In the prior funding period we obtained results strongly suggesting that oscillatory fluid flow due to loading is an important cellular physical signal for both osteoblasts and osteocytes. Utilizing our custom built dynamic flow system, we were able to show that oscillatory fluid flow can regulate cell metabolism via intracellular calcium mobilization, prostaglandin E2 release, and MAP kinase activity in the absence of other physical or biochemical signals. However, we have not uncovered the molecular mechanotransduction mechanism activated by oscillatory fluid flow. Candidates can be expected to experience load due to flow and have biochemical signaling potential. This conceptual model is supported by our observation made in the prior funding period that degradation of membrane proteoglycans extending into the flow field has a dramatic effect on the response to flow. Also, we have preliminary indications that actin and focal adhesion kinase (FAK) are involved fluid flow induced signaling. Thus, the central hypothesis of this five year project is that oscillatory fluid flow regulates bone cell metabolism via a molecular mechanism involving forces experienced by the cytoskeleton and transmitted through focal adhesion sites to integrins. To test this hypothesis we will undertake a systematic multilevel evaluation of cell structural proteins to include actin, integrins, and linker proteins both in terms of the effect of oscillatory flow on these proteins (aim 1) and the role of each in transducing the response to flow (aim 2). Additionally, strong evidence from our laboratory and others suggests specific involvement of focal adhesion kinase (FAK) tyrosine phosphorylation. This combined with recently developed molecular tools targeting FAK phosphorylation motivate us to perform a more in-depth investigation of two specific FAK signal pathways (aim 3). Finally, utilizing a novel microfabricated flow chamber, we will determine if the osteocyte process is a specialized structure with enhanced sensitivity to fluid shear forces (aim 4).

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR045989-09
Application #
7433185
Study Section
Special Emphasis Panel (ZRG1-MOSS-A (04))
Program Officer
Sharrock, William J
Project Start
2000-07-15
Project End
2009-05-31
Budget Start
2008-06-01
Budget End
2009-05-31
Support Year
9
Fiscal Year
2008
Total Cost
$370,954
Indirect Cost
Name
Stanford University
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
Downs, Matthew E; Nguyen, An M; Herzog, Florian A et al. (2014) An experimental and computational analysis of primary cilia deflection under fluid flow. Comput Methods Biomech Biomed Engin 17:2-10
Leucht, P; Monica, S D; Temiyasathit, S et al. (2013) Primary cilia act as mechanosensors during bone healing around an implant. Med Eng Phys 35:392-402
Nguyen, An M; Jacobs, Christopher R (2013) Emerging role of primary cilia as mechanosensors in osteocytes. Bone 54:196-204
Chen, Julia C; Jacobs, Christopher R (2013) Mechanically induced osteogenic lineage commitment of stem cells. Stem Cell Res Ther 4:107
Hoey, David A; Tormey, Shane; Ramcharan, Stacy et al. (2012) Primary cilia-mediated mechanotransduction in human mesenchymal stem cells. Stem Cells 30:2561-70
Castillo, Alesha B; Blundo, Jennifer T; Chen, Julia C et al. (2012) Focal adhesion kinase plays a role in osteoblast mechanotransduction in vitro but does not affect load-induced bone formation in vivo. PLoS One 7:e43291
Young, Y-N; Downs, M; Jacobs, C R (2012) Dynamics of the primary cilium in shear flow. Biophys J 103:629-39
Hoey, David A; Downs, Matthew E; Jacobs, Christopher R (2012) The mechanics of the primary cilium: an intricate structure with complex function. J Biomech 45:17-26
Case, N; Sen, B; Thomas, J A et al. (2011) Steady and oscillatory fluid flows produce a similar osteogenic phenotype. Calcif Tissue Int 88:189-97
Hoey, David A; Kelly, Daniel J; Jacobs, Christopher R (2011) A role for the primary cilium in paracrine signaling between mechanically stimulated osteocytes and mesenchymal stem cells. Biochem Biophys Res Commun 412:182-7

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