Leaky calcium channels in muscular dystrophy
Duchenne muscular dystrophy (DMD) is an X-linked disorder affecting 1 in 3,500 males characterized by progressive muscle weakness and death by age 30 to 40 due to cardiac or respiratory failure. DMD is caused by mutations in the gene encoding dystrophin. In normal skeletal muscle cells, dystrophin forms part of a protein complex that connects the internal “skeleton” of the cell to the surrounding external matrix. In DMD, loss of dystrophin disrupts the stability of the cell membrane, leading to an increase in calcium ions entering into the cell. The rise in the cytosolic levels of calcium activates the enzyme calpain. Calpain digests proteins, including those responsible for muscle contraction, thus rendering muscles much weaker.
Calcium can enter the muscle cell cytosol not only from the outside, but also from an internal compartment called the sarcoplasmic reticulum (SR), which normally serves to sequester the calcium. A calcium release channel called ryanodine receptor 1 (RyR1) regulates the movement of calcium out of the SR and into the cytosol. Investigators from the Leducq Transatlantic Network on Calcium Cycling and Novel Therapeutic Approaches for Heart Failure studied the mdx mouse, the standard animal model of DMD, to determine whether a disruption in the RyR1 complex might contribute to the cytosolic calcium overload seen in DMD.
Dr. Andrew Marks, the North American coordinator of the network, Dr. Alain Lacampagne, an associated member of the network, and their team published their findings in the March 2009 issue of Nature Medicine. RyR1 contains several sites that may be modified by the addition of a molecule of nitric oxide, a process called nitrosylation, and the investigators showed that there is a significant increase in RyR1 nitrosylation in mdx mice compared to normal mice. This hypernitrosylation leads to the dissociation of RyR1 from calstabin-1, a protein that normally stabilizes RyR1. As a consequence, RyR1 channels become “leaky” and allow excess calcium into the cytosol. To determine the source of the nitric oxide, the investigators measured the levels of the three types of nitric oxide synthase (enzymes that produce nitric oxide; nNOS, eNOS, and iNOS). The investigators found that, compared to normal mice, nNOS levels and eNOS levels are lower in mdx mice, but iNOS levels are much higher. Moreover, they found that iNOS is physically associated with RyR1 in the mdx mice.
The investigators then tested whether stabilizing the RyR1 calcium channels would protect against the muscle damage due to calpain. After only a few weeks of treatment with S107, a RyR1 calcium channel stabilizer, the mdx mice demonstrated decreases in calcium leak, calstabin depletion, calpain activity, and muscle cell death, and corresponding improvements in muscle function and exercise tolerance.
In conclusion, this work demonstrates for the first time that leaky hypernitrosylated RyR1 calcium channels may play a role in the progression of DMD. Stabilizing RyR1 and calstabin and blocking calcium leak from the SR may therefore be worthwhile strategies to protect against muscle damage.
Click on the title to access the article in Nature Medicine: Hypernitrosylated ryanodine receptor calcium release channels are leaky in dystrophic muscle.