CHAPTER THIRTEEN
MYOPATHOLOGY

PROGRESSIVE MUSCULAR DYSTROPHIES

Muscular dystrophies are genetically transmitted diseases characterized pathologically by degeneration and loss of myofibers and clinically by inexorably progressive weakness and, many of them, by elevated CK. The pattern of weakness, tempo of evolution, and mode of inheritance vary among different dystrophies. Over 30 genes causing muscular dystrophy are known presently. Muscular dystrophies are clinically classified into the following groups:

Dystrophinopathies (Duchenne and Becker muscular dystrophies)

Limb-Girdle dystrophies

Myotonic dystrophy

Facioscapulohumeral and scapuloperoneal dystrophy

Oculopharyngeal muscular dystrophy

Distal myopathies

Emery-Dreifuss muscular dystrophy

Congenital muscular dystrophies

Some of these groups contain several entities with different inheritance patterns. The most common muscular dystrophy in children is Duchene muscular dystrophy. In adults, the most common dystrophies are myotonic dystrophy and the limb girdle dysytrophies. The molecular pathogenesis and the basis for the genotypic and phenotypic diversity of muscular dystrophies are now beginning to be understood. The key structure in muscular dystrophies is the muscle membrane. Most muscular dystrophies are due to break down of the network of fibrous proteins that bind myofibers to the matrix and stabilize the sarcolemma during contraction and relaxation. Some of these proteins are located in the muscle fiber just inside the sarcolemma (dystrophin); others are embedded in the sarcolemma (sarcoglycans); and others are located in the basement membrane outside the sarcolemma (alpha-dystroglycan, merosin). Loss of the integrity of this network causes stress fractures of the sarcolemma to develop during muscle contraction. Influx of calcium through these breaks activates proteolytic enzymes leading to autodigestion of the sarcoplasm (myonecrosis). Defects of dystrophin cause the Duchenne and Becker muscular dystrophies. Abnormalities of sarcoglycans cause some limb girdle dystrophies. Deficiency of basement membrane proteins such as a2 laminin results in congenital muscular dystrophy. Based on these insights, the phenotypic classification is being replaced by a genetic-molecular classification. For instance, Duchenne and Becker muscular dystrophies are dystrophinopathies, several limb-girdle dystrophies are sarcoglycanopathies, etc. The clinical, pathological, and molecular aspects of the most common dystrophies are briefly described below.

DYSTROPHINOPATHIES

Duchenne muscular dystrophy. Left:Dystrophin immunostain; right:Spectrin (control) immunostain.

Duchenne and Becker muscular dystrophies (DMD-BMD) are caused by mutations of dystrophin, the largest known human gene, located on chromosome Xq21. They are transmitted in an X-linked pattern, i.e. from mother to son. Mothers and daughters are carriers. They have one normal and one abnormal copy of the gene. Male children who inherit the defective copy develop full blown disease. In DMD, dystrophin is absent. In BMD, it is severely reduced and of an abnormal molecular structure. The abnormality can be demonstrated by treating sections of muscle with antibodies to dystrophin (see figure above). In normal muscle, all muscle fibers show a strong reaction along the sarcolemma. In DMD (left), no reaction is seen. DMD muscle stained with antibodies to other membrane proteins such as spectrin (right) shows a normal reaction. In BMD, parts of muscle fibers have dystrophin, and other parts do not. The female carriers of DMD have a mixed population of myofibers, some with dystrophin and some without.

Early myonecrosis

The key pathology of DMD is myonecrosis. At an early phase, necrotic fibers appear homogeneous and deeply eosinophilic. Macrophages then enter necrotic myofibers and remove the debris. Breaks of the sarcolemma allow also CK and other intramuscular proteins to leak out into the interstitial fluid and serum in massive amounts. Myofibers are huge cells. The membrane changes and myonecrosis in dystrophinopathies do not involve the entire muscle fiber, but only parts of it. Myonuclei in unaffected segments divide, activate synthesis of new myofilaments and repair the damage. However, because of the inherent molecular defect, repair is incomplete and cannot keep up with necrosis. Gradually muscle is lost, causing severe weakness. Lost muscle is replaced by fat and fibrous tissue. The latter forms a fibrous scar that permeates muscle and causes stiffness and contractures.

These changes are most severe in DMD in which clinical abnormalities begin in early childhood. At an early stage, some muscles, especially the calves, may appear large (pseudohypertrophy) due to compensatory hypertrophy of non-affected myofibers and increased fat. As the disease progresses, muscle is gradually lost. Patients are usually confined to a wheelchair by 10-12 years. Death usually occurs by the end of the second decade due to respiratory insufficiency and other complications. In BMD, symptoms begin later and the disease is more protracted. Some patients have a nearly normal lifespan. Dystrophin mutations cause also dilated cardiomyopathy. Twenty percent to 40% of females with dystrophin mutations have mild muscle disease or a dilated left ventricle. Even if asymptomatic, they often show mild elevation of CK and subtle changes in the muscle biopsy.

Myonecrosis	Myofiber loss

The key laboratory abnormality of DMD and BMD is severe CK elevation. Because the fibers of each motor unit are destroyed gradually, the EMG shows low voltage and short duration motor unit potentials or polyphasic potentials corresponding to residual myofibers within each motor unit. The muscle biopsy shows myonecrosis, phagocytosis of necrotic fibers, regeneration, and non-specific structural changes (central nuclei, split fibers, atrophic and deformed fibers). Increased endomysial connective tissue and fat are also seen. In BMD, the changes are milder. In a 5- year-old boy with proximal weakness, pseudohypertrophy of the calves, a CK of 6,000 and the above biopsy findings, the diagnosis is hardly in doubt. Dystrophinopathy can be confirmed by immunohistochemistry and DNA analysis. The same methods can be used for carrier detection. Because dystrophin is also present in myocardial fibers, a similar process gradually damages the myocardium causing a clinically significant or fatal cardiomyopathy.

LIMB GIRDLE MUSCULAR DYSTROPHIES

The limb-girdle muscular dystrophies (LGMDs) are a genetically heterogeneous group. Ninety percent are autosomal recessive and 10% autosomal dominant. As a group, they are less frequent than the dystrophinopathies and are milder clinically, i.e., they begin in adolescence or adulthood and have a slower progression. However, a subset of autosomal recessive LGMDs which has been called “severe childhood autosomal recessive muscular dystrophy” (SCARMD) is almost as severe as Duchenne dystrophy is. The molecular defects that cause most LGMDs are now known. Many cases in the SCARMD group are caused by deficiencies of sarcoglycans. Other LGMDs are caused by mutations of other proteins the function of which is poorly understood. Most of these proteins have a close association with the sarcolemma suggesting that the pathogenesis of muscle damage has to do with a membrane abnormality. A relatively large group of autosomal recessive LGMDs are caused by mutation of calpain 3, a protease located in the contractile portion of myofibers. Caveolin-3 and dysferlin mutations also cause LGMD.

CONGENITAL MUSCULAR DYSTROPHIES

The most frequent of these dystrophies is caused by mutations of the extacellular matrix protein laminin a2 (merosin). Another group of CMDs is caused by mutations of enzymes that glycosylate alpha-dystroglycan. Abnormal glycosylation results in defective linkage of alpha-dystroglycan to matrix proteins, thus impairing the stability of the muscle membrane. Dystroglycan is also expressed in brain tissue, and is important for the normal migration and layering of cortical neurons. As a consequence, in addition to muscle disease, which is present at birth, many (CMD) patients also have neuronal migration defects, especially pachygyria, and visual abnormalities.

MYOTONIC DYSTROPHY

Ring fiber and central nuclei

Myotonic Dystrophy is an autosomal dominant muscular dystrophy characterized by weakness and stiffness, more pronounced in facial and distal muscles, and by increased muscle excitability. Atrophy and weakness of facial muscles, ptosis, and frontal baldness produce a characteristic facial appearance. Myotonia (prolonged muscle contraction) occurs spontaneously or is elicited by voluntary activity or by mild stimulation, such as tapping on a muscle (percussion myotonia). The EMG shows characteristic repetitive discharges. In many cases, a handshake is enough to establish the diagnosis (the myotonic patient cannot let go). Symptoms appear in adolescents or young adults, but no age is spared. At times, congenital myotonic dystrophy, transmitted from the mother, causes severe, even fatal hypotonia, weakness, and respiratory insufficiency in newborn babies. In addition to muscle disease, patients with myotonic dystrophy have cataracts, cardiac arrhythmias, testicular atrophy, and diabetes. Weakness is progressive. The biopsy shows atrophy of type 1 fibers, a profusion of central nuclei (normally myonuclei are under the sarcolemma), and ring fibers. None of these changes are diagnostic individually, but their combination strongly suggests myotonic dystrophy.

There are two genetic forms of myotonic dystrophy, DM1, and DM2. They are similar in most respects, except that in DM1 weakness is predominantly distal and in DM2 proximal. DM1 is caused by a CTG trinucleotide expansion in the DMPK (Dystrophia Myotonica Protein Kinase) gene on chromosome 19q13. In DM1, this gene is expanded over 37 CTG repeats. The more repeats, the more severe the dystrophy and the earlier the onset of symptoms. Thus, 100-150 repeats cause myotonia and cataracts, 150-1000 cause full blown myotonic dystrophy, and over 1500-2000 repeats cause neonatal myotonic dystrophy. As with other diseases caused by trinucleotide repeats, the onset of the disease is earlier with each successive generation (anticipation). DM2 is caused by a CCTG expansion of the ZNF9 (Zink Finger Protein 9) gene on 3q21. Neither mutation affects the coding portion of these proteins and it is not kown how these mutations affect muscle and other organs.

CONGENITAL MYOPATHIES

Congenital myopathies are primary muscle disorders. Unlike muscular dystrophies, which are caused by defects of the muscle membrane, most congenital myopathies are due to mutations of contractile and structural proteins, which result in structural abnormalities of myofibers and accumulation of abnormal proteins in the sarcoplasm. There are numerous congenital myopathies and types of inclusions. The most common ones are nemaline, centronuclear, central core, and myofibrillar myopathy.

Nemaline myopathy	Nemaline myopathy	Centronuclear myopathy	Centronuclear myopathy

Nemaline or rod body myopathy (Greek nema, thread) shows small and disorganized myofibers which contain rod-shaped structures composed of a-actinin, the main protein of Z-bands. It is caused by mutations of several genes that encode components of thin filaments. Centronuclear (myotubular myopathy) is characterized by small myofibers with central nuclei, and central areas without contractile filaments, like immature fetal muscle. The X-linked form of this myopathy may be fatal in infancy. Severe congenital myotonic dystrophy may have a similar appearance. In central core and multi-core myopathy, myofibers have one or more central areas of disorganized filaments without mitochondria. Myofibrillar or desmin-related myopathies are characterized by accumulation of abnormal fibrillar material composed of desmin and other proteins.

Congenital myopathies cause severe, sometimes fatal, hypotonia and weakness at birth. Once patients get over the neonatal period, the disease is usually either static or slowly progressive and may be compatible with a normal life span. Patients often have proximal and facial weakness, dysmorphic facial features, kyphoscoliosis, and other physical problems, and lag behind peers in physical prowess. The nomenclature of congenital myopathies is based on their pathological changes. They are clinically and genetically diverse. For instance, nemaline myopathy has infantile, juvenile, autosomal recessive, and autosomal dominant variants. The diagnosis can only be made by detecting the specific structural abnormality on a muscle biopsy. Congenital myopathies are rare, but they are important because they cause neonatal hypotonia. The differential diagnosis of neonatal hypotonia includes also perinatal asphyxia, metabolic disorders, congenital CNS abnormalities, and spinal muscular atrophy (Werdnig-Hoffmann disease).

Updated: September, 2006