Laboratory Issues

There are many molecular pathology laboratories offering testing for the homozygous deletion of SMN1. The majority of laboratories utilize RG-PCR of exon 7 and PCR of exon 8 in two separate reactions, with restriction enzyme digestion to differentiate SMN1 from SMN2 PCR products, and report results for both SMN1 exons 7 and 8. Some laboratories only test for exon 7 deletion. Carrier testing is being offered by a fewer number of laboratories. Proficiency testing for the homozygous SMN1 deletion is offered by the CAP. These proficiency specimens are sent to participants twice per year. External proficiency testing is not available for dosage carrier testing.


Mitochondria are semiautonomous replicating cellular organelles with their own genetic material. Each mitochondrion contains multiple copies of the mitochondrial DNA genome (mtDNA), with replication, transcription, and translation machineries separate from the cellular machineries for these functions. Human mtDNA encodes 13 polypeptides of the respiratory chain subunits, 28 ribosomal RNAs (rRNAs), and 22 transfer RNAs (tRNAs) in a circular double-stranded genome of approximately 16.5 kb.

Unlike nuclear DNA, in which each cell contains pairs of chromosomes, one of maternal and the other of paternal origin, mtDNA is inherited exclusively from the mother. This type of transmission is called maternal inheritance. Although both sexes are equally affected by mitochondrial diseases, inheritance of the disorder is from the mother. Mitochondrial mutations are often present in only some of the mtDNA molecules of a cell (heteroplasmy). Hetero-plasmy occurs because mitochondria segregate randomly into daughter cells during mitosis, which results in cells containing both mutant and wild-type mtDNA. Thus, the proportion of heteroplasmic mutation may vary widely between different tissues or even between different cells of the same tissue. The proportion of mutant to wild-type mtDNA plays a role in determining the clinical variability and severity often observed in the mitochondrial disorders. However, the phenoytpe-genotype correlation in the mitochondrial disorders is complex and is influenced by age, the type and extent of respiratory chain disruption caused by the mutation, and the tissue-specific threshold for the pathogenic effect.

The term "mitochondrial encephalomyopathies" is used to describe mitochondrial disorders in which both muscle and the central nervous system (CNS) are affected. These disorders are multisystemic, with diverse clinical features due to defects in the mitochondrial function. This chapter is restricted to those disorders in which the mutation event involves mtDNA, in contrast to the many nuclear genetic disorders that result in mitochondrial pathology. This occurs when the nuclear encoded protein functions in the mitochondria. This chapter discusses Kearns-Sayre syndrome (KSS), mitochondrial encephalomyopathy with lactic acidosis and strokelike episodes (MELAS), and myoclonic epilepsy with ragged-red fibers (MERRF).

Kearns-Sayre Syndrome

The common features of KSS include progressive external ophthalmoplegia (PEO), pigmentary degeneration of the retina, and defects of cardiac conduction.36 The typical affected patient presents before the age of 20 years with PEO and pitosis. This is followed by the pigmentary retinal degeneration and cardiac conduction block. Other features of the disorder may include ataxia, deafness, dementia, and diabetes mellitus. The most common type of mutation found in KSS is a deletion of mtDNA (Figure 7-4), and almost of all these deletions occur sporadically.37 Approximately one third of KSS cases are due to a common 4977 bp deletion, which is associated with direct repeats at the deletion junction. The severity of KSS depends on the extent of heteroplasmy and the tissue distribution of structurally altered mtDNA. An extreme KSS phenotype occurs when the frequency of deleted mtDNA in muscle cells is greater than 85%. In contrast, when lower levels of hetero-plasmy for the deletion are present, then PEO may be the only symptom.

Mitochondrial Encephalomyopathy with Lactic Acidosis and Strokelike Episodes

MELAS patients are usually normal at birth but develop stunted growth, intermittent vomiting, seizures, and recurrent cerebral insults resembling strokes during the first years of life.38 An episodic course follows, with recurrent strokelike episodes, only partial recovery, and eventual deterioration, with death from respiratory failure often before 20 years of age. Milder adult-onset cases have been reported. Approximately 80% of all MELAS cases are the result of an A3 243 G point mutation in the mtDNA gene encoding tRNALeu.39 The point mutation alters the normal structural conformation of the tRNA, thereby impairing protein synthesis. The A3243G mutation occurs in the het-eroplasmic state, with variation among different tissue types. When the mutation is present in greater than 90% of the mtDNA of the muscle tissue, there is increased likelihood of recurrent strokes and classic MELAS manifestations. However, when the heteroplasmy of the mutation is less than 90%, later onset and more moderate symptoms may occur.

Myoclonic Epilepsy with Ragged-Red Fibers

MERRF is a rare maternally inherited disorder in which the full expression includes muscle weakness, myoclonus, generalized seizures, ataxia, and deafness.40 The hallmark morphologic change seen in the muscle biopsy is the ragged-red fibers. The term "ragged-red fibers" refers to large clumps of abnormal mitochondria that accumulate beneath the sarcolemma and are stained red with the Gomori trichome stain. The majority of MERRF cases are the result of a point mutation (A8344G) in the tRNALys gene. The MERRF mutation, like the MELAS tRNALeu mutation, diminishes overall mitochondrial protein synthesis. Similar to the other mitochondrial disorders, a more classic MERRF phenotype is observed when the mutation is present at higher levels in the muscle and nerve.

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