MPC2 variants disrupt mitochondrial pyruvate metabolism and cause an early-onset mitochondriopathy.

Pyruvate is an essential metabolite produced by glycolysis in the cytosol and must be transported across the inner mitochondrial membrane into the mitochondrial matrix, where it is oxidized to fuel mitochondrial respiration. Pyruvate import is performed by the mitochondrial pyruvate carrier (MPC), a hetero-oligomeric complex composed by interdependent subunits MPC1 and MPC2. Pathogenic variants in the MPC1 gene disrupt mitochondrial pyruvate uptake and oxidation and cause autosomal-recessive early-onset neurological dysfunction in humans. The present work describes the first pathogenic variants in MPC2 associated with human disease in four patients from two unrelated families. In the first family, patients presented with antenatal developmental abnormalities and harboured a homozygous c.148T>C (p.Trp50Arg) variant. In the second family, patients that presented with infantile encephalopathy carried a missense c.2T>G (p.Met1?) variant disrupting the initiation codon. Patient-derived skin fibroblasts exhibit decreased pyruvate-driven oxygen consumption rates with normal activities of the pyruvate dehydrogenase complex and mitochondrial respiratory chain and no defects in mitochondrial content or morphology. Re-expression of wild-type MPC2 restored pyruvate-dependent respiration rates in patient-derived fibroblasts. The discovery of pathogenic variants in MPC2 therefore broadens the clinical and genetic landscape associated with inborn errors in pyruvate metabolism.

Adenosine kinase deficiency: Three new cases and diagnostic value of hypermethioninemia.

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.

Association of the CAG repeat polymorphism in mitochondrial polymerase gamma (POLG1) with male infertility: a case-control study in an Algerian population.

Polymorphisms in the have been speculated to be associated with male infertility. The main objective of our study was to CAG repeat polymorphism in POLG1 gene and male mitochondrial DNA polymerase gamma (POLG) assess the possible association of infertility in Algerian population. Genomic DNA from 89 infertile men and 84 controls was extracted using salting-out method. CAG repeat polymorphism was analyzed by the automated direct sequencing protocol. Statistical analysis was performed by Epi-info(r) (v6.0) software. A significant association with male infertility was found for CAG repeat polymorphism in heterozygous genotypes (10/≠10 vs 10/10: OR = 2.00 [0.99 - 4.05], p=0.03; "infertile vs control groups"; 10/≠10 vs 10/10: OR = 3.75[1.20-11.96], p=0.01 "oligoasthenoteratospermic group"). ALso, the results showed a significant association between the mordib allele (≠10) and male infertility (2.07 [01.07 - 04.02], p=0.01). Our results showed that POLG1 CAG repeat polymorphism might be a risk factor for male infertility in Algerian population. Investigations with larger sample sizes and representative population-based cases and matched controls are needed to validate our results.

Clinical, neuroimaging and biochemical findings in patients and patient fibroblasts expressing ten novel GFM1 mutations.

Pathogenic GFM1 variants have been linked to neurological phenotypes with or without liver involvement, but only a few cases have been reported in the literature. Here, we report clinical, biochemical, and neuroimaging findings from nine unrelated children carrying GFM1 variants, 10 of which were not previously reported. All patients presented with neurological involvement-mainly axial hypotonia and dystonia during the neonatal period-with five diagnosed with West syndrome; two children had liver involvement with cytolysis episodes or hepatic failure. While two patients died in infancy, six exhibited a stable clinical course. Brain magnetic resonance imaging showed the involvement of basal ganglia, brainstem, and periventricular white matter. Mutant EFG1 and OXPHOS proteins were decreased in patient's fibroblasts consistent with impaired mitochondrial translation. Thus, we expand the genetic spectrum of GFM1-linked disease and provide detailed clinical profiles of the patients that will improve the diagnostic success for other patients carrying GFM1 mutations.

Mitochondrial dysfunction caused by novel ATAD3A mutations.

Mitochondrial respiratory chain integrity depends on a number of proteins encoded by nuclear and mitochondrial genomes. Mutations of such factors can result in isolated or combined respiratory chain deficits, some of which can induce abnormal morphology of the mitochondrial network or accumulation of intermediary metabolites. Consequently, affected patients are clinically heterogeneous, presenting with central nervous system, muscular, or neurodegenerative disorders. ATAD3A is a nuclear-encoded ATPase protein of the AAA+ family and has been localized to the inner mitochondrial membrane. Recently reported mutations or large deletions in the ATDA3A gene in patients have been shown to induce altered mitochondrial structure and function and abnormal cholesterol metabolism in a recessive or dominant manner. Here, we report two siblings presenting axonal sensory-motor neuropathy associated with neonatal cataract. Genetic analyses identified two novel mutations in ATAD3A; a point mutation and an intronic 15 bp deletion affecting splicing and leading to exon skipping. Biochemical analysis in patient cells and tissues showed abnormal function of the mitochondrial respiratory chain in muscle and abnormal mitochondrial cristae structure. These new cases underline the large spectrum of biochemical and clinical presentations of ATAD3A deficiency and the different modes of inheritance, making it an atypical mitochondrial disorder.

Bovine and murine models highlight novel roles for SLC25A46 in mitochondrial dynamics and metabolism, with implications for human and animal health.

Neuropathies are neurodegenerative diseases affecting humans and other mammals. Many genetic causes have been identified so far, including mutations of genes encoding proteins involved in mitochondrial dynamics. Recently, the "Turning calves syndrome", a novel sensorimotor polyneuropathy was described in the French Rouge-des-Prés cattle breed. In the present study, we determined that this hereditary disease resulted from a single nucleotide substitution in SLC25A46, a gene encoding a protein of the mitochondrial carrier family. This mutation caused an apparent damaging amino-acid substitution. To better understand the function of this protein, we knocked out the Slc25a46 gene in a mouse model. This alteration affected not only the nervous system but also altered general metabolism, resulting in premature mortality. Based on optic microscopy examination, electron microscopy and on biochemical, metabolic and proteomic analyses, we showed that the Slc25a46 disruption caused a fusion/fission imbalance and an abnormal mitochondrial architecture that disturbed mitochondrial metabolism. These data extended the range of phenotypes associated with Slc25a46 dysfunction. Moreover, this Slc25a46 knock-out mouse model should be useful to further elucidate the role of SLC25A46 in mitochondrial dynamics.

Steroids in Stroke with Special Reference to Progesterone.

Both sex and steroid hormones are important to consider in human ischemic stroke and its experimental models. Stroke initiates a cascade of changes that lead to neural cell death, but also activates endogenous protective processes that counter the deleterious consequences of ischemia. Steroids may be part of these cerebroprotective processes. One option to provide cerebroprotection is to reinforce these intrinsic protective mechanisms. In the current review, we first summarize studies describing sex differences and the influence of steroid hormones in stroke. We then present and discuss our recent results concerning differential changes in endogenous steroid levels in the brains of male and female mice and the importance of progesterone receptors (PR) during the early phase after stroke. In the third part, we give an overview of experimental studies, including ours, that provide evidence for the pleiotropic beneficial effects of progesterone and its promising cerebroprotective potential in stroke. We also highlight the key role of PR signaling as well as potential additional mechanisms by which progesterone may provide cerebroprotection.

Pattern multiplicity and fumarate hydratase (FH)/S-(2-succino)-cysteine (2SC) staining but not eosinophilic nucleoli with perinucleolar halos differentiate hereditary leiomyomatosis and renal cell carcinoma-associated renal cell carcinomas from kidney tumors without FH gene alteration.

Hereditary leiomyomatosis and renal cell carcinoma syndrome is characterized by an increased risk of agressive renal cell carcinoma, often of type 2 papillary histology, and is caused by FH germline mutations. A prominent eosinophilic macronucleolus with a perinucleolar clear halo is distinctive of hereditary leiomyomatosis and renal cell carcinoma syndrome-associated renal cell carcinoma according to the 2012 ISUP and 2016 WHO kidney tumor classification. From an immunohistochemistry perspective, tumors are often FH-negative and S-(2-succino)-cysteine (2SC) positive. We performed a pathology review of 24 renal tumors in 23 FH mutation carriers, and compared them to 12 type 2 papillary renal cell carcinomas from FH wild-type patients. Prominent eosinophilic nucleoli with perinucleolar halos were present in almost all FH-deficient renal cell carcinomas (23/24). Unexpectedly, they were also present in 58% of type 2 papillary renal cell carcinomas from wild-type patients. Renal cell carcinoma in mutation carriers displayed a complex architecture with multiple patterns, typically papillary, tubulopapillary, and tubulocystic, but also sarcomatoid and rhabdoid. Such pattern diversity was not seen in non-carriers. FH/2SC immunohistochemistry was informative as all hereditary leiomyomatosis and renal cell carcinoma-associated renal cell carcinomas were either FH- or 2SC+. For FH and 2SC immunohistochemistries taken separately, sensitivity of negative anti-FH immunohistochemistry was 87.5% and specificity was 100%. For positive anti-2SC immunohistochemistry, sensitivity, and specificity were 91.7% and 91.7%, respectively. All FH wild-type renal cell carcinoma were FH-positive, and all but one were 2SC-negative. In conclusion, multiplicity of architectural patterns, rhabdoid/sarcomatoid components and combined FH/2SC staining, but not prominent eosinophilic nucleoli with perinucleolar halos, differentiate hereditary leiomyomatosis and renal cell carcinoma-associated renal cell carcinoma from type 2 papillary renal cell carcinoma with efficient FH gene. Our findings are crucial in identifying who should be referred to Cancer Genetics clinics for genetic counseling and testing.

Long-term liver disease in methylmalonic and propionic acidemias.

BACKGROUND AND OBJECTIVES: Patients affected with methylmalonic acidemia (MMA) and propionic acidemia (PA) exhibit diverse long-term complications and poor outcome. Liver disease is not a reported complication. The aim of this study was to characterize and extensively evaluate long-term liver involvement in MMA and PA patients. PATIENTS AND METHODS: We first describe four patients who had severe liver involvement during the course of their disease. Histology showed fibrosis and/or cirrhosis in 3 patients. Such liver involvement led us to retrospectively collect liver (clinical, laboratory and ultrasound) data of MMA (N = 12) or PA patients (N = 16) from 2003 to 2016. RESULTS: Alpha-fetoprotein (αFP) levels were increased in 8/16 and 3/12 PA and MMA patients, respectively, and tended to increase with age. Moderate and recurrent increase of GGT was observed in 4/16 PA patients and 4/12 MMA patients. Abnormal liver ultrasound with either hepatomegaly and/or hyperechoic liver was observed in 7/9 PA patients and 3/9 MMA patients. CONCLUSIONS: These data demonstrate that approximately half of the patients affected by MMA or PA had signs of liver abnormalities. The increase of αFP with age suggests progressive toxicity, which might be due to the metabolites accumulated in PA and MMA. These metabolites (e.g., methylmalonic acid and propionic acid derivatives) have previously been reported to have mitochondrial toxicity; this toxicity is confirmed by the results of histological and biochemical mitochondrial analyses of the liver in two of our MMA patients. In contrast to the moderate clinical, laboratory or ultrasound expression, severe pathological expression was found for three of the 4 patients who underwent liver biopsy, ranging from fibrosis to cirrhosis. These results emphasize the need for detailed liver function evaluation in organic aciduria patients, including liver biopsy when liver disease is suspected. TAKE HOME MESSAGE: MMA and PA patients exhibit long-term liver abnormalities.

Diagnostic approach to neurotransmitter monoamine disorders: experience from clinical, biochemical, and genetic profiles.

BACKGROUND AND AIM: To improve the diagnostic work-up of patients with diverse neurological diseases, we have elaborated specific clinical and CSF neurotransmitter patterns. METHODS: Neurotransmitter determinations in CSF from 1200 patients revealed abnormal values in 228 (19%) cases. In 54/228 (24%) patients, a final diagnosis was identified. RESULTS: We have reported primary (30/54, 56%) and secondary (24/54, 44%) monoamine neurotransmitter disorders. For primary deficiencies, the most frequently mutated gene was DDC (n = 9), and the others included PAH with neuropsychiatric features (n = 4), PTS (n = 5), QDPR (n = 3), SR (n = 1), and TH (n = 1). We have also identified mutations in SLC6A3, FOXG1 (n = 1 of each), MTHFR (n = 3), FOLR1, and MTHFD (n = 1 of each), for dopamine transporter, neuronal development, and folate metabolism disorders, respectively. For secondary deficiencies, we have identified POLG (n = 3), ACSF3 (n = 1), NFU1, and SDHD (n = 1 of each), playing a role in mitochondrial function. Other mutated genes included: ADAR, RNASEH2B, RNASET2, SLC7A2-IT1 A/B lncRNA, and EXOSC3 involved in nuclear and cytoplasmic metabolism; RanBP2 and CASK implicated in post-traductional and scaffolding modifications; SLC6A19 regulating amino acid transport; MTM1, KCNQ2 (n = 2), and ATP1A3 playing a role in nerve cell electrophysiological state. Chromosome abnormalities, del(8)(p23)/dup(12) (p23) (n = 1), del(6)(q21) (n = 1), dup(17)(p13.3) (n = 1), and non-genetic etiologies (n = 3) were also identified. CONCLUSION: We have classified the final 54 diagnoses in 11 distinctive biochemical profiles and described them through 20 clinical features. To identify the specific molecular cause of abnormal NT profiles, (targeted) genomics might be used, to improve diagnosis and allow early treatment of complex and rare neurological genetic diseases.

UQCRC2 mutation in a patient with mitochondrial complex III deficiency causing recurrent liver failure, lactic acidosis and hypoglycemia.

An isolated mitochondrial complex III (CIII) defect constitutes a rare cause of mitochondrial disorder. Here we present the second case involving UQCRC2 gene, which encodes core protein 2, one of the 11 structural subunits of CIII. The patient has the same mutation (c.547C>T; p.Arg183Trp) as the first case and presented with neonatal lactic acidosis, hypoglycemia and severe episodes of liver failure. Our study expands the few reported cases of CIII deficiency of nuclear origin.

Mutation in the AGK gene in two siblings with unusual Sengers syndrome.

Sengers syndrome is a rare autosomal recessive metabolic disorder caused by lack of acylglycerol kinase due to mutations in the AGK gene. It is characterized by congenital cataract, hypertrophic cardiomyopathy, myopathy and lactic acidosis. Two clinical forms have been described: a severe neonatal form, and a more benign form displaying exercise intolerance. We describe two siblings with congenital cataract, cardiomyopathy, hypotonia, intellectual disability and lactic acidosis. Whole exome sequencing revealed a homozygous c.1035dup mutation in the two siblings, supporting a diagnosis of Sengers syndrome. Our patients presented an intermediate form with intellectual deficiency, an unusual feature in Sengers syndrome. This permitted a prenatal diagnosis for a following pregnancy.

Mutations in CYC1, encoding cytochrome c1 subunit of respiratory chain complex III, cause insulin-responsive hyperglycemia.

Many individuals with abnormalities of mitochondrial respiratory chain complex III remain genetically undefined. Here, we report mutations (c.288G>T [p.Trp96Cys] and c.643C>T [p.Leu215Phe]) in CYC1, encoding the cytochrome c1 subunit of complex III, in two unrelated children presenting with recurrent episodes of ketoacidosis and insulin-responsive hyperglycemia. Cytochrome c1, the heme-containing component of complex III, mediates the transfer of electrons from the Rieske iron-sulfur protein to cytochrome c. Cytochrome c1 is present at reduced levels in the skeletal muscle and skin fibroblasts of affected individuals. Moreover, studies on yeast mutants and affected individuals' fibroblasts have shown that exogenous expression of wild-type CYC1 rescues complex III activity, demonstrating the deleterious effect of each mutation on cytochrome c1 stability and complex III activity.

Prevalence of rare mitochondrial DNA mutations in mitochondrial disorders.

BACKGROUND: Mitochondrial DNA (mtDNA) diseases are rare disorders whose prevalence is estimated around 1 in 5000. Patients are usually tested only for deletions and for common mutations of mtDNA which account for 5-40% of cases, depending on the study. However, the prevalence of rare mtDNA mutations is not known. METHODS: We analysed the whole mtDNA in a cohort of 743 patients suspected of manifesting a mitochondrial disease, after excluding deletions and common mutations. Both heteroplasmic and homoplasmic variants were identified using two complementary strategies (Surveyor and MitoChip). Multiple correspondence analyses followed by hierarchical ascendant cluster process were used to explore relationships between clinical spectrum, age at onset and localisation of mutations. RESULTS: 7.4% of deleterious mutations and 22.4% of novel putative mutations were identified. Pathogenic heteroplasmic mutations were more frequent than homoplasmic mutations (4.6% vs 2.8%). Patients carrying deleterious mutations showed symptoms before 16 years of age in 67% of cases. Early onset disease (<1 year) was significantly associated with mutations in protein coding genes (mainly in complex I) while late onset disorders (>16 years) were associated with mutations in tRNA genes. MTND5 and MTND6 genes were identified as 'hotspots' of mutations, with Leigh syndrome accounting for the large majority of associated phenotypes. CONCLUSIONS: Rare mitochondrial DNA mutations probably account for more than 7.4% of patients with respiratory chain deficiency. This study shows that a comprehensive analysis of mtDNA is essential, and should include young children, for an accurate diagnosis that is now accessible with the development of next generation sequencing technology.

Mitochondrial complex I deficiency of nuclear origin I. Structural genes.

Complex I (or NADH-ubiquinone oxidoreductase), is by far the largest respiratory chain complex with 38 subunits nuclearly encoded and 7 subunits encoded by the mitochondrial genome. Its deficiency is the most frequently encountered in mitochondrial disorders. Here, we summarize recent data obtained on architecture of complex I, and review the pathogenic mutations identified to date in nuclear structural complex I genes. The structural NDUFS1, NDUFS2, NDUFV1, and NDUFS4 genes are mutational hot spot genes for isolated complex I deficiency. The majority of the pathogenic mutations are private and the genotype-phenotype correlation is inconsistent in the rare recurrent mutations.

Mitochondrial complex I deficiency of nuclear origin II. Non-structural genes.

Complex I deficiency is the most frequent cause of respiratory chain diseases. This large multiprotein complex is composed in human of 45 structural subunits, of which 7 are mitochondrial-encoded and 38 are nuclear-encoded. Most of the pathological mutations responsible for complex I deficiencies have been identified to date in complex I structural subunits. Numerous studies from last decade gave some insight into the biogenesis of this huge multi subunit complex of double genetic origin. A sequential incorporation of the structural subunits as well as ten complex I assembly factors has been described. Here, we present a short overview of the human complex I biogenesis and we review the pathological mutations identified to date in eight of the ten known complex I assembly factors.

Novel FH mutations in families with hereditary leiomyomatosis and renal cell cancer (HLRCC) and patients with isolated type 2 papillary renal cell carcinoma.

BACKGROUND: Hereditary leiomyomatosis and renal cell cancer (HLRCC) is an autosomal dominant disorder predisposing humans to cutaneous and uterine leiomyomas; in 20% of affected families, type 2 papillary renal cell cancers (PRCCII) also occur with aggressive course and poor prognosis. HLRCC results from heterozygous germline mutations in the tumour suppressor fumarate hydratase (FH) gene. METHODS: As part of the French National Cancer Institute (INCa) 'Inherited predispositions to kidney cancer' network, sequence analysis and a functional study of FH were preformed in 56 families with clinically proven or suspected HLRCC and in 23 patients with isolated PRCCII (5 familial and 18 sporadic). RESULTS: The study identified 32 different germline FH mutations (15 missense, 6 frameshifts, 4 nonsense, 1 deletion/insertion, 5 splice site, and 1 complete deletion) in 40/56 (71.4%) families with proven or suspected HLRCC and in 4/23 (17.4%) probands with PRCCII alone, including 2 sporadic cases. 21 of these were novel and all were demonstrated as deleterious by significant reduction of FH enzymatic activity. In addition, 5 asymptomatic parents in 3 families were confirmed as carrying disease-causing mutations. CONCLUSIONS: This study identified and characterised 21 novel FH mutations and demonstrated that PRCCII can be the only one manifestation of HLRCC. Due to the incomplete penetrance of HLRCC, the authors propose to extend the FH mutation analysis to every patient with PRCCII occurring before 40 years of age or when renal tumour harbours characteristic histologic features, in order to discover previously ignored HLRCC affected families.

Clinical and biochemical heterogeneity associated with fumarase deficiency.

Fumarase deficiency (FD), caused by biallelic alteration of the Fumarase Hydratase gene (FH), and a rare metabolic disorder that affects the Krebs cycle, causes severe neurological impairment and fumaric aciduria. Less than 30 unrelated cases are known to date. In addition, heterozygous mutations of the FH gene are responsible for hereditary leiomyomatosis and renal cell cancer (HLRCC). We report three additional patients with dramatically different clinical presentations of FD and novel missense mutations in the FH gene. One patient had severe neonatal encephalopathy, polymicrogyria, <1% enzyme activity, and mildly increased levels of urinary fumarate. The second patient had microcephaly, mental retardation, 20% of fumarase activity, and intermediate levels of urinary fumarate. The third patient had mild mental retardation, polymicrogyria, 42-61% enzyme activity in different cell types and massive amounts of urinary fumarate. In silico analysis predicted minor yet significant structural changes in the encoded proteins. The nuclear translocation of hypoxia-inducible factor (HIF)-1alpha (HIF1A) in cultured fibroblasts was similar to controls. These results extend the range of clinical and biochemical variation associated with FD, supporting the notion that patients with moderate increases in fumarate excretion should be investigated for this disease. The tumoral risk in the patients and their relatives requires adequate screening protocols.

Carboplatin: a new cause of severe type B lactic acidosis secondary to mitochondrial DNA damage.

In adults, type B lactic acidosis is rare and generally associated with a toxin, particularly metformin or antiretroviral nucleosides analogues. We report a case of lactic acidosis caused by carboplatin in a 50-year-old woman suffering from primary peritoneal carcinoma. She was admitted for severe lactic acidosis (pH 6.77, lactate 19 mmol/L) associated with multiple organ failure (PaO2/FiO2 96, creatinine 231 µmol/L, aspartate aminotransferase > 25,000 UI, factor V 13%) occurring during the sixth carboplatin cycle. In the absence of sepsis, internal bleeding, alcohol poisoning, or other causes of lactic acidosis, the hypothesis of mitochondrial DNA (mtDNA) damage secondary to carboplatin and subsequent mitochondrial dysfunction leading to increase in glycolysis and lactic acidosis was suspected. L-Carnitine therapy associated with aggressive intensive care support led to a progressive improvement (pH 7.29, bicarbonate 24 mmol/L, lactate 7.8 mmol/L), but life support was withdrawn on day 7 because of peritoneal relapse. A respiratory chain dysfunction of enzyme activities encoded by mtDNA and multiple mtDNA deletions were found in muscle and liver tissue. It is generally accepted that carboplatin toxicity results in bone marrow suppression, renal dysfunction, or neurotoxicity and that platinating agents have no direct mitochondrial effect. However, although very unusual, emergency physicians must be aware that carboplatin can cause mitochondrial toxicity and trigger lactic acidosis.