Regulation of Signaling Molecules in Endothelial Cells by Steady Flow and Pulsatile Flow
Date Issued
2010
Date
2010
Author(s)
Kuo, Jen-Yuan
Abstract
Shear stress generated by blood flow regulates vascular endothelial cell structure and function, thus playing a crucial role in the cardiovascular physiology and atherosclerosis process. Since the blood flow in blood vessels is pulsatile, the magnitude of shear stress applied on the vessel wall alters periodically. We used the roller pump and the syringe pump to generate steady flow and pulsatile flow, respectively. We investigated and compared the influence of the two types of flow on the signaling molecules, including protein kinase Cα (PKCα), PKCε, ERK1/2, β-catenin, Akt, eNOS and reactive oxygen species (ROS) in bovine arotic endothelial cells (BAECs).
The flow chamber system, which consisted of roller pump and syringe pump, was able to produce steady flow and pulsatile flow with shear stress magnitude of 12 dyn/cm2 and 12±4 dyn/cm2, respectively. The two types of flow did not have significant effects upon the phosphorylation of PKCα and PKCε. The phosphorylation of ERK1/2, as the downstream of PKC, was transiently induced by the stimulation of the flow. Both steady flow and pulsatile flow activated Akt with the maximum activation at 10 minutes and maintained for 30 minutes after BAECs subjected to the flow. The stability of β-catenin was regulated by its phosphorylation at serine/threonine residues by Axin complex and PKCα. After BAECs were exposed to the two types of flow, there were no significant changes in the phosphorylation of β-catenin. We also used the confocal microscopy to identify the location of β-catenin in BAECs and observed that β-catenin continuously appeared at adherens junctions. Known that eNOS activity was regulated by Akt, and Akt was activated under the both types of flow, the phosphorylation of eNOS at Ser1179 increased 4 times as compared with basal level, as expected. The phosphorylation of eNOS at Ser635 increased 3 to 5 times as compared with basal level, and the phosphorylation sustained longer. After BAECs were stimulated by the two types of flow for 30 minutes, the ROS levels in BAECs increased 2.5 to 3 times as compared with basal level.
In summary, the steady flow and the pulsatile flow might activate PKCα and PKCε by the way other than phosphorylation. Both types of flow activated ERK1/2 and Akt, which promoted cell survival and prevented cell apoptosis. Both types of flow also activated eNOS to counteract the elevated ROS levels and providing protective effects for cardiovascular system. Slightly increased ROS levels might function as signaling molecules to mediate cellular signal transduction.
Subjects
steady flow
pulsatile flow
shear
endothelial cells
signaling molecules
Type
thesis
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