Very long-/ and long Chain-3-Hydroxy Acyl CoA Dehydrogenase Deficiency correlates with deregulation of the mitochondrial fusion/fission machinery.

Children diagnosed with Long-Chain-3-Hydroxy-Acyl-CoA-Dehydrogenase-Deficiency (LCHADD) or Very-Long-Chain-3-Hydroxy-Acyl-CoA-Dehydrogenase-Deficiency (VLCADD) frequently present with hypertrophic cardiomyopathy or muscle weakness which is caused by the accumulation of fatty acid metabolites due to inactivating mutations in the mitochondrial trifunctional protein. By analyzing mitochondrial morphology we uncovered that mutations within the HADHA or the ACADVL gene not only affect fatty acid oxidation, but also cause significant changes in the DNM1L/MFN2 ratio leading to the significant accumulation of truncated and punctate mitochondria in contrast to network-like mitochondrial morphology in controls. These striking morphological abnormalities correlate with changes in OXPHOS, an imbalance in ROS levels, reduced mitochondrial respiration, reduced growth rates and significantly increased glucose uptake per cell, suggesting that HADHA and ACADVL mutations shift cellular energy household into glycolysis. Experiments using the NOX2-specific inhibitor Phox-I2 suggest that NOX2 is activated by accumulating long-chain fatty acids and generates ROS, which in turn changes mitochondrial morphology and activity. We thereby provide novel insights into the cellular energy household of cells from LCHADD/VLCADD patients and demonstrate for the first time a connection between fatty acid metabolism, mitochondrial morphology and ROS in patients with these rare genetic disorders.

Mitochondrial trifunctional protein deficiency in human cultured fibroblasts: effects of bezafibrate.

Mitochondrial trifunctional protein (MTP) deficiency caused by HADHA or HADHB gene mutations exhibits substantial molecular, biochemical, and clinical heterogeneity and ranks among the more severe fatty acid oxidation (FAO) disorders, without pharmacological treatment. Since bezafibrate has been shown to potentially correct other FAO disorders in patient cells, we analyzed its effects in 26 MTP-deficient patient fibroblasts representing 16 genotypes. Overall, the patient cell lines exhibited variable, complex, biochemical profiles and pharmacological responses. HADHA-deficient fibroblasts showed markedly reduced alpha subunit protein levels together with decreased beta-subunit abundance, exhibited a -86 to -96% defect in LCHAD activity, and produced large amounts of C14 and C16 hydroxyacylcarnitines. In control fibroblasts, exposure to bezafibrate (400 µM for 48 h) increased the abundance of HADHA and HADHB mRNAs, immune-detectable alpha and beta subunit proteins, activities of LCHAD and LCKAT, and stimulated FAO capacities, clearly indicating that MTP is pharmacologically up-regulated by bezafibrate in human fibroblasts. In MTP-deficient patient fibroblasts, which were found markedly FAO-deficient, bezafibrate improved FAO capacities in six of 26 (23%) cases, including three cell lines heterozygous for the common c1528G > C mutation. Altogether, our results strongly suggest that, due to variable effects of HADHA and HADHB mutations on MTP abundance and residual activity, improvement of MTP deficiency in response to bezafibrate was achieved in a subset of responsive genotypes.

A comprehensive HADHA c.1528G>C frequency study reveals high prevalence of long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency in Poland.

Isolated long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD) is associated with c.1528G>C substitution in the HADHA gene, since most patients have the prevalent mutation on at least one allele. As it is known that the disease is relatively frequent in Europe, especially around the Baltic Sea, and that the majority of Polish LCHADD patients originate from the coastal Pomeranian province, partly inhabited by an ancient ethnic group, the Kashubians, we aimed to determine the carrier frequency of the prevalent HADHA mutation in various districts of Poland with special focus on the Kashubian district. A total of 6,854 neonatal dried blood samples from the entire country, including 2,976 Pomeranian neonates of Kashubian origin, were c.1528G>C genotyped. Fifty-nine heterozygous carriers for the prevalent c.1528G>C substitution (41 Pomeranian children) were detected in the studied group. Our data reveal a geographically skewed distribution of the c.1528C allele in the Polish population; in the northern Pomeranian province the carrier frequency is 1:73, which is the highest frequency ever reported, whereas in the remaining regions it is 1:217. Hence, the incidence of LCHADD in Poland is predicted to be 1:118,336 versus 1:16,900 in the Pomeranian district. Despite the relative rarity of the disease, screening for LCHADD in neonates born in the northern part of Poland, especially those of Kashubian origin, is justified. Our data allow us to suggest a probable Kashubian origin of the prevalent c.1528G>C mutation.