A collaborative research project was begun in 1994 between the Department of Anaesthesia and Intensive Care, Palmerston North Hospital, and the Institute of Molecular BioSciences, Massey University, Palmerston North.
The ultimate aim of this project is to develop DNA-based tests that will identify susceptibility to Malignant Hyperthermia (MH). The approach is multi-disciplinary and includes clinical diagnosis as well as molecular genetic analysis of DNA prepared from individuals of families known to be susceptible to Malignant Hyperthermia (MHS).
DNA samples from families with MHS members has been used for genetic screening. As a causative factor in MH, the ryanodine receptor gene (RYR1) has been the most widely studied in overseas laboratories. Several mutations in the RYR1 gene which result in altered amino acid residues in the skeletal muscle ryanodine receptor have been shown to be associated with MHS. Our initial approach was to search for these mutations in the New Zealand families known to be susceptible to MHS. The most common of these mutations have been studied in each of the 37 New Zealand families for which we have been able to collect blood samples. To date we have identified three of these mutations in three separate small families. In the course of this work we have identified three novel mutations in another three families. From the location of these mutations and the co-segregation of the mutation with the MHS phenotype, we are confident that these novel mutations are causative of MH.
The major thrust of this research project has been focused on one very large Maori family. Several members of this family have suffered fatal reactions to anaesthesia due to MH. We began the study by screening individuals of this family for the known mutations, none of which were detected. We then embarked on a genetic linkage analysis by examining seven markers that are within or close to the RYR1 locus. The results of this study indicated that MHS is almost certainly linked to RYR1 in this family. DNA sequence analysis of the entire coding region of the ryanodine receptor gene has identified another novel mutation that results in an amino acid change in the protein. We have now confirmed that affected members of this family carry this mutation and that it does not occur in the general population. In the course of this work we identified an additional novel mutation close by in yet another Maori family. We are now in a position to offer genetic diagnosis of MH for several New Zealand families.
Current Research Projects
- Screening the three mutation "hot-spot" regions of the RYR1 gene by RT-PCR of RNA prepared from muscle biopsy followed by direct DNA sequence analysis using an ABI Prism 377-64 and Big Dye terminators.
- Confirmation of the causative nature of the novel mutations by biochemical and physiological assays of calcium release using recombinant DNA methodologies and tissue culture of mammalian cells.
- Genome wide scan of a discordant branch of a large Maori family to identify alternative genetic loci associated with the MH phenotype.
Stowell, K. M., Brown, R. L., James, D., Couchman, K.G., Hodges, M. and Pollock, A. N. Malignant Hyperthermia in New Zealand. New Zealand Bioscience May p12-17 (1999)
Chamley, D., Pollock, A. N., Stowell, K. M. and Brown, R. L. Malignant Hyperthermia in Infancy and identification of a novel RYR1 mutation. British Journal of Anaesthesia, 84, 500-504 (2000)
Brown, R. L., Pollock, A. N., Couchman, K. G., Hodges, M., Waaka, R., Lynch, P., McCarthy, T. V., and Stowell, K. M. A novel ryanodine receptor mutation and genotype-phenotype correlation in a large malignant hyperthermia New Zealand Maori pedigree. Human Molecular Genetics, 9, 1515-1524. (2000)
Davis, M., James, D., Norgate, M., Marston, R., Laing, N., Perlman, D., Brown, R., Dickson, A, Pollock, N., Stowell, K. Malignant Hyperthermia Associated with congenital abnormalities and a Novel RYR1 mutation in New Zealand and Australian Pedigrees. British Journal of Anaesthesia, 88(4), 508-515 (2002)
Pollock, N., Couchman, K., Langton, E., Stowell K., Waddington M. Suspected Malignant Hyperthermia Reactions in New Zealand. Anaesthesia Intensive Care, 30, 453-611 (2002)
Australian and New Zealand College of Anaesthetists
New Zealand Lotteries Grants Board
Palmerston North Medical Research Foundation
The Paykel Trust
Massey University Research Fund