News Release

Top: When aortic endothelial cells were stretched in the up-and-down orientation shown here, they grew "stress fibers" (red) in a "healty" alignment perpendicular to the axis of stretch. Bottom: When researchers inhibited a protein called Rho in aortic endothelial cells, stress fibers grew in an "unhealthy" direction parallel to the axis of stretch.

Bioengineering researchers at UCSD’s Jacobs School of Engineering reported in the cover story of the Nov. 1 issue of Proceedings of the National Academy of Sciences (PNAS) that arterial endothelial cells subjected to repeated stretching (10 percent of their length, 60 times per minute) produced intracellular arrays of parallel “stress fibers” in a few hours.

The tests were performed on endothelial cells lining the aorta of a cow, but the endothelial cells of the human aorta are expected to react similarly. The stress fibers were made of actin, a fibrous protein that is part of the machinery that gives muscle its ability to contract. Actin also gives virtually all cells their ability to make an internal “cytoskeleton.” The stress fibers of endothelial cells in arteries are aligned parallel to the long axis of blood vessels, and this alignment is perpendicular to the direction of rhythmic stretching caused by a beating heart. Such an orientation of stress fibers is a hallmark of healthy blood vessels, but scientists currently understand few of the factors responsible for generating that configuration.

Shu Chien, left, a UCSD professor of bioengineering, and research scientist Shunichi Usami adhered endothelial cells to a transparent layer of silicone rubber, a sample of which is being held by Usami, and stretched the cells from 1 percent to 10 percent of their length.

Each of these bovine aorta endothelial cells were stretched along an axis that runs up and down in these photographs. An unstretched cell, top, produced "stress fiber" filaments in random directions, but as the stretching increased from 3 percent of a cell length, middle, to 10 percent of a cell length, bottom, stress fibers were increasingly likely to be aligned perpendiculat to the stretching force.

In the most significant finding in the PNAS article, which was made available online Oct. 24, Chien’s group reported that when an intracellular protein called Rho was chemically inhibited, stress fibers grew in the “wrong” direction; they grew parallel rather than perpendicular to the direction of cell stretching. Without Rho, the cells lost their ability to orient stress fibers properly.

“Rho is a very important molecule,” said Chien. “It works in response to, and in concert with, physical stretching to generate the healthy alignment of stress fibers.” Indeed, when Chien’s group used a genetic technique to increase the activity of Rho, those cells grew stress fibers in the healthy direction at a lower threshold of stretching.

   “Until now, it has not been shown that there is an equivalence and cooperation between mechanical and biochemical stimuli to regulate the proper orientation of these stress fibers,” said Kaunas. “Indeed, we found that the stress fibers oriented in such a way to control their level of stress – not too little and not too much.”

Chien and Kaunas collaborated with UCSD research scientist Shunichi Usami, who contributed to the design of the miniature workout chambers. Silicone rubber membranes inside the chambers were coated with a protein that allowed the endothelial cells to adhere to the membranes in a manner similar to how they attach to underlying blood vessel tissue in the body. The researchers isolated endothelial cells from the bovine aorta, grew the cells in culture flasks, and seeded them onto the silicone membranes. After the cells grew into confluent layers, a piston-like “indenter” was programmed to repeatedly 

The cover image of the Nov. 1, 2005, issue of PNAS shows  stress fibers in endothelial cells.

The researchers also demonstrated that inhibition of either Rho or a related protein called Rho kinase resulted in loss of the healthy alignment of stress fibers as well as alignment of adhesion sites where those stress fibers would attach to the cell membrane.

“These new results clearly show that Rho and physical stresses cooperate to produce healthy alignments of stress fibers,” said Chien. “We need to understand how cells can sense the mechanical force and achieve this beneficial effect through the activation of Rho, and we also need to identify other proteins that may be involved in this feedback control mechanism.”