ABSTRACT
Introduction: Spinal muscular atrophy (SMA) is caused by homozygous loss of the SMN1 gene with SMN2 gene copy number correlating with disease severity. Rarely SMA is caused by a deletion on one allele and a pathogenic variant on the other. The pathogenic missense variant c.5C>G (p.Ala2Gly) correlates with a mild disease phenotype that does not correlate with SMN2 copy number. In a mouse model the c.5C>G transgene produces SMN that is thought to form partially functional SMN complexes, but levels in humans have not yet been investigated. Methods: We identified two patients with mild SMA caused by a heterozygous deletion of SMN1 and the heterozygous variant, c.5C>G. Molecular findings were confirmed with deletion/duplication analysis and Sanger sequencing. Skin fibroblasts were collected and cultured, and SMN expression was analyzed using immunofluorescence. Results: Two patients with slowly progressing mild weakness were confirmed to have heterozygous pathogenic missense variant c.5C>G and a heterozygous deletion of SMN1. Their clinical presentation revealed much milder disease progression than patients with matched SMN2 copy number. Analysis of the patients' fibroblasts revealed much higher numbers of SMN nuclear complexes than a patient with a homozygous SMN1 deletion and matched SMN2 copy number. Conclusions: These case reports reinforce that the rare c.5C>G variant causes mild disease. Furthermore, the analysis of SMA nuclear gems in patient samples supports the theory that the p.Ala2Gly SMN can form partially functional SMN complexes that may carry out essential cellular functions and result in mild disease.
ABSTRACT
Treatment of phenylketonuria (PKU) has evolved since the initial introduction of a phenylalanine (Phe) restricted diet. The most recent option for adults affected with PKU is treatment with an alternate enzyme, phenylalanine ammonia lyase (PAL), that metabolizes excess Phe. Proper management of all patients with PKU relies on accurate measurement of Phe levels in blood, to comply with guidance intended to minimize the neurological symptoms. Recently, our laboratory was notified of discrepant results for a patient with PKU who is treated with pegvaliase. Two specimens were collected at the same time but yielded unexpectedly different Phe concentrations. After exclusion of specimen mix-ups or analytical errors, we suspected that there was residual pegvaliase activity in the specimens continuing to degrade Phe after collection. To investigate this possibility, we performed spiking studies that showed the degradation of Phe over time at ambient temperatures. Sample preparation by protein crash appears to deactivate pegvaliase and prevents further Phe degradation. However, because pegvaliase deactivation would be required immediately following blood collection, appropriate mitigation measures must be implemented, including stringent pre-analytical requirements, alternate sample matrices such as dried blood spots, or point of care testing. Until then, health care professionals need to be cautious in their interpretation of Phe levels in their patients with PKU that are treated with pegvaliase.
ABSTRACT
Creatine transporter deficiency has been described with normal or uninformative levels of creatine and creatinine in plasma, while urine has been the preferred specimen type for biochemical diagnosis. We report a cohort of untreated patients with creatine transporter deficiency and abnormal plasma creatine panel results, characterized mainly by markedly decreased plasma creatinine. We conclude that plasma should be considered a viable specimen type for the biochemical diagnosis of this disorder, and abnormal results should be followed up with further confirmatory testing.
Subject(s)
Brain Diseases, Metabolic, Inborn , Creatine , Creatine/deficiency , Creatinine , Mental Retardation, X-Linked , Plasma Membrane Neurotransmitter Transport Proteins , Plasma Membrane Neurotransmitter Transport Proteins/deficiency , Humans , Creatine/blood , Creatine/urine , Creatinine/blood , Creatinine/urine , Plasma Membrane Neurotransmitter Transport Proteins/genetics , Plasma Membrane Neurotransmitter Transport Proteins/blood , Male , Female , Mental Retardation, X-Linked/genetics , Mental Retardation, X-Linked/blood , Mental Retardation, X-Linked/diagnosis , Child , Child, Preschool , Nerve Tissue Proteins/genetics , Nerve Tissue Proteins/blood , Nerve Tissue Proteins/deficiency , Infant , Adolescent , Membrane Transport Proteins/genetics , Membrane Transport Proteins/deficiency , Membrane Transport Proteins/blood , AdultABSTRACT
Inborn errors of metabolism (IEMs) encompass a diverse group of disorders that can be difficult to classify due to heterogenous clinical, molecular, and biochemical manifestations. Untargeted metabolomics platforms have become a popular approach to analyze IEM patient samples because of their ability to detect many metabolites at once, accelerating discovery of novel biomarkers, and metabolic mechanisms of disease. However, there are concerns about the reproducibility of untargeted metabolomics research due to the absence of uniform reporting practices, data analyses, and experimental design guidelines. Therefore, we critically evaluated published untargeted metabolomic platforms used to characterize IEMs to summarize the strengths and areas for improvement of this technology as it progresses towards the clinical laboratory. A total of 96 distinct IEMs were collectively evaluated by the included studies. However, most of these IEMs were evaluated by a single untargeted metabolomic method, in a single study, with a limited cohort size (55/96, 57%). The goals of the included studies generally fell into two, often overlapping, categories: detecting known biomarkers from many biochemically distinct IEMs using a single platform, and detecting novel metabolites or metabolic pathways. There was notable diversity in the design of the untargeted metabolomic platforms. Importantly, the majority of studies reported adherence to quality metrics, including the use of quality control samples and internal standards in their experiments, as well as confirmation of at least some of their feature annotations with commercial reference standards. Future applications of untargeted metabolomics platforms to the study of IEMs should move beyond single-subject analyses, and evaluate reproducibility using a prospective, or validation cohort.
Subject(s)
Metabolism, Inborn Errors , Humans , Reproducibility of Results , Prospective Studies , Metabolism, Inborn Errors/diagnosis , Metabolism, Inborn Errors/metabolism , Metabolomics/methods , Biomarkers/metabolismABSTRACT
Measurement of plasmalogens is useful for the biochemical diagnosis of rhizomelic chondrodysplasia punctata (RCDP) and is also informative for Zellweger spectrum disorders (ZSD). We have developed a test method for the simultaneous quantitation of C16:0, C18:0, and C018:1 plasmalogen (PG) species and their corresponding fatty acids (FAs) in dried blood spots (DBS) and erythrocytes (RBC) by using capillary gas chromatography-mass spectrometry. Normal reference ranges for measured markers and 10 calculated ratios were established by the analysis of 720 and 473 unaffected DBS and RBC samples, respectively. Determination of preliminary disease ranges was made by using 45 samples from 43 unique patients: RCDP type 1 (DBS: 1 mild, 17 severe; RBC: 1 mild, 6 severe), RCDP type 2 (DBS: 2 mild, 1 severe; RBC: 2 severe), RCDP type 3 (DBS: 1 severe), RCDP type 4 (RBC: 2 severe), and ZSD (DBS: 3 severe; RBC: 2 mild, 7 severe). Postanalytical interpretive tools in Collaborative Laboratory Integrated Reports (CLIR) were used to generate an integrated score and a likelihood of disease. In conjunction with a review of clinical phenotype, phytanic acid, and very long-chain FA test results, the CLIR analysis allowed for differentiation between RCDP and ZSD. Data will continue to be gathered to improve CLIR analysis as more samples from affected patients with variable disease severity are analyzed. The addition of DBS analysis of PGs may allow for at-home specimen collection and second-tier testing for newborn screening programs.
Subject(s)
Chondrodysplasia Punctata, Rhizomelic , Peroxisomal Disorders , Zellweger Syndrome , Infant, Newborn , Humans , Plasmalogens , Chondrodysplasia Punctata, Rhizomelic/genetics , Peroxisomal Disorders/diagnosis , Phytanic AcidABSTRACT
Intraluminal crystalloids are a common finding within malignant prostatic acini and are infrequently identified within benign glands. The proteomic composition of these crystalloids remains poorly understood and may provide insight regarding prostate cancer pathogenesis. Laser microdissection-assisted liquid chromatography-tandem mass spectrometry (LMD-LC-MS/MS) was performed to compare proteomic composition of corpora amylacea within benign acini (n = 9), prostatic adenocarcinoma-associated crystalloids (n = 8), benign (n = 8), and malignant prostatic acini (n = 6). The expression of candidate biomarkers was then measured in urine specimens from patients with (n = 8) and without prostate cancer (n = 10) using ELISA, and immunohistochemistry-based expression in adjacent prostate cancer and benign glands was assessed in 56 whole-slide sections from radical prostatectomy specimens. LMD-LC-MS/MS revealed enrichment for the C-terminal portion of growth and differentiation factor 15 (GDF15) in prostatic crystalloids. Although urinary GDF15 levels were higher in patients with prostatic adenocarcinoma compared to those without (median: 1561.2 versus 1101.3, arbitrary units), this did not meet statistical significance (P = 0.07). Immunohistochemistry for GDF15 revealed occasional positivity in benign glands (median H-score: 30, n = 56), and diffuse positivity in prostatic adenocarcinoma (median H-score: 200, n = 56, P < 0.0001). No significant difference was identified within different prognostic grade groups of prostatic adenocarcinoma, or within malignant glands with large cribriform morphology. Our results show that the C-terminal portion of GDF15 is enriched in prostate cancer-associated crystalloids, and higher GDF15 expression is seen in malignant rather than benign prostatic acini. Improved understanding of the proteomic composition of prostate cancer-associated crystalloids provides the rationale for evaluating GDF15 as a urine-based biomarker of prostate cancer.
Subject(s)
Adenocarcinoma , Prostatic Neoplasms , Male , Humans , Chromatography, Liquid , Proteomics , Tandem Mass Spectrometry , Prostatic Neoplasms/metabolism , Crystalloid Solutions , Adenocarcinoma/pathologyABSTRACT
Clinical variability and substantial overlap between mitochondrial disorders and other genetic disorders and inborn errors make the clinical and metabolic diagnosis of mitochondrial disorders quite challenging. Evaluating specific laboratory markers is essential in the diagnostic process, but mitochondrial disease can be present in the absence of any abnormal metabolic markers. In this chapter, we share the current consensus guidelines for metabolic investigations, including investigations in blood, urine, and the cerebral spinal fluid and discuss different diagnostic approaches. As personal experience might significantly vary and there are different recommendations published as diagnostic guidelines, the Mitochondrial Medicine Society developed a consensus approach based on literature review for metabolic diagnostics in a suspected mitochondrial disease. According to the guidelines, the work-up should include the assessment of complete blood count, creatine phosphokinase, transaminases, albumin, postprandial lactate and pyruvate (lactate/pyruvate ratio when the lactate level is elevated), uric acid, thymidine, amino acids, acylcarnitines in blood, and urinary organic acids (especially screening for 3-methylglutaconic acid). Urine amino acid analysis is recommended in mitochondrial tubulopathies. CSF metabolite analysis (lactate, pyruvate, amino acids, and 5-methyltetrahydrofolate) should be included in the presence of central nervous system disease. We also suggest a diagnostic strategy based on the mitochondrial disease criteria (MDC) scoring system in mitochondrial disease diagnostics; evaluating muscle-, neurologic-, and multisystem involvement, and the presence of metabolic markers and abnormal imaging. The consensus guideline encourages a primary genetic approach in diagnostics and only suggests a more invasive diagnostic approach with tissue biopsies (histology, OXPHOS measurements, etc.) after nonconclusive genetic testing.
Subject(s)
Mitochondrial Diseases , Humans , Mitochondrial Diseases/diagnosis , Mitochondria , Amino Acids , Pyruvic Acid , Lactic AcidABSTRACT
The clinical and largely unpredictable heterogeneity of phenotypes in patients with mitochondrial disorders demonstrates the ongoing challenges in the understanding of this semi-autonomous organelle in biology and disease. Previously, we used the gene-breaking transposon to create 1200 transgenic zebrafish strains tagging protein-coding genes (Ichino et al., 2020), including the lrpprc locus. Here, we present and characterize a new genetic revertible animal model that recapitulates components of Leigh Syndrome French Canadian Type (LSFC), a mitochondrial disorder that includes diagnostic liver dysfunction. LSFC is caused by allelic variations in the LRPPRC gene, involved in mitochondrial mRNA polyadenylation and translation. lrpprc zebrafish homozygous mutants displayed biochemical and mitochondrial phenotypes similar to clinical manifestations observed in patients, including dysfunction in lipid homeostasis. We were able to rescue these phenotypes in the disease model using a liver-specific genetic model therapy, functionally demonstrating a previously under-recognized critical role for the liver in the pathophysiology of this disease.
Subject(s)
Disease Models, Animal , Liver Diseases , Mitochondrial Diseases , Animals , Canada , Genetic Therapy , Liver Diseases/genetics , Liver Diseases/therapy , Mitochondrial Diseases/genetics , Mitochondrial Diseases/therapy , Neoplasm Proteins/genetics , Zebrafish/geneticsABSTRACT
BACKGROUND: Primary hyperoxaluria (PH) type 3 (PH3) is caused by mutations in the hydroxy-oxo-glutarate aldolase 1 gene. PH3 patients often present with recurrent urinary stone disease in the first decade of life, but prior reports suggested PH3 may have a milder phenotype in adults. This study characterized clinical manifestations of PH3 across the decades of life in comparison with PH1 and PH2. METHODS: Clinical information was obtained from the Rare Kidney Stone Consortium PH Registry (PH1, n = 384; PH2, n = 51; PH3, n = 62). RESULTS: PH3 patients presented with symptoms at a median of 2.7 years old compared with PH1 (4.9 years) and PH2 (5.7 years) (P = 0.14). Nephrocalcinosis was present at diagnosis in 4 (7%) PH3 patients, while 55 (89%) had stones. Median urine oxalate excretion was lowest in PH3 patients compared with PH1 and PH2 (1.1 versus 1.6 and 1.5 mmol/day/1.73 m2, respectively, P < 0.001) while urine calcium was highest in PH3 (112 versus 51 and 98 mg/day/1.73 m2 in PH1 and PH2, respectively, P < 0.001). Stone events per decade of life were similar across the age span and the three PH types. At 40 years of age, 97% of PH3 patients had not progressed to end-stage kidney disease compared with 36% PH1 and 66% PH2 patients. CONCLUSIONS: Patients with all forms of PH experience lifelong stone events, often beginning in childhood. Kidney failure is common in PH1 but rare in PH3. Longer-term follow-up of larger cohorts will be important for a more complete understanding of the PH3 phenotype.
Subject(s)
Hyperoxaluria, Primary , Hyperoxaluria , Nephrolithiasis , Renal Insufficiency , Female , Humans , Hyperoxaluria, Primary/diagnosis , Hyperoxaluria, Primary/genetics , Male , Mutation , PhenotypeABSTRACT
OBJECTIVE: Epalrestat, an aldose reductase inhibitor increases phosphomannomutase (PMM) enzyme activity in a PMM2-congenital disorders of glycosylation (CDG) worm model. Epalrestat also decreases sorbitol level in diabetic neuropathy. We evaluated the genetic, biochemical, and clinical characteristics, including the Nijmegen Progression CDG Rating Scale (NPCRS), urine polyol levels and fibroblast glycoproteomics in patients with PMM2-CDG. METHODS: We performed PMM enzyme measurements, multiplexed proteomics, and glycoproteomics in PMM2-deficient fibroblasts before and after epalrestat treatment. Safety and efficacy of 0.8 mg/kg/day oral epalrestat were studied in a child with PMM2-CDG for 12 months. RESULTS: PMM enzyme activity increased post-epalrestat treatment. Compared with controls, 24% of glycopeptides had reduced abundance in PMM2-deficient fibroblasts, 46% of which improved upon treatment. Total protein N-glycosylation improved upon epalrestat treatment bringing overall glycosylation toward the control fibroblasts' glycosylation profile. Sorbitol levels were increased in the urine of 74% of patients with PMM2-CDG and correlated with the presence of peripheral neuropathy, and CDG severity rating scale. In the child with PMM2-CDG on epalrestat treatment, ataxia scores improved together with significant growth improvement. Urinary sorbitol levels nearly normalized in 3 months and blood transferrin glycosylation normalized in 6 months. INTERPRETATION: Epalrestat improved PMM enzyme activity, N-glycosylation, and glycosylation biomarkers in vitro. Leveraging cellular glycoproteome assessment, we provided a systems-level view of treatment efficacy and discovered potential novel biosignatures of therapy response. Epalrestat was well-tolerated and led to significant clinical improvements in the first pediatric patient with PMM2-CDG treated with epalrestat. We also propose urinary sorbitol as a novel biomarker for disease severity and treatment response in future clinical trials in PMM2-CDG. ANN NEUROL 20219999:n/a-n/a.
Subject(s)
Congenital Disorders of Glycosylation/diagnosis , Enzyme Inhibitors/therapeutic use , Phosphotransferases (Phosphomutases)/deficiency , Rhodanine/analogs & derivatives , Sorbitol/urine , Thiazolidines/therapeutic use , Adolescent , Adult , Aged , Biomarkers/urine , Child , Child, Preschool , Congenital Disorders of Glycosylation/drug therapy , Congenital Disorders of Glycosylation/urine , Female , Glycosylation , Humans , Infant , Male , Middle Aged , Patient Acuity , Phosphotransferases (Phosphomutases)/urine , Prognosis , Rhodanine/therapeutic use , Young AdultABSTRACT
BACKGROUND: Glutaric acidemia type I (GA1) is an organic acidemia that is often unrecognized in the newborn period until patients suffer an acute encephalopathic crisis, which can be mistaken for nonaccidental trauma. Presymptomatic identification of GA1 patients is possible by newborn screening (NBS). However, the biochemical "low-excretor" (LE) phenotype with nearly normal levels of disease metabolites can be overlooked, which may result in untreated disease and irreversible neurological sequelae. The LE phenotype is also a potential source of false negative (FN) NBS results that merits further investigation. METHODS: Samples from six LE GA1 patients were analyzed by biochemical and molecular methods and newborn screen outcomes were retrospectively investigated. RESULTS: Five LE GA1 patients were identified that had normal NBS results and three of these presented clinically with GA1 symptoms. One additional symptomatic patient was identified who did not undergo screening. Semiquantitative urine organic acid analysis was consistent with a GA1 diagnosis in two (33%) of the six patients, while plasma glutarylcarnitine was elevated in four (67%) of the six and urine glutarylcarnitine was elevated in four (80%) of five patients. Five GCDH variants were identified in these patients; three of which have not been previously linked to the biochemical LE phenotype. CONCLUSIONS: The data presented here raise awareness of potential FN NBS results for LE GA1 patients. The LE phenotype is not protective against adverse clinical outcomes, and the possibility of FN NBS results calls for high vigilance amongst clinicians, even in the setting of a normal NBS result.
ABSTRACT
Barth syndrome is an X-linked recessive disorder caused by pathogenic variants in TAZ, which leads to a reduction in cardiolipin with a concomitant elevation of monolysocardiolipins. There is a paucity of studies characterizing changes in individual species of monolysocardiolipins, dilysocardiolipins and cardiolipin in Barth syndrome using high resolution untargeted lipidomics that can accurately annotate and quantify diverse lipids. We confirmed the structural diversity monolysocardiolipins, dilysocardiolipins and cardiolipin and identified individual species that showed previously unreported alterations in BTHS. Development of mass spectrometry-based targeted assays for these lipid biomarkers should provide an important tool for clinical diagnosis of Barth syndrome.
Subject(s)
Barth Syndrome/blood , Cardiolipins/blood , Chromatography, Liquid/methods , Tandem Mass Spectrometry/methods , Adolescent , Cardiolipins/chemistry , Cardiolipins/classification , Cell Line , Child , Humans , MaleABSTRACT
Phosphoglucomutase 1 (PGM1) catalyzes the interconversion of glucose-6-phosphate to glucose-1-phosphate and is a key enzyme of glycolysis, glycogenesis, and glycogenolysis. PGM1 deficiency (OMIM: 614921) was initially defined as a glycogen storage disorder (type XIV), and later re-classified as a PGM1-congenital disorder of glycosylation (PGM1-CDG). Serum transferrin (Tf) glycan isoform analysis by liquid chromatography-mass spectrometry (LC-MS) is used as a primary diagnostic screen tool, and reveals a very unique CDG profile described as a mixture of CDG-type I and CDG-type II patterns. Oral d-galactose supplementation shows significant clinical and metabolic improvements, which are indicated by the Tf glycan isoform normalization over time in patients with PGM1-CDG. Thus, there is a need for biomarkers to guide d-galactose dosage in patients in order to maintain effective and safe drug levels. Here, we present a simplified algorithm called PGM1-CDG Treatment Monitoring Index (PGM1-TMI) for assessing the response of PGM1-CDG patients to d-galactose supplementation. For our single-center cohort of 16 PGM1-CDG patients, the Tf glycan profile analysis provided the biochemical diagnosis in all of them. In addition, the PGM1-TMI was reduced in PGM1-CDG patients under d-galactose supplementation as compared with their corresponding values before treatment, indicating that glycosylation proceeds towards normalization. PGM1-TMI allows tracking Tf glycan isoform normalization over time when the patients are on d-galactose supplementation.
Subject(s)
Galactose/therapeutic use , Glycogen Storage Disease/drug therapy , Adult , Biomarkers/metabolism , Child , Child, Preschool , Cohort Studies , Dose-Response Relationship, Drug , Drug Monitoring , Female , Galactose/administration & dosage , Galactose/adverse effects , Glycoproteins/metabolism , Humans , Infant , Male , Mass Spectrometry , Phosphoglucomutase/metabolism , Young AdultABSTRACT
INTRODUCTION: Nonaccidental trauma (NAT) is considered when pediatric patients present with intracranial injuries and a negative history of an accidental injury or concomitant medical diagnosis. The evaluation of NAT should include the consideration of possible medical causes including coagulation, hematologic, metabolic and other genetic disorders, as well as witnessed and unwitnessed accidental injuries. CASE PRESENTATION: We present a 7-month-old male with spells and incidental findings of bilateral subdural hematomas, retinal hemorrhages, and secondary macrocephaly, leading to investigation for NAT. Biochemical analysis showed excretion of a large amount of D-2-hydroxyglutaric in urine consistent with a biochemical diagnosis of D-2-hydroxyglutaric aciduria, a rare neurometabolic disorder characterized by developmental delay, epilepsy, hypotonia, and psychomotor retardation. None of these symptoms were present in our patient at the time of diagnosis. Molecular genetic testing revealed a pathogenic splice site variant (c.685-2A>G) and a variant of uncertain significance (c.1256G>T) with evidence of pathogenicity in the D2HGDH gene, consistent with a molecular diagnosis of D-2-hydroxyglutaric aciduria type I (OMIM #600721). CONCLUSION: Since several metabolic disorders, including D-2-hydroxyglutaric aciduria type I, can present solely with symptoms suggestive of NAT (subdural and retinal hemorrhages), an early metabolic evaluation by urine organic acid analysis should be included in clinical protocols evaluating NAT. A methodical and nonjudgmental approach coordinated between pediatricians and metabolic specialists is also necessary to ensure that rare genetic conditions are not overlooked to prevent devastating social, legal, and financial consequences of suspected child abuse.
ABSTRACT
Mitochondrial metabolism is increasingly implicated in psychopathologies and mood disorders, including post-traumatic stress disorder (PTSD). We recently reported that mice exposed to a novel paradigm for the induction of PTSD-like behavior displayed reduced mitochondrial electron transport chain (mtETC) complex activity as well as decreased multi-system fatty acid oxidation (FAO) flux. Based on these results, we hypothesized that stressed and PTSD-like animals would display evidence of metabolic reprogramming in both cerebellum and plasma consistent with increased energetic demand, mitochondrial metabolic reprogramming, and increased oxidative stress. We performed targeted metabolomics in both cerebellar tissue and plasma, as well as untargeted nuclear magnetic resonance (NMR) spectroscopy in the cerebellum of 6 PTSD-like and 7 resilient male mice as well as 7 trauma-naïve controls. We identified numerous differences in amino acids and tricarboxylic acid (TCA) cycle metabolite concentrations in the cerebellum and plasma consistent with altered mitochondrial energy metabolism in trauma exposed and PTSD-like animals. Pathway analysis identified metabolic pathways with significant metabolic pathway shifts associated with trauma exposure, including the tricarboxylic acid cycle, pyruvate, and branched-chain amino acid metabolism in both cerebellar tissue and plasma. Altered glutamine and glutamate metabolism, and arginine biosynthesis was evident uniquely in cerebellar tissue, while ketone body levels were modified in plasma. Importantly, we also identified several cerebellar metabolites (e.g. choline, adenosine diphosphate, beta-alanine, taurine, and myo-inositol) that were sufficient to discriminate PTSD-like from resilient animals. This multilevel analysis provides a comprehensive understanding of local and systemic metabolite fingerprints associated with PTSD-like behavior, and subsequently altered brain bioenergetics. Notably, several transformed metabolic pathways observed in the cerebellum were also reflected in plasma, connecting central and peripheral biosignatures of PTSD-like behavior. These preliminary findings could direct further mechanistic studies and offer insights into potential metabolic interventions, either pharmacological or dietary, to improve PTSD resilience.
ABSTRACT
Mitochondrial disorders represent a large collection of rare syndromes that are difficult to manage both because we do not fully understand biochemical pathogenesis and because we currently lack facile markers of severity. The m.3243A>G variant is the most common heteroplasmic mitochondrial DNA mutation and underlies a spectrum of diseases, notably mitochondrial encephalomyopathy lactic acidosis and stroke-like episodes (MELAS). To identify robust circulating markers of m.3243A>G disease, we first performed discovery proteomics, targeted metabolomics, and untargeted metabolomics on plasma from a deeply phenotyped cohort (102 patients, 32 controls). In a validation phase, we measured concentrations of prioritized metabolites in an independent cohort using distinct methods. We validated 20 analytes (1 protein, 19 metabolites) that distinguish patients with MELAS from controls. The collection includes classic (lactate, alanine) and more recently identified (GDF-15, α-hydroxybutyrate) mitochondrial markers. By mining untargeted mass-spectra we uncovered 3 less well-studied metabolite families: N-lactoyl-amino acids, ß-hydroxy acylcarnitines, and ß-hydroxy fatty acids. Many of these 20 analytes correlate strongly with established measures of severity, including Karnofsky status, and mechanistically, nearly all markers are attributable to an elevated NADH/NAD+ ratio, or NADH-reductive stress. Our work defines a panel of organelle function tests related to NADH-reductive stress that should enable classification and monitoring of mitochondrial disease.
Subject(s)
MELAS Syndrome/blood , Adolescent , Adult , Aged , Aged, 80 and over , Alanine/blood , Biomarkers/blood , Child , Child, Preschool , Female , Growth Differentiation Factor 15/blood , Humans , Hydroxybutyrates/blood , Lactic Acid/blood , MELAS Syndrome/genetics , Male , Middle Aged , Mutation , Severity of Illness IndexABSTRACT
Glycosylation is a critical post/peri-translational modification required for the appropriate development and function of the immune system. As an example, abnormalities in glycosylation can cause antibody deficiency and reduced lymphocyte signaling, although the phenotype can be complex given the diverse roles of glycosylation. Human MGAT2 encodes N-acetylglucosaminyltransferase II, which is a critical enzyme in the processing of oligomannose to complex N-glycans. Complex N-glycans are essential for immune system functionality, but only one individual with MGAT2-CDG has been described to have an abnormal immunologic evaluation. MGAT2-CDG (CDG-IIa) is a congenital disorder of glycosylation (CDG) associated with profound global developmental disability, hypotonia, early onset epilepsy, and other multisystem manifestations. Here, we report a 4-year old female with MGAT2-CDG due to a novel homozygous pathogenic variant in MGAT2, a 4-base pair deletion, c.1006_1009delGACA. In addition to clinical features previously described in MGAT2-CDG, she experienced episodic asystole, persistent hypogammaglobulinemia, and defective ex vivo mitogen and antigen proliferative responses, but intact specific vaccine antibody titers. Her infection history has been mild despite the testing abnormalities. We compare this patient to the 15 previously reported patients in the literature, thus expanding both the genotypic and phenotypic spectrum for MGAT2-CDG.
Subject(s)
Arrhythmias, Cardiac/genetics , Congenital Disorders of Glycosylation/genetics , Immune System Diseases/genetics , N-Acetylglucosaminyltransferases/genetics , Arrhythmias, Cardiac/complications , Arrhythmias, Cardiac/immunology , Arrhythmias, Cardiac/pathology , Child, Preschool , Congenital Disorders of Glycosylation/complications , Congenital Disorders of Glycosylation/immunology , Congenital Disorders of Glycosylation/pathology , Female , Glycosylation , Homozygote , Humans , Immune System Diseases/complications , Immune System Diseases/immunology , Immune System Diseases/pathology , Mutation/genetics , N-Acetylglucosaminyltransferases/immunology , PhenotypeABSTRACT
Acylcarnitine analysis is a useful test for identifying patients with inborn errors of mitochondrial fatty acid ß-oxidation and certain organic acidemias. Plasma is routinely used in the diagnostic workup of symptomatic patients. Urine analysis of targeted acylcarnitine species may be helpful in the diagnosis of glutaric acidemia type I and other disorders in which polar acylcarnitine species accumulate. For newborn screening applications, dried blood spot acylcarnitine analysis can be performed as a multiplex assay with other analytes, including amino acids, succinylacetone, guanidinoacetate, creatine, and lysophosphatidylcholines. Tandem mass spectrometric methodology, established more than 30 years ago, remains a valid approach for acylcarnitine analysis. The method involves flow-injection analysis of esterified or underivatized acylcarnitines species and detection using a precursor-ion scan. Alternative methods utilize liquid chromatographic separation of isomeric and isobaric species and/or detection by selected reaction monitoring. These technical standards were developed as a resource for diagnostic laboratory practices in acylcarnitine analysis, interpretation, and reporting.
Subject(s)
Genetics, Medical , Laboratories , Carnitine/analogs & derivatives , Genomics , Humans , Infant, Newborn , United StatesABSTRACT
BACKGROUND: (+)-Epicatechin (EPI) induces mitochondrial biogenesis and antioxidant metabolism in muscle fibers and neurons. We aimed to evaluate safety and efficacy of (+)-EPI in pediatric subjects with Friedreich's ataxia (FRDA). METHODS: This was a phase II, open-label, baseline-controlled single-center trial including 10 participants ages 10 to 22 with confirmed FA diagnosis. (+)-EPI was administered orally at 75 mg/d for 24 weeks, with escalation to 150 mg/d at 12 weeks for subjects not showing improvement of neuromuscular, neurological or cardiac endpoints. Neurological endpoints were change from baseline in Friedreich's Ataxia Rating Scale (FARS) and 8-m timed walk. Cardiac endpoints were changes from baseline in left ventricular (LV) structure and function by cardiac magnetic resonance imaging (MRI) and echocardiogram, changes in cardiac electrophysiology, and changes in biomarkers for heart failure and hypertrophy. RESULTS: Mean FARS/modified (m)FARS scores showed nonstatistically significant improvement by both group and individual analysis. FARS/mFARS scores improved in 5/9 subjects (56%), 8-m walk in 3/9 (33%), 9-peg hole test in 6/10 (60%). LV mass index by cardiac MRI was significantly reduced at 12 weeks (P = .045), and was improved in 7/10 (70%) subjects at 24 weeks. Mean LV ejection fraction was increased at 24 weeks (P = .008) compared to baseline. Mean maximal septal thickness by echocardiography was increased at 24 weeks (P = .031). There were no serious adverse events. CONCLUSION: (+)-EPI was well tolerated over 24 weeks at up to 150 mg/d. Improvement was observed in cardiac structure and function in subset of subjects with FRDA without statistically significant improvement in primary neurological outcomes. SYNOPSIS: A (+)-epicatechin showed improvement of cardiac function, nonsignificant reduction of FARS/mFARS scores, and sustained significant upregulation of muscle-regeneration biomarker follistatin.