and others (45) to be a potent biophysical stimulus. In MC3T3-E1 preosteoblasts, fluid flow induces the mobilization of intracellular calcium (Ca i 2ϩ ), activates ERK1/2, and increases osteopontin mRNA levels (11,72,73). Recent evidence has shown that exposing the osteocytic cell line MLO-Y4 to physiological levels of fluid flow induces similar responses (1, 6, 52).Although the mechanisms by which osteoblasts and osteocytes respond to mechanical stimuli are being elucidated, little is known about how bo...

Brief Reports: TRPM7 Senses Mechanical Stimulation Inducing Osteogenesis in Human Bone Marrow Mesenchymal Stem Cells

“…This finding was consistent with the heterologously expressed TRPM7 in HEK293T cells [20]. [3], indicating that the IP 3 receptor (IP 3 R) may play an essential role. However, the exact IP 3 R subtype mediating Ca 21 release is still unknown.…”

“…This proposal is indirectly supported by another study showing that cilia mediated fluid flow mechanostimulaiton in human MSCs, and increased osteogenic gene expressions [27]. Fluid flow was also reported to induce an increase in intracellular calcium in human MSCs [3]. Furthermore, TRP channels located in the cilia can transduce the mechanical stimulation to calcium signals [28].…”

“…Accumulating evidence suggests that mechanical stimuli, such as pressure [2], shear stress [3], and stretch [4], modulate MSC differentiation through mobilization of second messengers, such as inositol trisphosphate (IP 3 ) [5] and Ca 21 [6]. Spontaneous Ca 21 oscillations and stimulation induce a cytosolic Ca 21 increase, which has been well defined to regulate the expression level of transcription related genes [7,8].…”

From streaming‐potentials to shear stress: 25 years of bone cell mechanotransduction

“…One hour of oscillatory fluid flow exposure induces a significant increase in MSC proliferation via the activation of cell signaling cascades similar to those activated in more mature bone cells. 102 We demonstrated that fluid flow stimulates the activation of MAP kinase signaling as well as an increase in intracellular calcium concentration that leads to the activation of calcineurin, and pharmacological inhibition of these pathways abolished the effect of fluid flow on MSC proliferation. MSCs actively release ATP in response to a fluid flow stimulus, and this molecule leads to the activation of the signaling cascades necessary for fluid flow to induce proliferation.…”

Calcium Signalling and Calcium Transport in Bone Disease

“…Skeletal flexion also promotes chondrocyte growth [Drescher et al, 2003]. Osteoblastic cellular responses include cell proliferation [Riddle et al, 2006], as well as diverse, generally anabolic, effects on differentiationand activity.…”