Acute Arrhythmias in a Long-Chain 3-Hydroxyacyl-CoA Dehydrogenase Deficiency (LCHADD) Mouse Model
Biochemical/Metabolic and Therapeutics
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Introduction:
Introduction: People with long-chain fatty acid oxidation disorders (LC-FAODs), such as LCHADD (OMIM# 609016), frequently present dilated or hypertrophic cardiomyopathy and can suffer life-threatening arrhythmias. LCHADD is caused by a missense pathogenic variant, c.1528G>C, of the HADHA gene encoding the alpha subunit of the mitochondrial trifunctional protein. In a recent retrospective analysis, we reported an increased incidence of major cardiac involvement in adolescent and young adults with LCHADD where heart failure and life-threatening arrhythmias remain the leading cause of mortality. The etiology of cardiac arrhythmias among people with LC-FAODs is not fully elucidated; therefore, we examined the mechanisms of impaired cardiac function in an LCHADD mouse model.
Methods:
Methods: We previously determined that the LCHADD mice (Hadha c.1528G>C homozygotes) recapitulate human disease by oxidizing more glucose and less fat compared to wildtype (WT) mice and accumulated LCHADD-specific long-chain 3-hydroxy-acylcarnitines in plasma. LCHADD mice have lower ketones when fasted, exhaust earlier during treadmill exercise and have decreased visual performance and retinopathy. On echocardiography, LCHADD mice have early concentric remodeling progressing to hypertrophic and dilated cardiomyopathy with low ejection fraction. To test for arrhythmias, we measured heart rate (HR), blood pressure (BP) and single-lead ECG on 9-month-old LCHADD and WT mice (n= 4-5 per genotype and sex) under mild sedation and stimulation with a single dose of a beta-agonist (isoproterenol; 3 mg/kg).
Results:
Results: LCHADD mice have significantly increased frequency of atrial premature beats, premature ventricular contractions, atrial flutter, atrial fibrillation, and non-sustained ventricular tachycardia after isoproterenol injection compared to WT mice, according to the Lambeth Conventions Guidelines. While HR was similar at baseline and increased with isoproterenol, it did not differ between groups. However, all LCHADD mice returned to baseline faster than WT after isoproterenol injection suggesting an inefficient sympathetic response. Long QRS and long QT intervals were seen in LCHADD male and female mice. Heart weight to body weight ratio was significantly increased in LCHADD hearts. Frozen and fixed heart sections demonstrated significantly increased cardiomyocyte cross-sectional area by H&E, increased collagen deposition by Masson Trichrome, decreased glycogen deposits by PAS and significantly increased lipid deposition by Oil Red O and confirmed by increased cardiac triglycerides. Plasma fatty acids were not different suggesting cardiac tissue lipid accumulation with normal circulating fatty acids. There was global sympathetic denervation in LCHADD hearts compared to WT visualized by tyrosine hydroxylase immunostaining. RNA-seq results on atria and ventricles found differentially expressed genes by KEGG pathway analysis: increased gene expression of glycolytic and glutathione synthesis enzymes, decreased gene expression of TCA cycle enzymes, and decreased gene expression of Ca++ signaling and cardiac muscle contraction genes. RT-PCR confirmed these findings.
Conclusion:
Conclusions: We demonstrate evidence of hypertrophic cardiomyopathy with diffuse fibrosis, accumulation of lipids, and lower glycogen storage in the absence of obesity. Increased gene expression of glycolytic and glutathione enzymes suggests a shift from fatty acid oxidation toward glycolysis with chronic oxidative stress. Chronic energy deficiency and lipotoxicity likely influence decreased gene expression of Ca++ signaling and cardiac contraction proteins. Long QRS and QT intervals with global denervation predispose to repolarization abnormalities increasing susceptibility to arrhythmias and elevating risk of sudden cardiac arrest and death. Further studies will be conducted to determine if current treatment options like low-fat diet and triheptanoin can modify the arrhythmogenic cardiomyopathy found in LCHADD.
Introduction: People with long-chain fatty acid oxidation disorders (LC-FAODs), such as LCHADD (OMIM# 609016), frequently present dilated or hypertrophic cardiomyopathy and can suffer life-threatening arrhythmias. LCHADD is caused by a missense pathogenic variant, c.1528G>C, of the HADHA gene encoding the alpha subunit of the mitochondrial trifunctional protein. In a recent retrospective analysis, we reported an increased incidence of major cardiac involvement in adolescent and young adults with LCHADD where heart failure and life-threatening arrhythmias remain the leading cause of mortality. The etiology of cardiac arrhythmias among people with LC-FAODs is not fully elucidated; therefore, we examined the mechanisms of impaired cardiac function in an LCHADD mouse model.
Methods:
Methods: We previously determined that the LCHADD mice (Hadha c.1528G>C homozygotes) recapitulate human disease by oxidizing more glucose and less fat compared to wildtype (WT) mice and accumulated LCHADD-specific long-chain 3-hydroxy-acylcarnitines in plasma. LCHADD mice have lower ketones when fasted, exhaust earlier during treadmill exercise and have decreased visual performance and retinopathy. On echocardiography, LCHADD mice have early concentric remodeling progressing to hypertrophic and dilated cardiomyopathy with low ejection fraction. To test for arrhythmias, we measured heart rate (HR), blood pressure (BP) and single-lead ECG on 9-month-old LCHADD and WT mice (n= 4-5 per genotype and sex) under mild sedation and stimulation with a single dose of a beta-agonist (isoproterenol; 3 mg/kg).
Results:
Results: LCHADD mice have significantly increased frequency of atrial premature beats, premature ventricular contractions, atrial flutter, atrial fibrillation, and non-sustained ventricular tachycardia after isoproterenol injection compared to WT mice, according to the Lambeth Conventions Guidelines. While HR was similar at baseline and increased with isoproterenol, it did not differ between groups. However, all LCHADD mice returned to baseline faster than WT after isoproterenol injection suggesting an inefficient sympathetic response. Long QRS and long QT intervals were seen in LCHADD male and female mice. Heart weight to body weight ratio was significantly increased in LCHADD hearts. Frozen and fixed heart sections demonstrated significantly increased cardiomyocyte cross-sectional area by H&E, increased collagen deposition by Masson Trichrome, decreased glycogen deposits by PAS and significantly increased lipid deposition by Oil Red O and confirmed by increased cardiac triglycerides. Plasma fatty acids were not different suggesting cardiac tissue lipid accumulation with normal circulating fatty acids. There was global sympathetic denervation in LCHADD hearts compared to WT visualized by tyrosine hydroxylase immunostaining. RNA-seq results on atria and ventricles found differentially expressed genes by KEGG pathway analysis: increased gene expression of glycolytic and glutathione synthesis enzymes, decreased gene expression of TCA cycle enzymes, and decreased gene expression of Ca++ signaling and cardiac muscle contraction genes. RT-PCR confirmed these findings.
Conclusion:
Conclusions: We demonstrate evidence of hypertrophic cardiomyopathy with diffuse fibrosis, accumulation of lipids, and lower glycogen storage in the absence of obesity. Increased gene expression of glycolytic and glutathione enzymes suggests a shift from fatty acid oxidation toward glycolysis with chronic oxidative stress. Chronic energy deficiency and lipotoxicity likely influence decreased gene expression of Ca++ signaling and cardiac contraction proteins. Long QRS and QT intervals with global denervation predispose to repolarization abnormalities increasing susceptibility to arrhythmias and elevating risk of sudden cardiac arrest and death. Further studies will be conducted to determine if current treatment options like low-fat diet and triheptanoin can modify the arrhythmogenic cardiomyopathy found in LCHADD.