Cardiomyopathy and carnitine deficiency

https://doi.org/10.1016/j.ymgme.2008.02.002Get rights and content

Abstract

Carnitine is essential for the transfer of long-chain fatty acids across the mitochondrial membrane for subsequent β-oxidation. A defect in the high-affinity carnitine transporter OCTN2 causes autosomal recessive primary carnitine deficiency that can present with hypoketotic hypoglycemia, mainly in infancy or cardiomyopathy. Heterozygotes for primary carnitine deficiency can have mildly reduced plasma carnitine levels and can develop benign cardiac hypertrophy. In animal models, heterozygotes for this disease have a higher incidence of cardiomyopathy with aging. This study tested whether heterozygosity for primary carnitine deficiency was associated with cardiomyopathy. The frequency of mutations in the SLC22A5 gene encoding the OCTN2 carnitine transporter was determined in 324 patients with cardiomyopathy and compared to that described in the normal population. Missense variations identified in normal controls and patients with cardiomyopathy were expressed in Chinese Hamster Ovary cells to confirm a functional effect. Exons 2–10 of the SLC22A5 gene were amplified by PCR in the presence of LCGreen I and analyzed by dye-binding/high-resolution thermal denaturation. Exon 1 of the gene was sequenced in all patients. Heterozygosity for a few variants (L144F, T264M, I312V, E317K, and R488H) was found in 6/324 patients with cardiomyopathy. Expression of these variants in CHO cells indicated that T264M decreased, E317K increased, while L144F, I312V, and R488H did not significantly affect carnitine transport. Expression in CHO cells of all the variants identified in a normal population indicated that only two had a functional effect (L17F and Y449D), while L144F, V481I, V481F, M530V, and P549S did not change significantly carnitine transport. The frequency of variants affecting carnitine transport was 2/324 patients with cardiomyopathy (0.61%) not significantly different from frequency of 3/270 (1.11%) in the general population. These results indicate that heterozygosity for primary carnitine deficiency is not more frequent in patients with unselected types of cardiomyopathy and is unlikely to be an important cause of cardiomyopathy in humans.

Section snippets

Materials and methods

The protocol for DNA studies was approved by the University of Colorado and by the University of Utah Institutional Review Board. Informed consent was obtained from all patients or their parents prior to DNA studies. GenBank sequence AB016625.1 was used as reference for the gene, NM_003060.2 was used as the reference sequence for the cDNA. Genomic DNA was obtained from 324 patients with cardiomyopathy part of the cardiomyopathy registry [14], [15]. About 10% of these patients had hypertrophic

Results

Direct DNA sequencing of exon 1 of the SLC22A5 gene did not identify any significant variation in the DNA of patients with cardiomyopathy. Screening of the remaining exons (2–10) of the SLC22A5 gene by high-resolution melting analysis in 324 patients with cardiomyopathy identified 38 variants out of 2916 exons screened. An example of the melting curve of selected exons is shown in Fig. 1. DNA sequencing identified polymorphisms not affecting the amino acid residues and described in the normal

Discussion

Primary carnitine deficiency impairs the accumulation of carnitine within organs and tissues [9]. In the heart, carnitine is essential for normal fatty acid β-oxidation and even partial deficiency could lead to organ dysfunction. Heterozygous animal models of primary carnitine deficiency have a higher rate of cardiomyopathy with aging [11], [12]. Humans heterozygous for primary carnitine deficiency have mildly reduced plasma carnitine levels due to increased urinary losses [8] and might develop

Acknowledgments

We thank Idaho Technology Inc. for allowing the use of the HR-1 High-Resolution Melter and for valuable help in optimizing the screening assay. We also thank the Familial Cardiomyopathy Study subjects for their participation. This study was supported by Grant-in-Aid 0455086Y from the American Heart Association and in part by Grants DK 53824 and 1K23Hl67915-01A1 from the National Institutes of Health.

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