Smooth Muscle or Skeletal Muscle? Stem Cells Use Myocardin to Decide

University of Rochester Medical Center, October 12, 2007

According to a study published recently in the Proceedings of the National Academy of Sciences (PNAS), researchers have discovered a key protein that controls how stem cells "choose" to become smooth muscle cells that support blood vessels or skeletal muscle cells that move limbs. The results may point in the direction of new treatments for diseases that involve the creation of new blood vessels from stem cell reserves that would otherwise replace worn out skeletal muscle in addition to providing insight into the development of muscle types in the human fetus. New treatments could also be developed for cancer and atherosclerosis.

Physician researchers should start watching to see if a previously undetected side effect exists since the newly discovered mechanism also suggests that some current cancer treatments may weaken muscle.

With as many as 400 cell types in millions of combinations, humans develop from a single cell into a complex being thanks to stem cells. The potential to develop into every kind of human cell is locked within the fertilized embryo, or original single human cell. As we develop in the womb, successive generations of stem cells specialize (differentiate), with each group able to become fewer and fewer cell types.

The ability to become smooth muscle, skeletal muscle, blood, or bone is a characteristic of a set of mostly differentiated stem cells. Ready to differentiate into replacement parts depending on the stimuli they receive, many human tissues keep a reserve of stem cells on hand in adulthood. The stem cells take the required route if the body signals that skeletal muscle needs replacing. The same stem cells may become smooth muscle that supports the lining of blood vessels if tissues signal for more blood vessels.

A transcription factor called Myocardin may be the master regulator of whether stem cells become skeletal or smooth muscle. The discovery was made by a team of collaborating researchers at the University of Texas Southwestern Medical Center, and the Aab Cardiovascular Research Institute at the University of Rochester School of Medicine & Dentistry. Myocardin is a transcription factor, a protein designed to associate with a section of the DNA code, and to turn the expression of that gene on or off. Smooth muscle cells were originally thought to be the only tissue affected by Myocardin which was believed to be a protein that promoted cell growth by turning regulatory genes on. Myocardin is shown to also turn off genes that make skeletal muscle in the PNAS report.

"These findings could eventually lead to stem-cell based therapies where researchers take control of what the stem cell does once implanted through the action of transcription factors like myocardin, unlike current therapies that "hope" the stem cell will take a correct differentiation path to fight disease," said Joseph M. Miano, Ph.D., senior author of the paper and associate professor within the Aab Cardiovascular Research Institute at the University of Rochester Medical Center "More specifically, many diseases are driven by whether stem cells decide to become skeletal muscle, or instead to become part of new blood vessel formation. These discoveries have created a new wing of medical research that seeks to understand the genetic signals that turn on such stem cell replacement programs."

Atherosclerosis, or hardening of the arteries, for instance, becomes likely to cause heart attack or stroke when cholesterol-driven plaques that build up inside of arteries become fragile. Tissue death occurs when clots that block arteries develop from these plaques interact with circulating factors due to arterial rupture. In theory, the plaques could be strengthened to prevent rupture by injecting stem cells programmed to become smooth muscle said Miano.

Conversely, in order to grow, tumors must be able to grow blood vessels. New blood vessels are built by turning on vascular endothelial growth factor (VEGF), the tumor accomplishes this by sending signals for stem cells to form smooth muscle in combination with other signals.

Would manipulating myocardin along with VEGF interfere with tumor growth by cutting off its blood supply?

Do current VEGF-based treatments kick myocardin into action, creating smooth muscle instead of continually repairing worn out skeletal muscle?

Since VEGF is used experimentally to treat peripheral artery disease and coronary artery disease, is this treatment reducing the skeletal muscle strength of these patients?

Miano's team found that myocardin is a bi-functional developmental switch with the ability to both turn off the genes that turn stem cells into skeletal muscle as well as turn on a set of genes that turns stem cells into smooth muscle. Providing the biological context that made sense of Miano's finding was accomplished by applying the same idea to the development of the fetus via transgenic mouse studies by the team at Southwestern.

A group of cells in the human fetus known to develop into skeletal muscle are know as the somite. This has been the focus of research at many institutions. Myocardin is expressed briefly in the somite during development in mice, but then disappears from that region of the fetus. This was determined during cell lineage and tracking studies performed by the Southwestern team. This current data leads to the surprising theory that both skeletal and smooth muscle cells come from the same stem cell region. To make the new human's supply of smooth muscle cells, Myocardin briefly switches on. Blood vessels are formed when the cells migrate to another area. Then allowing the somite to continue differentiating into skeletal muscle, Myocardin quickly shuts off. Skeletal muscle would not develop properly if the turn off did not occur.

Seeking to define ancient sections of our genetic code that may soon be as important to medical science as genes has been the focus of many teams including Miano's in recent years. How small regulatory DNA sequences tell genes where, when and to what degree to "turn on" in combination with enzymes that seek them out has been the concern of this new wave of research. This is in contrast to putting the spotlight on on how genes work.

The workhorses that make up the body's organs and carry its signals; genes are the chains of deoxyribonucleic acids (DNA) that encode instructions for the building of proteins. The potential to create new classes of treatment for nerve disorders and heart failure are a side effect of the growing knowledge of how regulatory sequences control gene behavior. Once thought of as "junk DNA", regulatory sequences are emerging as an important part of the non-gene majority of human genetic material. The complete set of DNA sequences that regulate the precise turning on and off of genes is referred to as theregulome and it's study presents a new frontier in genetic research.

In an article by Miano and team published February 2006 in the journal Genome Research, they described one such regulatory sequence: the CArG box. The nucleotide building blocks of DNA chains may contain any one of four nucleobases: adenine (A), thymine (T), guanine (G) and cytosine (C). Any sequence of code starting with 2 Cs, followed by any combination of 6 As or Ts, and ending in 2 Gs is a CArG box.

Occurring approximately three million times throughout the human DNA blueprint, all together there are 1,216 variations of CArG box according to Miano. CArG boxes exert their influence over genes because they are "shaped" to partner with a nuclear protein called serum response factor (SRF) and several other proteins within a genetic regulatory network, including Myocardin. As many as sixty genes so far have been found to be influenced by the CArG-SRF, including many involved in heart cell and blood vessel function.

Past studies had determined that myocardin is a co-factor with SRF in CArG-Box mediated genetic regulation of stem cells. Through CArG box interaction, researchers believed myocardin partnered with SRF to turn on smooth muscle genes until now. But serving as a potent silencer of gene expression for the stem cell to skeletal muscle gene program, the current findings suggest that myocardin has a second role, independent of its partnership with CARG-SRF.

"With its dual action, myocardin is an early example of the efficiency and elegance of the system of genetic controls, where one factor has more than one complementary effect on the development of the body," said Eric Olson, Ph.D., chair of the Department of Molecular Biology at the University of Texas Southwestern Medical Center in Dallas, and also senior author of the study.