When muscle fibers fail to properly seal tiny tears in their membrane, as occurs in some forms of muscular dystrophy, the resulting leakage can change the surrounding environment and attract immune cells that contribute to further muscle damage, a mouse study suggests.
The findings suggest that the tissue surrounding muscle fibers may play an active role in driving inflammation and disease progression, rather than simply providing structural support.
“The main finding of this work is that prolonged membrane leak due to defective repair creates a unique muscle environment,” Elizabeth McNally, MD, PhD, director of Northwestern University’s Center for Genetic Medicine and the study’s co-lead author, said in a university press release.
The findings could help researchers identify new treatment strategies for muscular dystrophy that target inflammation and changes in the tissue surrounding muscle fibers to slow disease progression.
The study, “Excess muscle plasma membrane leak disrupts ECM [extracellular matrix] content and shifts macrophage-mediated muscle repair,” was published in the JCI Insight.
Mice lacking two key proteins had more muscle damage
Muscle fibers routinely develop small tears in their outer membrane as they contract. Normally, the body quickly repairs these injuries, preventing the contents of the fibers from escaping into the surrounding tissue.
Two proteins, dysferlin and annexin A6, help muscle fibers repair these tears. Problems with this repair process are known to cause certain forms of muscular dystrophy.
“In humans, there is a form of muscular dystrophy that comes from having mutations in the dysferlin gene. In this form of disease, patients have very leaky muscles, which can be measured by detecting muscle proteins in the blood. For these patients, their muscles become dysfunctional, causing them to have weak muscles that cannot support walking,” McNally said.
However, it’s not exactly known how chronic membrane leakage from impaired membrane repair may contribute to disease. To find out, researchers studied mice lacking either dysferlin or annexin A6. They also created mice lacking both dysferlin and annexin A6, referred to as DysfA6 mice, which had particularly severe problems repairing damaged muscle membranes.
Compared with mice lacking dysferlin alone, DysfA6 mice had more muscle damage, inflammation, tissue scarring, fat deposits, and immune cell infiltration, along with poorer muscle function and higher blood levels of creatine kinase, a marker of muscle damage. A dye used to detect leaky muscle fibers entered 19 times more fibers in DysfA6 mice, confirming substantially greater membrane leakage.
Damaged muscle fibers signaled for macrophages to ‘eat’ them
The researchers next looked at the immune cells present in the damaged muscles. An analysis of more than 82,000 individual cells found that macrophages and related immune cells made up the largest immune cell population in dysferlin-deficient and DysfA6 muscles.
Macrophages are immune cells that normally help clean up damaged or dead cells and support tissue repair. But in the mice with persistent membrane leakage, the researchers found evidence that these cells were contributing to the disease process. In particular, macrophages carrying the markers Mertk and Trem2 were drawn to the leaky muscle.
The researchers found that damaged muscle fibers produced signals that encouraged macrophages to “eat” them, a process known as phagocytosis. In lab experiments, macrophages were more likely to engulf muscle fibers lacking dysferlin than healthy fibers or fibers from a mouse model of Duchenne muscular dystrophy.
“A key finding is that leaky muscle fibers draw macrophages to come and ‘take a bite’ in a process called phagocytosis,” McNally said. “The excess macrophages and their tendency to phagocytose leaky muscle appears to contribute to making the disease process much worse.”
The researchers also found that persistent leakage altered the extracellular matrix, the network of proteins and other molecules surrounding and supporting muscle fibers. In particular, proteins called annexin A1 and annexin A2 accumulated in the surroundings of the most severely affected fibers.
Collectively, our findings demonstrate that the prolonged leak due to defective membrane repair not only alters the muscle microenvironment but also redefines the immune-[extracellular matrix] interface, offering mechanistic insight into how defective repair propagates chronic inflammation and [tissue scarring] in muscular dystrophy.
This altered environment, in turn, was found to attract more macrophages and cause them to adopt features associated with inflammatory activity.
The findings suggest that persistent membrane leakage may therefore create a damaging cycle: muscle fibers become leaky, the surrounding environment changes, macrophages are attracted to the area, and the resulting immune response may contribute to additional muscle damage and inflammation.
“Collectively, our findings demonstrate that the prolonged leak due to defective membrane repair not only alters the muscle microenvironment but also redefines the immune-[extracellular matrix] interface, offering mechanistic insight into how defective repair propagates chronic inflammation and [tissue scarring] in muscular dystrophy,” the researchers wrote.
The scientists said the findings could point to new treatment strategies targeting the immune signals or changes in surrounding area that develop when muscle fibers are not repaired adequately and become chronically leaky.
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