Background Histopathology & Immunohistochemistry

BACKGROUND AND CLINICAL INFORMATION: Head

Summary: Nemaline myopathy affects predominantly infants and children although cases with fatal neonatal onset and adult onset are also known. This disease is generally considered benign but significant disability may be present. Cardiorespiratory insufficency can also occur in infantile cases. Four clinical forms are being recognized. Childhood cases usually start as infantile hypotonia, some of them can be extremely severe and fatal. Muscles of proximal and distal limbs, neck and trunk are affected. The bulbar muscles innervated by cranial nerves are spared but the facial muscles may be severely affected. It is a genetically diverse group of diseases that share the common feature of nemaline rod (nemaline bodies) formation. The pattern of inheritance is unclear and has been suggested to be autosomal dominant or autosomal recessive; a variety of mutations have been described.

Clinical forms: 4 major clinical forms

Fatal infantile form Classic childhood form Childhood autosomodominant form Late onset or adult form

Fatal infantile nemaline myopathy: Assisted respiration is usually necessary from birth and patients usually die in 6 months because of pneumonia. These patients may also have arthrogryposis and cardiomyopathy.

Childhood or “classic” nemaline myopathy:

· General: This entity is considered benign although disability may be present although respiratory deficiency particularly nocturnal hypoventilation may cause early death. Mild hypotonia may be present in the newborn period and muscular hypotonia is always during the first year of life. Most cases present as a mild, non-progressive myopathy although weakness may be severe. Motor developmental milestones are delayed but new motor skills are acquired as the child grows. Serum CPK is normal.

· Muscle affected: Muscles of proximal and distal limbs, neck and face are also affected.

· Characteristic facial features: Long face, high arched palate, and a tent shaped mouth.

· Skeletal: Associated skeletal changes such as kyphoscoliosis, pigeon chest, pes cavus. Scoliosis and lordosis may complicate the clinical situation.

· Respiratory problem: Patients may develop respiratory deficiency and are susceptible to repeated severe respiratory infections.

· Cardic problem: They are more commonly seen in cases that are over 20 years of age [Muller-Hocker J et al., 2000].

· Asymptomatic carrier: Asymptomatic family members may have nemaline bodies in muscle fibers.

Late onset or adult form of nemaline myopathy: This is rare. Patients usually present as a limb-girdle syndrome with onset in the fifthe and sixth decade. Distal weakness may predominate and the lower extremities are usually more affected. Lumbar lordosis is particularly prominent. “Marfanoid” body habitus is iseen in adolescent and adult cases.

Childhood autosomal dominant form:

Hyaline and nemaline rods: Two sublings with cardioneuromyopathy with hyaline masses and nemaline rods have been reported. [Selcen D et., Ann Neurol. 2002 Feb;51(2):224-34.].

Genetics: Several genes are involved in nemaline myopathy. [Bornemann A and goebel HH, 2001, Brain Pathology 11:206-217]. Nemaline with mutation in TPM3 or TPM2 is less common than with mutation in NEB gene. Almost all of the involved genes (TPM3, NEB, ACTA1, TPM2, Troponin T1) are components of thin filament.

NEM 1 (on 1q21-23): a-tropomyosin (TPM3). A missense mutation (Met9Arg) on this gene is associated with a juvenile slowly progressive form in one family [Laing NG et al., 1992; Tan P et al., 1999], related to autosomal dominant and recessive diseases.
NEM 2 (on 2q21.2-22): Nebulin (NEB) gene, related to autosomsal recessive diseases.
NEM 3 (on 1q42.1): Sarcomeric actin (ACTA1) gene [Jungbluth H et al., 2001], related to autosomal dominant and recessive diseases.
NEM 4 (on 9p13.2): b-tropomyosin (TPM2) gene.
Troponin T1 (in 19q13.4): Mutations have been described in one family [Johnston JJ et al., 2000]. This is seen in the Armish families [Johnston JJ et al., 2000].
Ryanodine receptor gene: In one family, mutation of ryanodine receptor gene have caused myopathy that have features of both central core disease and nemaline myopathy [Scacheri PC et al., 2000]. Ryanodine receptor is involved in central core disease.

HISTOPATHOLOGY AND IMMUNOHISTOCHEMISTRY: Head

Childhood cases:

Nemaline bodies (nemaline rods): Nemaline bodies have biochemical and structural properties of Z-discs and arise as a product of polymerization or proliferation of the lattice of the Z-disc. Although nemaline bodies are the hallmark of nemaline body diseases, they are also seen in fibers that have lost thick filaments, myopathy associated with certain types of HIV virus, some cases of dermatomyositis and polymyositis, and severe steroid myopathy.

· Routine sections: The proportion of fibers being affected is highly variable among different muscle and also within the same biopsy. The diameter of muscle fibers can vary from atrophic to hypertrophic. In childhood cases, nemaline bodies are seen in most type 1 fibers and in many cases also in some type 2 fibers. They probably do not occur in intrafusal fibers. With HE stain, they may appear as vaguely defined pink refractile bodies. Nemaline bodies bearing cells may be quite atrophic. With Gomori’s modified trichrome, they appear as very dense dark red granules or tiny rods that are sharply demarcated. Althogh they tend to accumulate beneath the sarcolemma, centrally located rods are not uncommon. Nuclear rods can also be present. They generally measure about 1-3 mm in width and 3-6 mm in length.

· Histochemistry: Nemaline bodies have no enzyme reactivity. This distinguishes them from ragged red fibers.

· Immunostaining: The rods are reactive to a-actinin and to a-actin. Desin may be demonstrated around the rods but not within the rods.

· Semithin sections: Rods are densely osmophilic and sharply demarcated. Their origin from Z-disc can be well demonstrated in longitudinal sections.

· EM: They have longitudinal striations with periodicity of 8 nm. Small nemaline bodies are often connected with the Z-discs but larger nemaline bodies are more often not. Thin filaments protrude from the end of nemaline bodies that appears like thin filaments protruding from Z-disc. They have a tetragonal arrangment on cross sections with the squares about 100 nm in dimensiona and the filaments about 3 nm in diameter. Nuclear nemaline bodies do not have protruding thin filaments.

Fiber affected: There is usually a paucity or complete absence of type 2 fibers. The type 1 fibers tend to be small.

Changes with age: As the age of the patient increase, there is increased variation of fiber diameter with both atrophic and hypertrophied fibers seen. The number of type 2 fibers diminishes and more centrally located rods are seen.

Fatal infantile nemaline myopathy and adult nemaline myopathy have slightly different histologic fetures.

Cardiac muscle: Nemaline bodies may be seen in cardiac muscle.