Thermo-sensitive mitochondrial trifunctional protein deficiency presenting with episodic myopathy.

Mitochondrial trifunctional protein (MTP) is involved in long-chain fatty acid ß-oxidation (lcFAO). Deficiency of one or more of the enzyme activities as catalyzed by MTP causes generalized MTP deficiency (MTPD), long-chain hydroxyacyl-CoA dehydrogenase deficiency (LCHADD), or long-chain ketoacyl-CoA thiolase deficiency (LCKATD). When genetic variants result in thermo-sensitive enzymes, increased body temperature (e.g. fever) can reduce enzyme activity and be a risk factor for clinical decompensation. This is the first description of five patients with a thermo-sensitive MTP deficiency. Clinical and genetic information was obtained from clinical files. Measurement of LCHAD and LCKAT activities, lcFAO-flux studies and palmitate loading tests were performed in skin fibroblasts cultured at 37°C and 40°C. In all patients (four MTPD, one LCKATD), disease manifested during childhood (manifestation age: 2-10 years) with myopathic symptoms triggered by fever or exercise. In four patients, signs of retinopathy or neuropathy were present. Plasma long-chain acylcarnitines were normal or slightly increased. HADHB variants were identified (at age: 6-18 years) by whole exome sequencing or gene panel analyses. At 37°C, LCHAD and LCKAT activities were mildly impaired and lcFAO-fluxes were normal. Remarkably, enzyme activities and lcFAO-fluxes were markedly diminished at 40°C. Preventive (dietary) measures improved symptoms for most. In conclusion, all patients with thermo-sensitive MTP deficiency had a long diagnostic trajectory and both genetic and enzymatic testing were required for diagnosis. The frequent absence of characteristic acylcarnitine abnormalities poses a risk for a diagnostic delay. Given the positive treatment effects, upfront genetic screening may be beneficial to enhance early recognition.

The spectrum of peripheral neuropathy in disorders of the mitochondrial trifunctional protein.

Peripheral neuropathy is a known irreversible long-term complication of long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD) and mitochondrial trifunctional protein deficiency (MTPD), two inherited disorders of mitochondrial long-chain fatty acid oxidation. The underlying pathophysiology of neuropathy is still not fully understood. We report electrophysiological studies and neurological findings in a series of 8 LCHAD-deficient and 11 MTP-deficient patients. The median age at time of the study was 8.0 years (0.5-25 years). The overall prevalence of neuropathy was 58% with neuropathic symptoms being slightly more common in MTPD compared to LCHADD (70% vs 50%, respectively). Onset of neuropathy was significantly earlier in MTPD patients compared to LCHADD patients (median age at onset 4.7 vs 15.3 years, respectively, P = .047). In four patients, isolated peripheral neuropathy was the first and only presenting symptom, and in all four the diagnosis was missed by newborn screening. About half of the patients (45.5%) had a sensorimotor neuropathy, while 27.3% showed a pure motor form and another 27.3% an isolated sensory form. Despite early diagnosis by newborn screening and early initiation of therapy, peripheral neuropathy cannot be prevented in all patients with LCHADD/MTPD and has severe impact on the life of affected patients. Electrophysiology classifies LCHADD/MTPD neuropathy as axonal with secondary demyelination. A novel observation is that in patients with acute, fulminant onset of neuropathy, symptoms can be partly reversible. Further studies are needed to elucidate the underlying pathophysiology of axonal damage and possible therapeutic targets.

Pharmacological inhibition of carnitine palmitoyltransferase 1 restores mitochondrial oxidative phosphorylation in human trifunctional protein deficient fibroblasts.

BACKGROUND: Mitochondrial Trifunctional Protein deficiency (TFPD) is a severe genetic disease characterized by altered energy metabolism and accumulation of long-chain (LC) acylcarnitines in blood and tissues. This accumulation could impair the mitochondrial oxidative phosphorylation (OxPhos), contributing to the non-optimal outcome despite conventional diet therapy with medium-chain triglycerides (MCT). METHOD: Acylcarnitine and OxPhos parameters were measured in TFPD-fibroblasts obtained from 8 children and cultured in medium mimicking fasting (LCFA) or conventional treatment (MCT), with or without Etomoxir (ETX) an inhibitor of carnitine palmitoyltransferase 1 (CPT1) activity, and were compared to results obtained with fibroblasts from 5 healthy-control children. The effects of various acylcarnitines were also tested on control fibroblasts. RESULTS: In the LCFA-condition, TFPD-fibroblasts demonstrated a large accumulation of LC-acylcarnitines associated with decreased O2-consumption (63±3% of control, P<0.001) and ATP production (67±5%, P<0.001) without modification of coupling efficiency. A dose-dependent decrease in O2-consumption was reproduced in control fibroblasts by addition of increasing dose of LC-acylcarnitines, while it was almost preserved with MC-acylcarnitines. The MCT-condition reduced LC-acylcarnitine accumulation and partially improved O2-consumption (80±3%, P<0.01) in TFPD-fibroblasts. The addition of ETX in both LCFA- and MCT-conditions normalized acylcarnitine profiles and restored O2-consumption and ATP production at the same levels than control. CONCLUSION: Accumulation of LC-acylcarnitines plays a major role in the pathophysiology of TFPD, reducing OxPhos capacities. These deleterious effects could be partially prevented by MCT-therapy and totally corrected by ETX. Inhibition of CPT1 may be view as a new therapeutic target for patients with a severe form of TFPD.

Mitochondrial bioenergetics deregulation caused by long-chain 3-hydroxy fatty acids accumulating in LCHAD and MTP deficiencies in rat brain: a possible role of mPTP opening as a pathomechanism in these disorders?

Long-chain 3-hydroxylated fatty acids (LCHFA) accumulate in long-chain 3-hydroxy-acyl-CoA dehydrogenase (LCHAD) and mitochondrial trifunctional protein (MTP) deficiencies. Affected patients usually present severe neonatal symptoms involving cardiac and hepatic functions, although long-term neurological abnormalities are also commonly observed. Since the underlying mechanisms of brain damage are practically unknown and have not been properly investigated, we studied the effects of LCHFA on important parameters of mitochondrial homeostasis in isolated mitochondria from cerebral cortex of developing rats. 3-Hydroxytetradecanoic acid (3 HTA) reduced mitochondrial membrane potential, NAD(P)H levels, Ca(2+) retention capacity and ATP content, besides inducing swelling, cytochrome c release and H2O2 production in Ca(2+)-loaded mitochondrial preparations. We also found that cyclosporine A plus ADP, as well as ruthenium red, a Ca(2+) uptake blocker, prevented these effects, suggesting the involvement of the mitochondrial permeability transition pore (mPTP) and an important role for Ca(2+), respectively. 3-Hydroxydodecanoic and 3-hydroxypalmitic acids, that also accumulate in LCHAD and MTP deficiencies, similarly induced mitochondrial swelling and decreased ATP content, but to a variable degree pending on the size of their carbon chain. It is proposed that mPTP opening induced by LCHFA disrupts brain bioenergetics and may contribute at least partly to explain the neurologic dysfunction observed in patients affected by LCHAD and MTP deficiencies.

Acute fatty liver of pregnancy associated with fetal mitochondrial trifunctional protein deficiency.

Acute fatty liver of pregnancy (AFLP) is a devastating disorder of the maternal liver in the third trimester. Recent studies have demonstrated an association between AFLP and fetal fatty acid oxidation disorders. Here, we report a case of AFLP caused by fetal mitochondrial trifunctional protein (TFP) deficiency. A 21-year-old parous woman presented with nausea, genital bleeding and abdominal pain at 33 weeks of gestation. Laboratory data revealed hepatic failure and disseminated intravascular coagulopathy. The patient underwent emergency cesarean section and was diagnosed with AFLP from the clinical characteristics. She was successfully treated with frequent plasma exchange. The newborn presented severe heart failure and died on the 39th day after birth. Tandem mass spectrometry indicated long-chain fatty acid oxidation disorder. Gene analysis demonstrated homozygous mutation in exon 13 of HADHB, the gene responsible for mitochondrial TFP deficiency. The parents carried a heterozygous mutation at the same location in HADHB.

Mitochondrial trifunctional protein deficiency: a rare cause of adult-onset rhabdomyolysis.

INTRODUCTION: Mitochondrial trifunctional protein deficiency is a rare autosomal recessive disorder of mitochondrial fatty acid ß-oxidation that may be due to mutations in 2 different nuclear genes, HADHA and HADHB. Perturbation of this multienzyme complex compromises the oxidation of long-chain fatty acids, which leads to multiorgan dysfunction. Childhood- or adolescent-onset recurrent rhabdomyolysis is a common muscular manifestation and is preceded frequently by clinically overt peripheral neuropathy. METHODS: In this report we describe a patient with late adult-onset recurrent rhabdomyolysis. RESULTS: Despite normal sensory examination, nerve conduction studies showed a mild axonal peripheral neuropathy. The acylcarnitine profile showed elevated long-chain and 3-hydroxy long-chain acylcarnitine species. HADHA sequencing revealed known compound heterozygous mutations c.180+3A>G (p.Thr37SerfsX6) and c.1528G>C (p.Glu510Gln). During a 10-month follow-up period, he had no further episodes of rhabdomyolysis after appropriate dietary modifications. CONCLUSIONS: Mitochondrial trifunctional protein deficiency should be considered in patients with adult-onset recurrent rhabdomyolysis, especially in those with either clinically overt or subclinical peripheral neuropathy.

Reversible sensory neuropathy in mitochondrial trifunctional protein deficiency.

Axonal peripheral neuropathy is a common complication of mitochondrial trifunctional protein (MTP) deficiency and long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency that is usually considered progressive. Current treatment strategies are not able to fully prevent neuropathic symptoms in the majority of patients. We herein report three sisters with genetically proven MTP deficiency who were untreated until adolescence, when electrophysiological studies first revealed isolated axonal sensory neuropathy. Apart from mild exercise intolerance and missing deep tendon reflexes of the lower extremities, all three girls were clinically asymptomatic. A fat-reduced and fat-modified diet together with a reduction of the nocturnal fasting time resulted in complete normalisation of the electrophysiological studies after 1 year of dietary treatment. Our findings suggest that neuropathy might be responsive to dietary interventions in MTP patients at a very early stage of disease.

Hypoparathyroidism, neutropenia and nephrotic syndrome in a patient with mitochondrial trifunctional protein deficiency: A case report and review of the literature.

INTRODUCTION: Mitochondrial trifunctional protein (TFP) deficiency is an autosomal recessive disorder that causes a clinical spectrum of diseases ranging from severe infantile cardiomyopathy to mild chronic progressive neuromyopathy, however, parathyroid glands, hematologic system and kidney damage are not the common presentations of this disease. METHODS: We describe the clinical, biochemical and molecular features of the TFP deficiency patient at our institution. We also provide an extensive literature review of previous published cases with emphasis on the clinical/biochemical phenotype-genotype correlation of this disorder. RESULTS: Our case is a complete TFP deficiency patient dominated presented with hypoparathyroidism, neutropenia and nephrotic syndrome, which caused by compound heterozygoues variants in HADHB gene. Based on the retrospective study of 157 cases, TFP patients presented with diverse clinical, biochemical and molecular features. The onset age is typically before early childhood. Neuromuscular system is more vulnerable involved. Severe form is generally characterized by multiorgan involvement. A notable feature of severe and intermediate form is respiratory failure. Neuropathy and rhabdomyolysis are the typical manifestations of mild form. Increased long-chain 3-OH-acylcarnitines (C16-OH, C18:1-OH) are the most common biochemical finding. The mortality of the present study is as high as 57.9%, which is linked with the onset age, phenotype, mutation type and muscular histology. Mutations in HADHB are more frequent in Asian descent with complete TFP deficiency and usually presented with atypical presentations. The type of mutation, rather than residual enzyme activity seem to be more related to the phenotype and prognosis. The most common HADHA variant is 1528G > C, no common HADHB variant were detected. CONCLUSIONS: TFP deficiency is heterogeneous at both the molecular and phenotypic levels, generally a high mortality. Although there is no strict clinical/biochemical phenotype-genotype correlation, difference in ethnic and subunit mutations still have certain characteristics.

Mitochondrial trifunctional protein deficiency as a polyneuropathy etiology in childhood.

BACKGROUND: The mitochondrial trifunctional protein (MTP) is a multienzyme complex of the fatty acid betaoxidation cycle. Mitochondrial trifunctional protein deficiency (MTPD), a rare condition that leads to failure of converting certain fats to energy is characterized by decreased activity of three enzymes in the enzyme complex. Signs and symptoms of MTPD may present during infancy or later in life; those that begin after infancy include hypotonia, muscle pain, rhabdomyolysis, and peripheral neuropathy. We report a Turkish boy diagnosed with MTPD after being investigated for polyneuropathy of unknown origin since infancy. CASE: A 5.5-year-old male patient was admitted to our clinic with complaints of weakness in the arms and legs, physical inactivity compared to his peers, fatigue, weakness and, difficulty in climbing stairs since infancy. Electroneuromyography (ENMG) analysis showed moderate symmetric distal sensorimotor and axonal neuropathy. On the background of chronic polyneuropathy, the patient had acute relapsing episodes with progressively worsening severity in the follow-up period until 12.5 years of age. Whole exome sequencing (WES) was performed in the patient and, revealed that the patient had a homozygous c.1390G > A (p.Gly464Ser) pathogenic variant of the HADHB gene. Although rhabdomyolysis is a well defined accompanying clinical feature of MTPD, it was not present in our patient who only had worsening muscle weakness during attacks. CONCLUSION: On the background of chronic polyneuropathy and acute relapsing episodes triggered by fasting or illnesses and rhabdomyolysis physicians should suspect disorders of the fatty acid beta-oxidation cycle.

Charcot-Marie-Tooth Disease With Episodic Rhabdomyolysis Due to Two Novel Mutations in the ß Subunit of Mitochondrial Trifunctional Protein and Effective Response to Modified Diet Therapy.

A 29-year-old female experienced chronic progressive peripheral neuropathy since childhood and was diagnosed with Charcot-Marie-Tooth disease (CMT) at age 15. She developed recurrent, fever-induced rhabdomyolysis (RM) at age 24. EMG studies showed decreased amplitude of compound muscle action potential, declined motor conductive velocity, and absence of sensor nerve action potential. Acylcarnitine analysis revealed elevated C16-OH, C18-OH, and C18:1-OH. Muscle biopsy showed scattered foci of necrotic myofibers invaded by macrophages, occasional regenerating fibers, and remarkable muscle fiber type grouping. Whole-exome sequencing identified two novel heterozygous mutations: c.490G>A (p.G164S) and c.686G>A (p.R229Q) in HADHB gene encoding the ß-subunit of mitochondrial trifunctional protein (MTP). Reduction of long-chain fatty acid via dietary restrictions alleviated symptoms effectively. Our study indicates that the defect of the MTP ß-subunit accounts for both CMT and RM in the same patient and expands the clinical spectrum of disorders caused by the HADHB mutations. Our systematic review of all MTPD patients with dietary treatment indicates that the effect of dietary treatment is related to the age of onset and the severity of symptoms.

A Rare Presentation of Cardiomyopathy in Pregnancy.

Our patient presented in her third trimester of pregnancy with new onset of heart failure. A thorough workup in the initial postpartum period with detailed past medical history, advanced imaging modalities, and a multidisciplinary approach revealed a rare and treatable etiology of cardiomyopathy. (Level of Difficulty: Intermediate.).

Identification and functional characterization of mutations within HADHB associated with mitochondrial trifunctional protein deficiency.

Mitochondrial trifunctional protein (MTP) deficiency is a rare autosomal recessive disorder with several phenotypes. Neuromyopathic form of MTP deficiency is characterized by infantile or juvenile-onset, progressive peripheral neuropathy and rhabdomyolysis. To date, only one Chinese patient harboring homozygous c. 739C>T (p.R247C) in HADHB has been reported. Here, using whole exome sequencing (WES), we identified a compound heterozygote of c.407T>C (p.M136T) and c.421G>A (p.A141T) within HADHB in a Chinese MTP deficiency patient of neuromyopathic form. In vitro cell functional studies were performed to evaluate the effect of mutations on MTP complex expression and subcellular location, which revealed that p.M136T and p.A141T mutations compromised MTP complex stability but not altered subcellular localization, resulting in lower protein level at 37 °C but higher at 30 °C. These results indicated that both mutations were pathogenic through a loss-of-function mechanism and temperature-sensitive leading to their correlation with the mild phenotype. The current study broadens the genetic spectrum of HADHB and highlights the importance of screening fatty acid oxidation deficiency-related gene mutations among patients with intermittent rhabdomyolysis, as in the patient reported here, although extremely rare.

Evaluation of earlier versus later dietary management in long-chain 3-hydroxyacyl-CoA dehydrogenase or mitochondrial trifunctional protein deficiency: a systematic review.

BACKGROUND: Mitochondrial trifunctional protein (MTP) and long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiencies are rare fatty acid ß-oxidation disorders. Without dietary management the conditions are life-threatening. We conducted a systematic review to investigate whether pre-symptomatic dietary management following newborn screening provides better outcomes than treatment following symptomatic detection. METHODS: We searched Web of Science, Medline, Pre-Medline, Embase and the Cochrane Library up to 23rd April 2018. Two reviewers independently screened titles, abstracts and full texts for eligibility and quality appraised the studies. Data extraction was performed by one reviewer and checked by another. RESULTS: We included 13 articles out of 7483 unique records. The 13 articles reported on 11 patient groups, including 174 people with LCHAD deficiency, 18 people with MTP deficiency and 12 people with undifferentiated LCHAD/MTP deficiency. Study quality was moderate to weak in all studies. Included studies suggested fewer heart and liver problems in screen-detected patients, but inconsistent results for mortality. Follow up analyses compared long-term outcomes of (1) pre-symptomatically versus symptomatically treated patients, (2) screened versus unscreened patients, and (3) asymptomatic screen-detected, symptomatic screen-detected, and clinically diagnosed patients in each study. For follow up analyses 1 and 2, we found few statistically significant differences in the long-term outcomes. For follow up analysis 3 we found a significant difference for only one comparison, in the incidence of cardiomyopathy between the three groups. CONCLUSIONS: There is some evidence that dietary management following screen-detection might be associated with a lower incidence of some LCHAD and MTP deficiency-related complications. However, the evidence base is limited by small study sizes, quality issues and risk of confounding. An internationally collaborative research effort is needed to fully examine the risks and the benefits to pre-emptive dietary management with particular attention paid to disease severity and treatment group.

Disturbance of mitochondrial functions provoked by the major long-chain 3-hydroxylated fatty acids accumulating in MTP and LCHAD deficiencies in skeletal muscle.

The pathogenesis of the muscular symptoms and recurrent rhabdomyolysis that are commonly manifested in patients with mitochondrial trifunctional protein (MTP) and long-chain 3-hydroxy-acyl-CoA dehydrogenase (LCHAD) deficiencies is still unknown. In this study we investigated the effects of the major long-chain monocarboxylic 3-hydroxylated fatty acids (LCHFA) accumulating in these disorders, namely 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, on important mitochondrial functions in rat skeletal muscle mitochondria. 3HTA and 3HPA markedly increased resting (state 4) and decreased ADP-stimulated (state 3) and CCCP-stimulated (uncoupled) respiration. 3HPA provoked similar effects in permeabilized skeletal muscle fibers, validating the results obtained in purified mitochondria. Furthermore, 3HTA and 3HPA markedly diminished mitochondrial membrane potential, NAD(P)H content and Ca(2+) retention capacity in Ca(2+)-loaded mitochondria. Mitochondrial permeability transition (mPT) induction probably underlie these effects since they were totally prevented by cyclosporin A and ADP. In contrast, the dicarboxylic analogue of 3HTA did not alter the tested parameters. Our data strongly indicate that 3HTA and 3HPA behave as metabolic inhibitors, uncouplers of oxidative phosphorylation and mPT inducers in skeletal muscle. It is proposed that these pathomechanisms disrupting mitochondrial homeostasis may be involved in the muscle alterations characteristic of MTP and LCHAD deficiencies.