Analysis of human brain tissue derived from DBS surgery

Background: Transcriptomic and proteomic profiling of human brain tissue is hindered by the availability of fresh samples from living patients. Postmortem samples usually represent the advanced disease stage of the patient. Furthermore, the postmortem interval can affect the transcriptomic and proteomic profiles. Therefore, fresh brain tissue samples from living patients represent a valuable resource of metabolically intact tissue. Implantation of deep brain stimulation (DBS) electrodes into the human brain is a neurosurgical treatment for, e.g., movement disorders. Here, we describe an improved approach to collecting brain tissues from surgical instruments used in implantation of DBS device for transcriptomics and proteomics analyses. Methods: Samples were extracted from guide tubes and recording electrodes used in routine DBS implantation procedure to treat patients with Parkinson's disease, genetic dystonia and tremor. RNA sequencing was performed in tissues extracted from the recording microelectrodes and liquid chromatography-mass spectrometry (LC-MS) performed in tissues from guide tubes. To assess the performance of the current approach, the obtained datasets were compared with previously published datasets representing brain tissues. Results: Altogether, 32,034 RNA transcripts representing the unique Ensembl gene identifiers were detected from eight samples representing both hemispheres of four patients. By using LC-MS, we identified 734 unique proteins from 31 samples collected from 14 patients. The datasets are available in the BioStudies database (accession number S-BSST667). Our results indicate that surgical instruments used in DBS installation retain brain material sufficient for protein and gene expression studies. Comparison with previously published datasets obtained with similar approach proved the robustness and reproducibility of the protocol. Conclusions: The instruments used during routine DBS surgery are a useful source for obtaining fresh brain tissues from living patients. This approach overcomes the issues that arise from using postmortem tissues, such as the effect of postmortem interval on transcriptomic and proteomic landscape of the brain, and can be used for studying molecular aspects of DBS-treatable diseases.Peer reviewe

Cytosolic phosphoenolpyruvate carboxykinase deficiency : Expanding the clinical phenotype and novel laboratory findings

Cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C) deficiency due to the homozygous PCK1 variant has recently been associated with childhood-onset hypoglycemia with a recognizable pattern of abnormal urine organic acids. In this study, 21 children and 3 adult patients with genetically confirmed PEPCK-C deficiency were diagnosed during the years 2016 to 2019 and the available biochemical and clinical data were collected. All patients were ethnic Finns. Most patients (22 out of 24) had a previously published homozygous PCK1 variant c.925G>A. Two patients had a novel compound heterozygous PCK1 variant c.925G>A and c.716C>T. The laboratory results showed abnormal urine organic acid profile with increased tricarboxylic acid cycle intermediates and inadequate ketone body production during hypoglycemia. The hypoglycemic episodes manifested predominantly in the morning. Infections, fasting or poor food intake, heavy exercise, alcohol consumption, and breastfeeding were identified as triggering factors. Five patients presented with neonatal hypoglycemia. Hypoglycemic seizures occurred in half of the patients (12 out of 24). The first hypoglycemic episode often occurred at the age of 1-2 years, but it sometimes presented at a later age, and could re-occur during school age or adulthood. This study adds to the laboratory data on PEPCK-C deficiency, confirming the recognizable urine organic acid pattern and identifying deficient ketogenesis as a novel laboratory finding. The phenotype is expanded suggesting that the risk of hypoglycemia may continue into adulthood if predisposing factors are present.Peer reviewe

A multicenter study on Leigh syndrome: Disease course and predictors of survival

Background: Leigh syndrome is a progressive neurodegenerative disorder, associated with primary or secondary dysfunction of the mitochondrial oxidative phosphorylation. Despite the fact that Leigh syndrome is the most common phenotype of mitochondrial disorders in children, longitudinal natural history data is missing. This study was undertaken to assess the phenotypic and genotypic spectrum of patients with Leigh syndrome, characterise the clinical course and identify predictors of survival in a large cohort of patients. Methods. This is a retrospective study of patients with Leigh syndrome that have been followed at eight centers specialising in mitochondrial diseases in Europe; Gothenburg, Rotterdam, Helsinki, Copenhagen, Stockholm, Brussels, Bergen and Oulu. Results: A total of 130 patients were included (78 males; 52 females), of whom 77 patients had identified pathogenic mutations. The median age of disease onset was 7 months, w

HIDEA syndrome is caused by biallelic, pathogenic, rare or founder P4HTM variants impacting the active site or the overall stability of the P4H-TM protein

HIDEA syndrome is caused by biallelic pathogenic variants in P4HTM. The phenotype is characterized by muscular and central hypotonia, hypoventilation including obstructive and central sleep apneas, intellectual disability, dysautonomia, epilepsy, eye abnormalities, and an increased tendency to develop respiratory distress during pneumonia. Here, we report six new patients with HIDEA syndrome caused by five different biallelic P4HTM variants, including three novel variants. We describe two Finnish enriched pathogenic P4HTM variants and demonstrate that these variants are embedded within founder haplotypes. We review the clinical data from all previously published patients with HIDEA and characterize all reported P4HTM pathogenic variants associated with HIDEA in silico. All known pathogenic variants in P4HTM result in either premature stop codons, an intragenic deletion, or amino acid changes that impact the active site or the overall stability of P4H-TM protein. In all cases, normal P4H-TM enzyme function is expected to be lost or severely decreased. This report expands knowledge of the genotypic and phenotypic spectrum of the disease.publishedVersio

POLG1 p.R722H mutation associated with multiple mtDNA deletions and a neurological phenotype

<p>Abstract</p> <p>Background</p> <p>The c.2447G>A (p.R722H) mutation in the gene <it>POLG1 </it>of the catalytic subunit of human mitochondrial polymerase gamma has been previously found in a few occasions but its pathogenicity has remained uncertain. We set out to ascertain its contribution to neuromuscular disease.</p> <p>Methods</p> <p>Probands from two families with probable mitochondrial disease were examined clinically, muscle and buccal epithelial DNA were analyzed for mtDNA deletions, and the <it>POLG1, POLG2, ANT1 </it>and <it>Twinkle </it>genes were sequenced.</p> <p>Results</p> <p>An adult proband presented with progressive external ophthalmoplegia, sensorineural hearing impairment, diabetes mellitus, dysphagia, a limb myopathy and dementia. Brain MRI showed central and cortical atrophy, and ¹⁸F-deoxyglucose PET revealed reduced glucose uptake. Histochemical analysis of muscle disclosed ragged red fibers and cytochrome c oxidase-negative fibers. Electron microscopy showed subsarcolemmal aggregates of morphologically normal mitochondria. Multiple mtDNA deletions were found in the muscle, and sequencing of the <it>POLG1 </it>gene revealed a homozygous c.2447G>A (p.R722H) mutation. His two siblings were also homozygous with respect to the p.R722H mutation and presented with dementia and sensorineural hearing impairment. In another family the p.R722H mutation was found as compound heterozygosity with the common p.W748S mutation in two siblings with mental retardation, ptosis, epilepsy and psychiatric symptoms. The estimated carrier frequency of the p.R722H mutation was 1:135 in the Finnish population. No mutations in <it>POLG2</it>, <it>ANT1 </it>and <it>Twinkle </it>genes were found. Analysis of the POLG1 sequence by homology modeling supported the notion that the p.R722H mutation is pathogenic.</p> <p>Conclusions</p> <p>The recessive c.2447G>A (p.R722H) mutation in the linker region of the <it>POLG1 </it>gene is pathogenic for multiple mtDNA deletions in muscle and is associated with a late-onset neurological phenotype as a homozygous state. The onset of the disease can be earlier in compound heterozygotes.</p

The Genetic Landscape of Complex Childhood-Onset Hyperkinetic Movement Disorders

Acord transformatiu CRUE-CSICThis work was supported by an NIHR Professorship (to M.A.K.). M.A.K. has received funding from the Sir Jules Thorn Award for Biomedical Research and Wellcome Trust. B.P.-D. was supported by Instituto de Salud Carlos III, PI 18/01319 and PI21/00248, and has received funding from Beca José Castillejos (CAS14/00328). K.J.P. was supported by an MRC Clinician-Scientist Fellowship (511015) and was supported by the Dystonia Medical Research Foundation and Fight for Sight. S.S.M. has received funding from the Winston Churchill Memorial trust and Cerebral Palsy Alliance.Background and Objective: The objective of this study was to better delineate the genetic landscape and key clinical characteristics of complex, early-onset, monogenic hyperkinetic movement disorders. Methods: Patients were recruited from 14 international centers. Participating clinicians completed standardized proformas capturing demographic, clinical, and genetic data. Two pediatric movement disorder experts reviewed available video footage, classifying hyperkinetic movements according to published criteria. Results: One hundred forty patients with pathogenic variants in 17 different genes (ADCY5, ATP1A3, DDC, DHPR, FOXG1, GCH1, GNAO1, KMT2B, MICU1, NKX2.1, PDE10A, PTPS, SGCE, SLC2A1, SLC6A3, SPR, and TH) were identified. In the majority, hyperkinetic movements were generalized (77%), with most patients (69%) manifesting combined motor semiologies. Parkinsonism-dystonia was characteristic of primary neurotransmitter disorders (DDC, DHPR, PTPS, SLC6A3, SPR, TH); chorea predominated in ADCY5-, ATP1A3-, FOXG1-, NKX2.1-, SLC2A1-, GNAO1-, and PDE10A-related disorders; and stereotypies were a prominent feature in FOXG1- and GNAO1-related disease. Those with generalized hyperkinetic movements had an earlier disease onset than those with focal/segmental distribution (2.5 ± 0.3 vs. 4.7 ± 0.7 years; P = 0.007). Patients with developmental delay also presented with hyperkinetic movements earlier than those with normal neurodevelopment (1.5 ± 2.9 vs. 4.7 ± 3.8 years; P < 0.001). Effective disease-specific therapies included dopaminergic agents for neurotransmitters disorders, ketogenic diet for glucose transporter deficiency, and deep brain stimulation for SGCE-, KMT2B-, and GNAO1-related hyperkinesia. Conclusions: This study highlights the complex phenotypes observed in children with genetic hyperkinetic movement disorders that can lead to diagnostic difficulty. We provide a comprehensive analysis of motor semiology to guide physicians in the genetic investigation of these patients, to facilitate early diagnosis, precision medicine treatments, and genetic counseling. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society

Metabolic shift underlies recovery in reversible infantile respiratory chain deficiency.

Reversible infantile respiratory chain deficiency (RIRCD) is a rare mitochondrial myopathy leading to severe metabolic disturbances in infants, which recover spontaneously after 6-months of age. RIRCD is associated with the homoplasmic m.14674T>C mitochondrial DNA mutation; however, only ~ 1/100 carriers develop the disease. We studied 27 affected and 15 unaffected individuals from 19 families and found additional heterozygous mutations in nuclear genes interacting with mt-tRNAGlu including EARS2 and TRMU in the majority of affected individuals, but not in healthy carriers of m.14674T>C, supporting a digenic inheritance. Our transcriptomic and proteomic analysis of patient muscle suggests a stepwise mechanism where first, the integrated stress response associated with increased FGF21 and GDF15 expression enhances the metabolism modulated by serine biosynthesis, one carbon metabolism, TCA lipid oxidation and amino acid availability, while in the second step mTOR activation leads to increased mitochondrial biogenesis. Our data suggest that the spontaneous recovery in infants with digenic mutations may be modulated by the above described changes. Similar mechanisms may explain the variable penetrance and tissue specificity of other mtDNA mutations and highlight the potential role of amino acids in improving mitochondrial disease

Detailed Analysis of ITPR1 Missense Variants Guides Diagnostics and Therapeutic Design

BACKGROUND: The ITPR1 gene encodes the inositol 1,4,5-trisphosphate (IP3 ) receptor type 1 (IP3 R1), a critical player in cerebellar intracellular calcium signaling. Pathogenic missense variants in ITPR1 cause congenital spinocerebellar ataxia type 29 (SCA29), Gillespie syndrome (GLSP), and severe pontine/cerebellar hypoplasia. The pathophysiological basis of the different phenotypes is poorly understood. OBJECTIVES: We aimed to identify novel SCA29 and GLSP cases to define core phenotypes, describe the spectrum of missense variation across ITPR1, standardize the ITPR1 variant nomenclature, and investigate disease progression in relation to cerebellar atrophy. METHODS: Cases were identified using next-generation sequencing through the Deciphering Developmental Disorders study, the 100,000 Genomes project, and clinical collaborations. ITPR1 alternative splicing in the human cerebellum was investigated by quantitative polymerase chain reaction. RESULTS: We report the largest, multinational case series of 46 patients with 28 unique ITPR1 missense variants. Variants clustered in functional domains of the protein, especially in the N-terminal IP3 -binding domain, the carbonic anhydrase 8 (CA8)-binding region, and the C-terminal transmembrane channel domain. Variants outside these domains were of questionable clinical significance. Standardized transcript annotation, based on our ITPR1 transcript expression data, greatly facilitated analysis. Genotype-phenotype associations were highly variable. Importantly, while cerebellar atrophy was common, cerebellar volume loss did not correlate with symptom progression. CONCLUSIONS: This dataset represents the largest cohort of patients with ITPR1 missense variants, expanding the clinical spectrum of SCA29 and GLSP. Standardized transcript annotation is essential for future reporting. Our findings will aid in diagnostic interpretation in the clinic and guide selection of variants for preclinical studies. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society

Clinical, biochemical, cellular and molecular characterization of mitochondrial DNA depletion syndrome due to novel mutations in the MPV17 gene

Mitochondrial DNA (mtDNA) depletion syndromes (MDS) are severe autosomal recessive disorders associated with decreased mtDNA copy number in clinically affected tissues. The hepatocerebral form (mtDNA depletion in liver and brain) has been associated with mutations in the POLG, PEO1 (Twinkle), DGUOK and MPV17 genes, the latter encoding a mitochondrial inner membrane protein of unknown function. The aims of this study were to clarify further the clinical, biochemical, cellular and molecular genetic features associated with MDS due to MPV17 gene mutations. We identified 12 pathogenic mutations in the MPV17 gene, of which 11 are novel, in 17 patients from 12 families. All patients manifested liver disease. Poor feeding, hypoglycaemia, raised serum lactate, hypotonia and faltering growth were common presenting features. mtDNA depletion in liver was demonstrated in all seven cases where liver tissue was available. Mosaic mtDNA depletion was found in primary fibroblasts by PicoGreen staining. These results confirm that MPV17 mutations are an important cause of hepatocerebral mtDNA depletion syndrome, and provide the first demonstration of mosaic mtDNA depletion in human MPV17 mutant fibroblast cultures. We found that a severe clinical phenotype was associated with profound tissue-specific mtDNA depletion in liver, and, in some cases, mosaic mtDNA depletion in fibroblasts

Starting a DBS service for children:it’s not the latitude but the attitude — establishment of the paediatric DBS centre in Northern Finland

Abstract Objective: Paediatric movement disorder patients can benefit from deep brain stimulation (DBS) treatment and it should be offered in a timely manner. In this paper we describe our experience establishing a DBS service for paediatric patients. Methods: We set out to establish a paediatric DBS (pDBS) procedure in Oulu University Hospital in northern Finland, where up to this point DBS treatment for movement disorders had been available for adult patients. Collaboration with experienced centres aided in the process. Results: A multidisciplinary team was assembled and a systematic protocol for patient evaluation and treatment was created, with attention to special features of the regional health care system. All of our first paediatric patients had very severe movement disorders, which is typical for a new DBS centre. The patients benefitted from pDBS treatment despite variable aetiologies of movement disorders, which included cerebral palsy and rare genetic disorders with variants in PDE10A, TPK1 and ARX. We also present our high-quality paediatric MR-imaging protocol with tractography. Conclusions: Establishment of a pDBS centre requires expertise in classification of paediatric movement disorders, longstanding experience in adult DBS and a committed multidisciplinary team. Besides high-quality imaging and a skilled neurosurgery team, careful patient selection, realistic treatment goals and experience in rehabilitation are imperative in pDBS treatment