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August 1996
Volume 60 |
Number 8
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| Molecular Genetics
and Diagnosis of Malignant Hyperthermia |
Jeffrey E. Fletcher, Ph.D.
Malignant hyperthermia (MH) was identified as a genetic disorder
based on a family in Australia; it was first reported in Lancet
in the early 1960s by Denborough and collaborators.1
The first diagnostic tests were reported in the early 1970s based
on the greater in vitro sensitivity of fiber bundles from
biopsied vastus muscle to caffeine2
and to halothane.3 The contracture
tests underwent considerable refinement during the 1980s, and
the North American and European MH groups now have each established
a specific protocol to provide uniform testing. Alternative tests
have not withstood validation.
Studies of the genetics of MH progressed most rapidly over the
period of 1990-91. At that time, linkage was made to chromosome
19q13.1 close to or at the site encoding the
ryanodine receptor (ryr1), which is the Ca2+
release channel of skeletal muscle. These findings were published
independently in Nature in 1990 by MacLennan of Toronto4
and McCarthy of Ireland.5 Shortly
thereafter, the disorder was recognized as heterogeneous. In 1991,
Levitt6 first reported that more than
one gene could cause MH. This observation was quickly confirmed
by a number of groups internationally. Thus began an intensive
search for additional genes not linked with chromosome 19 that
could cause MH and account for 50 percent to 75 percent of the
occurrences of MH. Currently, linkage has been reported to chromosomes
3, 7, 17 and 19. Eight mutations in ryr1 and one mutation
in the sodium channel have been described in the literature as
possibly causal of MH. An additional eight mutations in ryr1
appear to have been found. To appreciate the significance of the
above findings, a few important terms need to be understood.
Phenotype in the simplest sense refers to whether or not a person
is susceptible to MH. In most cases, there are no obvious indicators
of MH susceptibility in the absence of anesthesia or a diagnostic
test. While a convincing episode of MH under anesthesia can be
sufficient to phenotype an individual, this approach obviously
limits the number of families and subjects within a family. These
events are rare, and considerable caution should be exercised
by anesthesiologists with other family members; i.e., the use
of triggering agents should be avoided. Experiencing anesthesia
without difficulty is not sufficient to warrant a negative diagnosis,
since many MH-susceptible patients undergo multiple anesthetics
without exhibiting signs of MH. This means that, in most cases,
we rely on the halothane and caffeine in vitro contracture
test for phenotyping. Although the sensitivity and specificity
of the test are quite acceptable, it is not perfect, and the invasive
and expensive nature of this test limits the number of subjects
phenotyped within a family.
Linkage analysis is a major tool used in "reverse genetics,"
which is an approach that does not require that the defective
protein be known to identify the gene causing a disorder. Linkage
refers to how close a particular genetic marker is to the gene
determining the MH phenotype. A number of factors go into this
determination, but the accuracy of linkage depends on the reliable
phenotyping of large families and on the occurrence of crossovers,
or the exchange of DNA between homologous segments of the paternal
and maternal chromosomes.
It is the requirement for large, accurately phenotyped pedigrees
that has been a major barrier to further progress in MH. Linkage
should not be interpreted as absolute proof that a particular
region of DNA or a protein encoded within that region is the causative
factor in MH. In the case of MH, the linkage to chromosome 19q13.1
has been verified by several laboratories, suggesting that a protein
encoded in that region of the genome, possibly ryr1, is
likely causative of MH. Linkage of MH to any other chromosomes
is far less certain and awaits further verification.
DNA can and frequently does have alterations that do not significantly
affect the function of the protein; these would be termed "polymorphisms."
Mutations are alterations in the genetic code that significantly
affect the function of a protein to the extent that a genetic
disorder could result. While most of the proposed "mutations"
described to date could eventually prove to be polymorphisms,
most investigators would agree that the "porcine" mutation
(conversion of arginine 614 to a cysteine) might be a cause of
human MH. The most controversial aspect surrounding this mutation
is that a pig inheriting only one copy of the porcine mutation
does not exhibit signs of MH, but the human disorder is inherited
dominantly. This mutation is rarely found in human MH (about 1
percent to 5 percent) and has occurred in subjects with normal
diagnostic contracture tests.
Likewise, the mutation has not been observed in some subjects
with a positive contracture test for MH in families in which other
susceptible members have the mutation. Also, one patient having
this mutation had 18 anesthetics (five with halothane and succinylcholine)
and only experienced muscle rigidity (no acidosis) a few times.
Similarly, the clinical picture for the sodium channel mutation
represented rigidity without acidosis, which does not present
a strong case for an MH episode. Overall, if all of the identified
"mutations" are truly causative of MH, then only 15
percent to 25 percent of the total mutations have been identified.
What is the impact of the foregoing on diagnosis? There have been
suggestions that the mutation could be used for diagnosis in families
in which specific "mutations" already have been identified.
While some investigators feel uncomfortable about this suggestion,
most would agree that the use of contracture testing in combination
with genetic testing could ultimately increase our understanding
of this disorder. In the meantime, we are asking why in some families
some subjects with these mutations have a normal diagnostic contracture
test for MH and others with a positive test for MH do not have
the mutation.
Are these mutations truly the cause of MH? Which tests are correct?
We are not certain at this time. Similar questions are being asked
regarding genetic screening in other fields.7
The muscle biopsy and contracture test will continue to be the
major means of diagnosis over the next several years. Large numbers
of families with many family members diagnosed by the contracture
test are required once again to accelerate the genetics studies.
The North American laboratories have begun to explore a large
cooperative venture, and strategies are being considered that
will hopefully provide needed momentum to research in this area.
Bibliography:
1. Denborough MA, Lovell RRH. Anaesthetic deaths
in a family. Lancet. 1960; 2:45.
2. Kalow W, Britt BA, Terreau ME, et al. Metabolic
error of muscle metabolism after recovery from malignant hyperthermia.
Lancet. 1970; 2:895-898.
3. Ellis FR, Harriman DGF, Keaney NP, et al. Halothane-induced
muscle contracture as a cause of hyperpyrexia. Br J Anaesth.
1971; 43:721-722.
4. MacLennan DH, Duff C, Zorzato F, et al. Ryanodine
receptor gene is a candidate for predisposition to malignant hyperthermia.
Nature. 1990; 343:559-561.
5. McCarthy TV, Healy JMS, Heffron JJA, et al.
Localisation of the malignant hyperthermia susceptibility locus
to human chromosome 19q12-13.2. Nature. 1990; 343:562-564.
6. Levitt RC, Nouri N, Jedlicka AE, et al. Evidence
for genetic heterogeneity in malignant hyperthermia susceptibility.
Genomics. 1991; 11:543-547.
7. Hubbard R, Lewontin RC. Pitfalls of genetic
testing. N Engl J Med. 1996; 334:1192-1194.
Jeffrey E. Fletcher, Ph.D., is Professor
of Anesthesiology at Allegheny University of the Health Sciences,
Philadelphia, Pennsylvania.
E-mail the author.
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