Self-healing engineered muscle grown in ‘bionic mouse’

Contracts as strongly as native neonatal skeletal muscle, a first
April 2, 2014

Engineered muscle fiber stained to observe growth after implantation into a mouse (credit: Duke University)

Duke University biomedical engineers have grown living skeletal muscle that resembles the real thing. It contracts powerfully and rapidly, integrates into mice quickly, and for the first time, demonstrates the ability to heal itself both inside the laboratory and inside an animal.

The researchers watched the muscle growth in real time through a window on the back of a living, walking mouse.

Both the lab-grown muscle and experimental techniques are important steps toward growing viable muscle for studying diseases and treating injuries, said Nenad Bursac, associate professor of biomedical engineering at Duke.

The results appear in the Proceedings of the National Academy of Sciences Early Edition March 31.

“The muscle we have made represents an important advance for the field,” Bursac said. “It’s the first time engineered muscle has been created that contracts as strongly as native neonatal skeletal muscle.”

Through years of perfecting their techniques, a team led by Bursac and graduate student Mark Juhas discovered that preparing better muscle requires two things—well-developed contractile muscle fibers and a pool of muscle stem cells, known as satellite cells.

Every muscle has satellite cells on reserve, ready to activate upon injury and begin the regeneration process. The key to the team’s success was successfully creating the microenvironments—called niches—where these stem cells await their call to duty.

“Simply implanting satellite cells or less-developed muscle doesn’t work as well,” said Juhas. “The well-developed muscle we made provides niches for satellite cells to live in, and, when needed, to restore the robust musculature and its function.”

To put their muscle to the test, the engineers ran it through a gauntlet of trials in the laboratory. By stimulating it with electric pulses, they measured its contractile strength, showing that it was more than 10 times stronger than any previous engineered muscles. They damaged it with a toxin found in snake venom to prove that the satellite cells could activate, multiply and successfully heal the injured muscle fibers.

This series of images shows the destruction and subsequent recovery of engineered muscle fibers that had been exposed to a toxin found in snake venom (credit: Duke University)

Then they moved it out of a dish and into a mouse.

The team inserted their lab-grown muscle into a small chamber placed on the backs of live mice. The chamber was then covered by a glass panel. Every two days for two weeks, they imaged the implanted muscles through the window to check on their progress.

By genetically modifying the muscle fibers to produce fluorescent flashes during calcium spikes—which cause muscle to contract— the researchers could watch the flashes become brighter as the muscle grew stronger.

The progress of veins slowly growing into implanted engineered muscle fibers (credit: Duke University)

“We could see and measure in real time how blood vessels grew into the implanted muscle fibers, maturing toward equaling the strength of its native counterpart,” said Juhas.

The engineers are now beginning work to see if their biomimetic muscle can be used to repair actual muscle injuries and disease.

This work was supported by a National Science Foundation Graduate Research Fellowship and the National Institute of Arthritis and Musculoskeletal and Skin Diseases.

Abstract of PNAS paper

Tissue-engineered skeletal muscle can serve as a physiological model of natural muscle and a potential therapeutic vehicle for rapid repair of severe muscle loss and injury. Here, we describe a platform for engineering and testing highly functional biomimetic muscle tissues with a resident satellite cell niche and capacity for robust myogenesis and self-regeneration in vitro. Using a mouse dorsal window implantation model and transduction with fluorescent intracellular calcium indicator, GCaMP3, we nondestructively monitored, in real time, vascular integration and the functional state of engineered muscle in vivo. During a 2-wk period, implanted engineered muscle exhibited a steady ingrowth of blood-perfused microvasculature along with an increase in amplitude of calcium transients and force of contraction. We also demonstrated superior structural organization, vascularization, and contractile function of fully differentiated vs. undifferentiated engineered muscle implants. The described in vitro and in vivo models of biomimetic engineered muscle represent enabling technology for novel studies of skeletal muscle function and regeneration.