Increased muscle coenzyme Q10 in riboflavin responsive MADD with ETFDH gene mutations due to secondary mitochondrial proliferation
Introduction
Multiple acyl-coenzyme A dehydrogenation deficiency (MADD, OMIM 231680), also known as glutaric aciduria type II, is an autosomal recessive inherited disorder with a heterogeneous phenotype. MADD cases are mostly divided between a lethal neonatal-onset form, with or without congenital anomalies, and a mild late-onset form. The latter is often due to mutations in ETFDH gene (electron transfer flavoprotein dehydrogenase, HGNC 3483), which encodes electron transfer flavoprotein:ubiquinone oxidoreductase (ETF:QO) and causes lipid storage myopathy. The clinical manifestations of late onset MADD are quite homogeneous including proximal myopathy and exercise intolerance, and it is worth mentioning that patients in both East and South China have frequently experienced episodes of vomiting and muscle pain [1], [2], [3].
ETF:QO, is synthesized as a 67-kDa precursor which is targeted to mitochondria and processed to a 64-kDa mature monomer integrated in the inner mitochondrial membrane. In the process of fatty acid β-oxidation and amino acids catabolism in mitochondria, ETF:QO is a critical element of the pathway by which electrons enter the coenzyme Q10 (CoQ10) pool, and then is passed on to complex III of the mitochondrial respiratory chain (MRC) (Fig. 1A) [4], [5]. Theoretically, a defect in ETF:QO protein can affect the transfer of all electrons produced by the β-oxidation enzymes, leading to the MADD phenotype.
ETF:QO contains three functional domains, respectively responsible for FAD, CoQ10, and 4Fe4S binding (Fig. 1B). In our previous study, we detected mutations in FAD and CoQ10 domains, but not in 4Fe4S domain [1]. Therefore, it seems that either both FAD and CoQ10 domain are important to maintain the enzyme function of ETF:QO or mutations in 4Fe4S domain might cause fetal death that couldn't be detected in late-onset MADD. FAD is the prothetic group of ETF:QO, by which electrons from β-oxidation are transferred to CoQ10 [4], [5].
It is therefore important to confirm CoQ10 levels in MADD patients. CoQ10 is synthesized in mitochondrial inner membrane and is composed of a benzoquinone and a decaprenyl side chain [6]. Four major functions are attributed to this compound: i. carriage of electrons from complexes I and II to complex III in the MRC; ii. potent lipophilic antioxidant defense in all cell membranes (in the reduced form, ubiquinol) [7]; iii. pyrimidine nucleoside biosynthesis, and iv. modulation of apoptosis and mitochondrial uncoupling protein [8].
However, the issue of CoQ10 levels in MADD patients is still a matter of debate. ETFDH mutations were reported to cause the myopathic form of coenzyme CoQ10 deficiency in five MADD patients [9]. While Liang et al. showed normal CoQ10 levels in two patients with ETFDH mutations [2]. Consequently, it is necessary to investigate changes in CoQ10 levels in a larger group of MADD patients with ETFDH mutations.
In this study, we characterized 34 patients with ETFDH mutations to further elucidate the probable functions of the cofactor CoQ10 in electron transfer pathway through ETF:QO. It is likely that this work will promote the understanding of MADD pathomechanisms and the therapeutic strategy of CoQ10 supplementation.
Section snippets
Patients and controls
Muscle biopsy was performed after obtaining written informed consent under study protocols approved by the medical Ethics Committee of Qilu Hospital, Shandong University. This study recruited 34 biopsied muscle samples from riboflavin responsive patients with ETFDH gene mutations and 32 normal controls who, after all tests were performed, were considered free of respiratory chain disease in the Neuromuscular Center of Qilu Hospital between 1995 and 2010. The recruitment criteria of 34 patients
CoQ10 levels and evaluation of mitochondrial mass
CoQ10 data from HPLC analysis was normalized to muscle mass (Fig. 2A), total protein (Fig. 2B) and citrate synthase (Fig. 2C). CoQ10 levels in 34 MADD patients with ETFDH gene mutations were increased to 1.92 and 1.64 folds compared with 17 normal controls (p < 0.001) when normalized to muscle mass and total protein, respectively. The CoQ10 pool in muscle from MADD patients seems enlarged.
However, when CoQ10 data was normalized to citrate synthase, a marker of mitochondrial mass, there was no
Discussion
In this work, we presented muscle CoQ10 levels in 34 MADD patients with ETFDH gene mutations who were well responsive to isolated riboflavin therapy. CoQ10 level was normalized to muscle mass, total protein and citrate synthase. None of our patients had been placed on CoQ10 or riboflavin therapy before this study. Patients with ETFDH gene mutations had increased CoQ10 levels compared with normal controls when normalized to muscle mass or total protein. But given the facts that the protein
Conflict of interest
None.
Acknowledgments
The authors thank the patients and their families for their contribution in this work. We are grateful to Dr. Liou CW of the Department of Neurology, Chang Gung Memorial Hospital, Taiwan and Dr. Guo RC of the Institute of Clinical Pharmacology, Qilu Hospital of Shandong University, Jinan for the technique support. The authors also thank Mr. Ash DB of Motor Neuron Center, Columbia University Medical Center, New York for his linguistic suggestions.
Details of funding
This work was supported by the
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These authors contributed equally to this work.