Cyclic Hydraulic Pressure and Fluid Flow Differentially Modulate Cytoskeleton Re-Organization in MC3T3 Osteoblasts

Abstract

Mechanical loads are essential towards maintaining bone mass and skeletal integrity. Such loads generate various stimuli at the cellular level, including cyclic hydraulic pressure (CHP) and fluid shear stress (FSS). To gain insight into the anabolic responses of osteoblasts to CHP and FSS, we subjected MC3T3-E1 preosteoblasts to either FSS (12 dynes/cm 2 ) or CHP varying from 0 to 68 kPa at 0.5 Hz. As with FSS, CHP produced a significant increase in ATP release over static controls within 5 min of onset. Cell stiffness examined by atomic force microscopy increased after 15 min of either CHP or FSS stimulation, which was attenuated when extracellular ATP was hydrolyzed with apyrase. As previously shown FSS induced polymerization of actins into stress fibers. However, the microtubule network was completely disrupted under FSS. In contrast, CHP appeared to maintain strong microtubule and f-actin networks. The purinergic signaling was found to be involved in the remodeling of f-actin, but not microtubule. Both CHP and FSS applied for 1 hour increased expression of COX-2. These data indicate that, while CHP and FSS produce similar anabolic responses, these stimuli have very different effects on the cytoskeleton remodeling and could contribute to loss of mechanosensitivity with extended loading.

Inserted rest period resensitizes MC3T3-E1 cells to fluid shear stress in a time-dependent manner via F-actin-regulated mechanosensitive channel(s)

“…12) In the meantime, F-actin begins to polymerize, gradually increasing the cell stiffness and the cellular mechanical threshold. 20,25) The combined effects of the 3-min sFSS stimulation and the following rest period reinforce F-actin stress fibers in 10 min of rest, but weaken F-actin stress fibers thereafter, and ultimately lead to the Several limitations exist in our study. First, we showed that F-actin regulated mechanosensitive channel(s) influenced mechanosensitivity recovery in MC3T3-E1 cells during a rest period.…”

“…20,24) In bone tissue, cells are exposed to a variety of mechanical stimulations, such as FSS, hydrostatic pressure, and stretching. sFSS stimulation has been shown to increase the polymerization of actin monomers, 25) F-actin alignment, 26) and microtubule remolding, 25) which indicate adaptation of cells to external stimulation. However, this may also cause an increase in the cellular mechanical threshold, and thus, decrease the mechanosensitivity in continuously loaded cells.…”

Purinergic signalling in the musculoskeletal system

“…A study also demonstrated that cyclic hydraulic pressure and fluid flow differentially modulate cytoskeleton re-organization in MC3T3 osteoblasts. This was suggested to contribute to the loss of mechanosensitivity with extended loading [197].…”

Enhanced chondrogenesis with upregulation of PKR using a novel hydrostatic pressure bioreactor

“…4) Previous studies on mouse chondrocytes have reported the use of hydrostatic pressures in the range 10-70 kPa applied at 0.25-1 Hz for short (5,15,30, and 60 min) or long periods (1 h daily for up to 19 days). 5,6) In this study, a similar range of pressure, 5-37 kPa at 0.5 Hz, was loaded 1 h daily for 3 days.…”