Dissertation

Gene Expression and Metabolism in Malignant Hyperthermia Susceptible and Normal Skeletal Muscle

Chang, Leon

2020

Overview

Malignant hyperthermia (MH) is an autosomal dominant, pharmacogenetic disorder primarily caused by RYR1 mutations which result in calcium dysregulation within skeletal muscle. Genetically susceptible individuals are at risk of potentially fatal hypermetabolic reactions when exposed to volatile anaesthetics and the muscle relaxant succinylcholine. MH susceptibility is diagnosed through the in vitro contracture test (IVCT) by challenging muscle biopsies with triggering agents, or by genetic screening for known familial mutations. Genome-wide gene expression was compared before and after IVCT, from MH-susceptible (MHS) and non-susceptible (MHN) skeletal muscle by RNA sequencing. A downregulation of genes related to oxidative phosphorylation (OXPHOS) was observed in MHS samples at baseline, suggesting a metabolic defect. Mitochondrial function was assessed by high resolution respirometry, measuring oxygen consumption rates in permeabilised muscle fibres. Results showed evidence of reduced OXPHOS capacity, complex II deficiency and increased mitochondrial content in MHS muscle at baseline. Exposure to halothane triggered a hypermetabolic response in MHS mitochondria which significantly increased oxygen consumption rates in several respiratory states, whilst MHN samples were unaltered. Genome-wide gene expression and mitochondrial function was also studied at baseline and after halothane challenge, using transgenic mouse models of MH to investigate RYR1 variant-specific effects. At baseline, fatty acid oxidation and mitochondria-related gene expression was downregulated in mice homozygous (HOM) for G2435R-RYR1 and heterozygous (HET) for T4826I-RYR1. In comparison to wild-type, mitochondria from G2435R-RYR1 HOM mice showed an increase in complex I-facilitated OXPHOS and reduced mitochondrial content at baseline. Mitochondria from transgenic mice also showed evidence of increased sensitivity to both halothane and calcium in comparison to wild-type. This study presents evidence of mitochondrial dysfunction in human and mouse MHS skeletal muscle, which is correlated with gene expression changes associated with oxidative metabolism. Functional defects in mitochondria are therefore potential contributors to phenotypic variability observed in MH.

• Publisher: ProQuest Dissertations & Theses
• Subject: Gene expression / Genetics / Genomes / Hyperthermia / Metabolism / Mitochondria / Musculoskeletal system / Mutation