Synergistic heterozygosity in mice with inherited enzyme deficiencies of mitochondrial fatty acid β-oxidation

doi:10.1016/j.ymgme.2004.09.006

Molecular Genetics and Metabolism

Volume 85, Issue 1, May 2005, Pages 7-11

https://doi.org/10.1016/j.ymgme.2004.09.006 Get rights and content

Abstract

We have used mice with inborn errors of mitochondrial fatty acid β-oxidation to test the concept of synergistic heterozygosity. We postulated that clinical disease can result from heterozygous mutations in more than one gene in single or related metabolic pathways. Mice with combinations of mutations in mitochondrial fatty acid β-oxidation genes were cold challenged to test their ability to maintain normal body temperature, a sensitive indicator of overall β-oxidation function. This included mice of the following genotypes: triple heterozygosity for mutations in very-long-chain acyl CoA dehydrogenase, long-chain acyl CoA dehydrogenase, and short-chain acyl CoA dehydrogenase genes (VLCAD+/−//LCAD+/−//SCAD+/−); double heterozygosity for mutations in VLCAD and LCAD genes (VLCAD+/−//LCAD+/−); double heterozygosity for mutations in LCAD and SCAD genes (LCAD+/−//SCAD+/−); single heterozygous mice (VLCAD+/−, LCAD+/−, SCAD+/−) and wild-type. We found that approximately 33% of mice with any of the combined mutant genotypes tested became hypothermic during a cold challenge. All wild-type and single heterozygous mice maintained normal body temperature throughout a cold challenge. Despite development of hypothermia in some double heterozygous mice, blood glucose concentrations remained normal. Biochemical screening by acylcarnitine and fatty acid analyses demonstrated results that varied by genotype. Thus, physiologic reduction of the β-oxidation pathway, characterized as cold intolerance, occurred in mice with double or triple heterozygosity; however, the derangement was milder than in mice homozygous for any of these mutations. These results substantiate the concept of synergistic heterozygosity and illustrate the potential complexity involved in diagnosis and characterization of inborn errors of fatty acid metabolism in humans.

Introduction

Inherited disorders of mitochondrial fatty acid β oxidation represent a challenging group of diseases to diagnose. Children affected by disorders of fatty acid oxidation often present clinically with a stereotypical phenotype. However, at times these patients have no specific diagnostic metabolite patterns, such as urinary organic acids or acylcarnitine profiles, or an unequivocal enzyme deficiency. Vockley and colleagues [1] postulated that these unclear cases for diagnosis may have a genotype representing a condition designated “synergistic heterozygosity.” They proposed that although inborn errors of metabolism are frequently autosomal recessive disorders, heterozygous mutations in two or more related pathways or steps in a single pathway could result in development of a disease phenotype [1]. To illustrate this point, several clinically relevant cases were discussed. Notably, “patient 8” [1] was an infant who died at 3 months of age after a mild bronchopneumonia. Postmortem studies on this infant’s cultured fibroblasts revealed a decreased ability to oxidize myristate (41% of control) and palmitate (71% of control). Molecular studies revealed heterozygous point mutations in both the carnitine transporter (OCTN2) and very long-chain acyl-CoA dehydrogenase genes (ACADVL). Since this is a potentially important disease mechanism that appears difficult to diagnose, we believed it was important to test the genetic/metabolic concept in the available mouse models of mitochondrial fatty acid oxidation enzyme deficiencies. We hypothesized that mice that are double or triple heterozygous for mutations in genes involved in mitochondrial fatty acid β-oxidation would display impaired fatty acid oxidation and energy generation, as evidenced by cold intolerance, and hypoglycemia.

Section snippets

Mice

Male and female BALB/cByJ [short-chain acyl CoA dehydrogenase (Acads) deficient-SCAD−/−] [2], C57BL/6NTTacfBR, 129S6/SvEvTac (B6, 129) (long-chain acyl CoA dehydrogenase (Acadl) deficient) (LCAD−/−) [3], B6,129- (very-long-chain acyl CoA dehydrogenase (Acadvl) deficient) (VLCAD−/−) [4], and wild-type controls were maintained at the University of Alabama at Birmingham. Double and triple heterozygous mice included the following genotypes: VLCAD+/−//LCAD+/−, LCAD+/−//SCAD+/−, or

Results

Within 5 h of starting the cold challenge, 36% of the LCAD+/−//SCAD+/− mice and 33% of the VLCAD+/−//LCAD+/− mice became lethally hypothermic (Fig. 1). Of the VLCAD+/−//LCAD+/−//SCAD+/−mice, 33% became lethally hypothermic (data not shown). These three groups were significantly different (P < 0.001) than the control groups of wild-type and the single heterozygous groups of mice. None of the wild-type mice became hypothermic (Fig. 1). Single heterozygous mice of each individual enzyme deficiency

Discussion

Cold challenge produces a reproducible phenotype in mice homozygous for any of the acyl-CoA dehydrogenase deficiencies [6]. Homozygous mice (LCAD−/−, SCAD−/−, and VLCAD−/−) uniformly experience an inability to thermoregulate when placed in the cold (4 °C) for more than 1 h. This result can be exacerbated by a fast prior to cold challenge. They also become hypoglycemic and presumably hypoketotic when exposed to the cold [6]. This abnormal phenotype makes these mice an excellent model for disorders

Acknowledgments

This work was supported by NIH grants RO1-RR02599, T-32 RR07003, and P30 DK56336.

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Synergistic heterozygosity in mice with inherited enzyme deficiencies of mitochondrial fatty acid β-oxidation

Abstract

Introduction

Section snippets

Mice

Results

Discussion

Acknowledgments

Mol. Genet. Metab.

Trends Genet.

Am. J. Hum. Genet.

Am. J. Hum. Genet.

Mol. Genet. Metab.

Short-chain acyl-coenzyme A dehydrogenase deficiency in mice

Pediatr. Res.

Targeted disruption of a mouse long-chain acyl-CoA dehydrogenase gene reveals crucial roles for fatty acid oxidation

Proc. Natl. Acad. Sci. USA

Gestational, pathologic, and biochemical differences between very long-chain acyl-CoA dehydrogenase deficiency and long-chain acyl CoA dehydrogenase deficiency in the mouse

Hum. Mol. Genet.