Mitochondrial PCK2 Missense Variant in Shetland Sheepdogs with Paroxysmal Exercise-Induced Dyskinesia (PED).

Four female Shetland Sheepdogs with hypertonic paroxysmal dyskinesia, mainly triggered by exercise and stress, were investigated in a retrospective multi-center investigation aiming to characterize the clinical phenotype and its underlying molecular etiology. Three dogs were closely related and their pedigree suggested autosomal dominant inheritance. Laboratory diagnostic findings included mild lactic acidosis and lactaturia, mild intermittent serum creatine kinase (CK) elevation and hypoglycemia. Electrophysiological tests and magnetic resonance imaging of the brain were unremarkable. A muscle/nerve biopsy revealed a mild type II fiber predominant muscle atrophy. While treatment with phenobarbital, diazepam or levetiracetam did not alter the clinical course, treatment with a gluten-free, home-made fresh meat diet in three dogs or a tryptophan-rich, gluten-free, seafood-based diet, stress-reduction, and acetazolamide or zonisamide in the fourth dog correlated with a partial reduction in, or even a complete absence of, dystonic episodes. The genomes of two cases were sequenced and compared to 654 control genomes. The analysis revealed a case-specific missense variant, c.1658G>A or p.Arg553Gln, in the PCK2 gene encoding the mitochondrial phosphoenolpyruvate carboxykinase 2. Sanger sequencing confirmed that all four cases carried the mutant allele in a heterozygous state. The mutant allele was not found in 117 Shetland Sheepdog controls and more than 500 additionally genotyped dogs from various other breeds. The p.Arg553Gln substitution affects a highly conserved residue in close proximity to the GTP-binding site of PCK2. Taken together, we describe a new form of paroxysmal exercise-induced dyskinesia (PED) in dogs. The genetic findings suggest that PCK2:p.Arg553Gln should be further investigated as putative candidate causal variant.

A Nonsense Variant in the ACADVL Gene in German Hunting Terriers with Exercise Induced Metabolic Myopathy.

Several enzymes are involved in fatty acid oxidation, which is a key process in mitochondrial energy production. Inherited defects affecting any step of fatty acid oxidation can result in clinical disease. We present here an extended family of German Hunting Terriers with 10 dogs affected by clinical signs of exercise induced weakness, muscle pain, and suspected rhabdomyolysis. The combination of clinical signs, muscle histopathology and acylcarnitine analysis with an elevated tetradecenoylcarnitine (C14:1) peak suggested a possible diagnosis of acyl-CoA dehydrogenase very long chain deficiency (ACADVLD). Whole genome sequence analysis of one affected dog and 191 controls revealed a nonsense variant in the ACADVL gene encoding acyl-CoA dehydrogenase very long chain, c.1728C>A or p.(Tyr576*). The variant showed perfect association with the phenotype in the 10 affected and more than 500 control dogs of various breeds. Pathogenic variants in the ACADVL gene have been reported in humans with similar myopathic phenotypes. We therefore considered the detected variant to be the most likely candidate causative variant for the observed exercise induced myopathy. To our knowledge, this is the first description of this disease in dogs, which we propose to name exercise induced metabolic myopathy (EIMM), and the identification of the first canine pathogenic ACADVL variant. Our findings provide a large animal model for a known human disease and will enable genetic testing to avoid the unintentional breeding of affected offspring.

Clinical pattern, mutations and in vitro residual activity in 33 patients with severe 5, 10 methylenetetrahydrofolate reductase (MTHFR) deficiency.

BACKGROUND: Severe methylenetetrahydrofolate reductase (MTHFR) deficiency is a rare inborn defect disturbing the remethylation of homocysteine to methionine (<200 reported cases). This retrospective study evaluates clinical, biochemical genetic and in vitro enzymatic data in a cohort of 33 patients. METHODS: Clinical, biochemical and treatment data was obtained from physicians by using a questionnaire. MTHFR activity was measured in primary fibroblasts; genomic DNA was extracted from cultured fibroblasts. RESULTS: Thirty-three patients (mean age at follow-up 11.4 years; four deceased; median age at first presentation 5 weeks; 17 females) were included. Patients with very low (<1.5%) mean control values of enzyme activity (n = 14) presented earlier and with a pattern of feeding problems, encephalopathy, muscular hypotonia, neurocognitive impairment, apnoea, hydrocephalus, microcephaly and epilepsy. Patients with higher (>1.7-34.8%) residual enzyme activity had mainly psychiatric symptoms, mental retardation, myelopathy, ataxia and spasticity. Treatment with various combinations of betaine, methionine, folate and cobalamin improved the biochemical and clinical phenotype. During the disease course, patients with very low enzyme activity showed a progression of feeding problems, neurological symptoms, mental retardation, and psychiatric disease while in patients with higher residual enzyme activity, myelopathy, ataxia and spasticity increased. All other symptoms remained stable or improved in both groups upon treatment as did brain imaging in some cases. No clear genotype-phenotype correlation was obvious. DISCUSSION: MTHFR deficiency is a severe disease primarily affecting the central nervous system. Age at presentation and clinical pattern are correlated with residual enzyme activity. Treatment alleviates biochemical abnormalities and clinical symptoms partially.

An inherited episodic head tremor syndrome in Doberman pinscher dogs.

Episodic head tremor anecdotally occurs in the Doberman pinscher dog breed, but it is not described in sufficient detail in the literature. We evaluated 87 Doberman pinschers affected with episodic head tremor and appropriate controls. The data analyzed were collected through detailed questionnaires, elaborate telephone interviews, and video recordings. Affected dogs underwent clinical, neurological, and laboratory examination, and a detailed diagnostic workup was conducted in 5 affected dogs. Pedigrees of affected dogs were collected and reviewed. The affected dogs expressed individual phenotypes of either horizontal or vertical head movements, but rarely did a dog exhibit head movements in both directions. There was considerable variation in duration (10 seconds to 3 hours; median: 3 minutes), frequency of occurrence (1-20 episodes/day; median: 2/day) of head tremor and length of the period without head tremor (1-1,800 days; median: 60 days). Subtle dystonic posturing of the head and neck during head tremor was evident on video recordings of 5 dogs. Certain exceptional conditions such as illness, surgery, some medications, heat, pseudopregnancy, or pregnancy triggered episodes. Two main important forms of episodic head tremor were identified: a familial early-onset form (age < 1 year) that affected littermates and a sporadic form. Affected dogs were traced back to 1 common sire, also including sporadic cases. Episodic head tremor is an inherited, paroxysmal movement disorder that affects the Doberman pinscher breed. Identification of the causative genes in the future will allow us to obtain a more detailed description of the syndrome.

Multidimensional gas chromatography-mass spectrometry for tracer studies of fatty acid metabolism via stable isotopes in cultured human trophoblast cells.

The determination of placental fatty acid metabolism using stable isotope-labeled tracers was investigated in the human placental choriocarcinoma (JAR) cell line. Stable isotope incorporation was measured by MDGC-MS. The cultured trophoblast cells incorporated and metabolized the essential fatty acids to long-chain polyunsaturated fatty acids. The described method enables the detection of a low Delta(6)-desaturase activity in this human placental cell line. The developed MDGC-MS method allows the assessment of long-chain polyunsaturated fatty acid biosynthesis in cultured cells with high sensitivity and selectivity. In this respect, tracer studies with MDGC-MS will be a powerful tool to clarify the significance of placental fatty acid metabolism.

Antibiotic treatment-induced tubular dysfunction as a risk factor for renal stone formation in cystic fibrosis.

OBJECTIVES: Our purpose was to characterize the decisive pathophysiologic factors that lead to renal stone formation (nephrolithiasis) in patients with cystic fibrosis (CF). METHODS: Patients with CF (n = 96) were investigated with respect to lithogenic and inhibitory factors of urolithiasis and compared with 30 healthy control patients. They were subdivided into 2 groups, 86 without renal stones and 10 with renal stones. RESULTS: All stones were exclusively composed of calcium oxalate. As a major pathogenic factor, a urinary disequilibrium between promoting and inhibitory components of stone formation, characterized mainly by hypercalciuria, hyperoxaluria, and hypocitraturia, was found in the patients with nephrolithiasis. They tended to have lower plasma phosphate concentrations and an increased urinary phosphate excretion. The citrate/calcium ratio proved to be a valuable means to discriminate patients with renal stones from control patients. Patients with stones had ingested more cotrimoxazole and ceftazidim, cumulatively, than patients without stones. There was an inverse correlation between the amounts of antibiotics ingested and the percentage of tubular phosphate reabsorption (r = -0.91, P <.0046). CONCLUSION: Renal stone formation in patients with CF is caused by a disequilibrium between promoting and inhibitory components of stone formation, which is dominated by hypercalciuria, hyperoxaluria, and hypocitraturia. Treatment with cotrimoxazole and ceftazidim, primarily, may lead to renal proximal tubular damage with an ensuing sequence of phosphate loss, increase of parathyroid hormone secretion, increased 1,25-dihydroxyvitamin D3 formation, and absorptive hypercalciuria.