Background: Leigh syndrome (LS) is a rare disease caused by mitochondrial defects and has high phenotypic and genotypic heterogeneity. We analyzed the clinical symptoms, neuroimaging, muscular histopathology, and genotypes of 13 Chinese LS patients with mitochondrial DNA (mtDNA) mutations. Methods: Mutations in mtDNA were identified by targeted sequencing. The brain imaging features on magnetic resonance imaging (MRI) were analyzed. The levels of lactate in fasting blood and cerebrospinal fluid (CSF) were routinely tested. The levels of urinary organic acids, plasma amino acids, and acylcarnitines were examined with gas chromatography-mass spectrometry and tandem mass spectrometry. The histopathological traits of skeletal muscles were analyzed under microscope. Results: Among 13 patients, mutations of MT-NDs (n = 8) and MT-ATP6 (n = 4) genes were most common. Strabismus (8/13), muscle weakness (8/13), and ataxia (5/13) were also common, especially for the patients with late-onset age after 2 years old. However, respiratory distress was common in patients with early-onset age before 2 years old. The most frequently affected brain area in these patients was the brain stem (12/13), particularly the dorsal part of midbrain, followed by basal ganglia (6/13), thalamus (6/13), cerebellum (5/13), and supratentorial white matter (2/13). Besides, the elevated lactate levels in CSF (6/6) were more common than those in serum (7/13). However, the analysis of abnormal plasma amino acid and urinary organic acid showed limited results (0/3 and 1/4, respectively). Muscular histopathology showed mitochondrial myopathy in the three late-onset patients but not in the early-onset ones. Conclusions Noninvasive genetic screening is recommended for mtDNA mutations in MT-NDs and MT-ATP6 genes in patients with ophthalmoplegia, muscle weakness, ataxia, and respiratory disorder. Furthermore, the lactate detection in CSF and the brain MRI scanning are suggested as the diagnosis methods for LS patients with mtDNA mutations.
Child ; Child, Preschool ; Creatine Kinase ; blood ; Cytochrome-c Oxidase Deficiency ; DNA, Mitochondrial ; genetics ; Fasting ; blood ; cerebrospinal fluid ; Female ; Humans ; Infant ; Lactic Acid ; blood ; cerebrospinal fluid ; Leigh Disease ; diagnostic imaging ; genetics ; Magnetic Resonance Imaging ; Male ; Mutation ; genetics ; Neuroimaging ; methods

OBJECTIVE: : To screen genes involved in mitochondrial DNA (mtDNA) damage repair in rats with septic acute kidney injury (SAKI). METHODS: : Forty male C57BL/6J mice were randomly divided into SAKI group (=28) and sham operation group (=12). The SAKI mouse model was established by cecal ligation and puncture. Blood and kidney samples were collected at 8, 24, and 48 h after surgery. Serum creatinine and urea nitrogen were measured by a dry biochemical analyzer. Serum inflammatory cytokines were detected by ELISA. Histopathological changes were observed with HE staining. The mtDNA damage repair related genes were screened by RNA sequencing and bioinformatics analysis; the mRNA and protein expression levels of related genes were detected by real-time quantitative RT-PCR and immunohistochemisry, respectively. RESULTS: : Symptoms of sepsis were observed in SAKI group, and 16 out of 28 mice were died in the SAKI group; serum TNF-α, IL-6, creatinine and urea nitrogen levels were higher than those in the sham group (<0.05 or <0.01). Histopathological examination in SAKI group showed that renal tubular epithelial cells were swollen, inflammatory cells infiltrated, and a large number of cell vacuoles were seen, suggesting successful modeling. Mitochondrial DNA damage repair related genes and were screened out. The expression of these genes was detected by real-time RT-PCR, and the results were consistent with RNA sequencing trends. Immunohistochemical staining showed that Gadd45α was mainly expressed in the nucleus of renal tubular epithelial cells, and the positive rate of Gadd45α in SAKI group was higher than that in the sham operation group (<0.05). CONCLUSIONS : and genes are involved in mtDNA damage repair in rats with SAKI, indicating that these genes may be used as new targets for prevention and treatment of SAKI.
Acute Kidney Injury ; physiopathology ; Animals ; Blood Urea Nitrogen ; Creatinine ; blood ; DNA Repair ; genetics ; DNA, Mitochondrial ; Male ; Mice ; Mice, Inbred C57BL ; Random Allocation ; Rats ; Sepsis ; physiopathology

BACKGROUNDMitochondrial diseases are a group of energy metabolic disorders with multisystem involvements. Variable clinical features present a major challenge in pediatric diagnoses. We summarized the clinical spectrum of m.3243A>G mutation in Chinese pediatric patients, to define the common clinical manifestations and study the correlation between heteroplasmic degree of the mutation and clinical severity of the disease.

METHODSClinical data of one-hundred pediatric patients with symptomatic mitochondrial disease harboring m.3243A>G mutation from 2007 to 2013 were retrospectively reviewed. Detection of m.3243A>G mutation ratio was performed by polymerase chain reaction (PCR)-restriction fragment length polymorphism. Correlation between m.3243A>G mutation ratio and age was evaluated. The differences in clinical symptom frequency of patients with low, middle, and high levels of mutation ratio were analyzed by Chi-square test.

RESULTSSixty-six patients (66%) had suffered a delayed diagnosis for an average of 2 years. The most frequent symptoms were seizures (76%), short stature (73%), elevated plasma lactate (70%), abnormal magnetic resonance imaging/computed tomography (MRI/CT) changes (68%), vomiting (55%), decreased vision (52%), headache (50%), and muscle weakness (48%). The mutation ratio was correlated negatively with onset age (r = -0.470, P < 0.001). Myopathy was more frequent in patients with a high level of mutation ratio. However, patients with a low or middle level of m.3243A>G mutation ratio were more likely to suffer hearing loss, decreased vision, and gastrointestinal disturbance than patients with a high level of mutation ratio.

CONCLUSIONSOur study showed that half of Chinese pediatric patients with m.3243A>G mutation presented seizures, short stature, abnormal MRI/CT changes, elevated plasma lactate, vomiting, and headache. Pediatric patients with these recurrent symptoms should be considered for screening m.3243A>G mutation. Clinical manifestations and laboratory abnormalities should be carefully monitored in patients with this point mutation.

Adolescent ; Age of Onset ; Asian Continental Ancestry Group ; Chi-Square Distribution ; Child ; Child, Preschool ; DNA, Mitochondrial ; genetics ; Female ; Hearing Loss ; pathology ; physiopathology ; Humans ; Infant ; Lactic Acid ; blood ; Magnetic Resonance Imaging ; Male ; Mitochondrial Diseases ; blood ; genetics ; pathology ; physiopathology ; Mutation ; Point Mutation ; genetics ; Polymerase Chain Reaction ; Retrospective Studies ; Seizures ; pathology ; physiopathology

OBJECTIVETo investigate the relationship between the changes of the copy numbers of mtDNA in peripheral blood mono-nucle- ar cell(PBMC) and the disordered of antioxidant capacity of hepatocellular carcinoma (HCC) patients.

METHODSThe Ficoll Hypaque method was used to isolate the PBMC from blood specimens. The ND1 gene of the mitochondrial was amplified by real-time PCR; meantime β-actin was served as a quantitative standard marker; the difference of mtDNA copy number in PBMC was compared between HCC and healthy control group. The level of reactive oxygen species (ROS) in PBMC was determined by flow cytometry. The change of total antioxidant capacity (T- AOC) of plasma was detected by the biochemistry examination.

RESULTSThe copy numbers of ND1 gene in PBMC of HCC was 73% that of the healthy control group,which suggested a decrease of the copy numbers of mtDNA in HCC. The levels of ROS of PBMC in HCC was (417. 82 ± 110.62) and (301.82 ± 75.54) in control group, which showed that the levels of ROS of PBMC in HCC were significant higher than that in control group (P < 0.01).Plasma T-AOC in HCC was (1.30 ± 0.85), and (3.20 ± 1.62) in control. The T-AOC of plasma of HCC was significantly lower than in control group (P < 0.01).

CONCLUSIONThere was a certain relationship between the decrease of the copy numbers of mtDNA and the disordered antioxidant capacity in hepatocellular carcinoma, which may be associated with the development of hepatocellular carcinoma.

Actins ; Antioxidants ; metabolism ; Carcinoma, Hepatocellular ; blood ; genetics ; Case-Control Studies ; DNA Copy Number Variations ; DNA, Mitochondrial ; genetics ; Humans ; Leukocytes, Mononuclear ; metabolism ; Liver Neoplasms ; blood ; genetics ; Reactive Oxygen Species ; metabolism

BACKGROUND: A newborn screening (NBS) program has been utilized to detect asymptomatic newborns with inherited metabolic diseases (IMDs). There have been some bottlenecks such as false-positives and imprecision in the current NBS tests. To overcome these issues, we developed a multigene panel for IMD testing and investigated the utility of our integrated screening model in a routine NBS environment. We also evaluated the genetic epidemiologic characteristics of IMDs in a Korean population. METHODS: In total, 269 dried blood spots with positive results from current NBS tests were collected from 120,700 consecutive newborns. We screened 97 genes related to NBS in Korea and detected IMDs, using an integrated screening model based on biochemical tests and next-generation sequencing (NGS) called NewbornSeq. Haplotype analysis was conducted to detect founder effects. RESULTS: The overall positive rate of IMDs was 20%. We identified 10 additional newborns with preventable IMDs that would not have been detected prior to the implementation of our NGS-based platform NewbornSeq. The incidence of IMDs was approximately 1 in 2,235 births. Haplotype analysis demonstrated founder effects in p.Y138X in DUOXA2, p.R885Q in DUOX2, p.Y439C in PCCB, p.R285Pfs*2 in SLC25A13, and p.R224Q in GALT. CONCLUSIONS: Through a population-based study in the NBS environment, we highlight the screening and epidemiological implications of NGS. The integrated screening model will effectively contribute to public health by enabling faster and more accurate IMD detection through NBS. This study suggested founder mutations as an explanation for recurrent IMD-causing mutations in the Korean population.
Computational Biology ; DNA/chemistry/isolation & purification/metabolism ; Dried Blood Spot Testing ; Galactokinase ; Genomics ; Haplotypes ; High-Throughput Nucleotide Sequencing ; Humans ; Incidence ; Infant, Newborn ; Membrane Proteins/genetics ; Metabolic Diseases/*diagnosis/epidemiology/genetics ; Metabolism, Inborn Errors/diagnosis/epidemiology/genetics ; Mitochondrial Membrane Transport Proteins/genetics ; Neonatal Screening ; Polymorphism, Genetic ; Republic of Korea/epidemiology ; Sequence Analysis, DNA

OBJECTIVETo explore the role of mitochondrial DNA 5178 C/A (Mt5178) polymorphism of NADH-dehydrogenase subunit 2 (ND2) gene in type-2 diabetes mellitus (T2DM) among ethnic Han Chinese through a case-control study.

METHODSThe Mt5178C/A polymorphism was determined by sequencing 1103 T2DM patients and 791 healthy controls. Logistic regression analysis was conducted to estimate odds ratios (OR) and 95% confidence intervals (CI). To confirm the results, a meta-analysis was conducted based on published literature on the association of Mt5178 variant with T2DM.

RESULTSNo significant association was found between the Mt5178C/A variant and T2DM either by our study or the meta-analysis which included eight published studies. Nevertheless, it was found that the T2DM patients with 5178C genotype were at a higher risk for nephropathy complication (OR=1.49, 95%CI: 1.005-2.197, P<0.05) and at significantly lower risk for hypertension complication (OR=0.744, 95%CI: 0.556-0.996, P<0.05) compared with those carrying a 5178A genotype.

CONCLUSIONNo association was found between the Mt5178C/A polymorphism of mitochondrial ND2 gene with the increased risk of T2DM. However, the polymorphism may affect the development of nephropathy and hypertension complications among T2DM patients.

Adult ; Aged ; Blood Glucose ; metabolism ; Cholesterol ; blood ; Cholesterol, HDL ; blood ; DNA, Mitochondrial ; chemistry ; genetics ; Diabetes Complications ; blood ; genetics ; Diabetes Mellitus, Type 2 ; blood ; complications ; genetics ; Diabetic Nephropathies ; blood ; genetics ; Fasting ; blood ; Female ; Humans ; Hypertension ; blood ; complications ; genetics ; Male ; Meta-Analysis as Topic ; Middle Aged ; Odds Ratio ; Polymorphism, Single Nucleotide ; Sequence Analysis, DNA ; Triglycerides ; blood

PURPOSE: Recently, mitochondrial DNA 4977bp deletion (mtDNA4977-mut), a somatic mutation related to oxidative stress, has been shown to be associated with atrial fibrillation (AF). We hypothesized that patient age, as well as electroanatomical characteristics of fibrillating left atrial (LA), vary depending on the presence of mtDNA4977-mut in peripheral blood among patients with non-valvular AF. MATERIALS AND METHODS: Analyzing clinical and electroanatomical characteristics, we investigated the presence of the mtDNA4977-mut in peripheral blood of 212 patients (51.1+/-13.2 years old, 83.5% male) undergoing catheter ablation for non-valvular AF, as well as 212 age-matched control subjects. RESULTS: The overall frequency of peripheral blood mtDNA4977-mut in patients with AF and controls was not significantly different (24.5% vs. 19.3%, p=0.197). When the AF patient group was stratified according to age, mtDNA4977-mut was more common (47.4% vs. 20.0%, p=0.019) in AF patients older than 65 years than their age-matched controls. Among AF patients, those with mtDNA4977-mut were older (58.1+/-11.9 years old vs. 48.8+/-11.9 years old, p<0.001). AF patients positive for the mtDNA mutation had greater LA dimension (p=0.014), higher mitral inflow peak velocity (E)/diastolic mitral annular velocity (Em) ratio (p<0.001), as well as lower endocardial voltage (p=0.035), and slower conduction velocity (p=0.048) in the posterior LA than those without the mutation. In multivariate analysis, E/Em ratio was found to be significantly associated with the presence of mtDNA4977-mut in peripheral blood. CONCLUSION: mtDNA4977-mut, an age-related somatic mutation detected in the peripheral blood, is associated with advanced age and electro-anatomical remodeling of the atrium in non-valvular AF.
Adult ; Aged ; Atrial Fibrillation/blood/*genetics/*physiopathology ; Atrial Remodeling/*genetics ; Base Pairing/*genetics ; Case-Control Studies ; DNA, Mitochondrial/*blood/*genetics ; Female ; Heart Atria/pathology/physiopathology ; Humans ; Kaplan-Meier Estimate ; Logistic Models ; Male ; Middle Aged ; Mutation Rate ; Phenotype ; Sequence Deletion/*genetics

OBJECTIVETo achieve early diagnosis for inheritable hearing loss and determine carrier rate of deafness causing gene mutations in order to provide information for premarital, prenatal and postnatal genetic counseling.

METHODSA total of 17 000 dried heel blood spots of normal newborns in Chengdu were collected with informed consent obtained from their parents. Genomic DNA was extracted from dried blood spots using Qiagen DNA extraction kits. Microarrays with 9 common mutation loci of 4 deafness-associated genes in Chinese population were used. Nine hot mutations including GJB2 (35delG, 176del16, 235delC and 299delAT), GJB3 (538C> T), SLC26A4 (IVS 7-2A> G, 2168A> G), and mitochondrial DNA 12S rRNA (1555A> G, 1494C> T) were detected by PCR amplification and microarray hybridization. Mutations detected by microarray were verified by Sanger DNA sequencing.

RESULTSOf the 17 000 new-borns, 542 neonates had mutations of the 4 genes. Heterozygous mutations of GJB2, at 235delC, 299delAT, and 176del16 were identified in 254, 55, and 15 newborns, respectively. Two newborns had homozygous mutation of GJB2, 235delC. Heterozygous mutations at 538C> T of GJB3, 2168A> G and IVS 7-2A> G of SLC26A4 were found in 23, 17 and 128 newborns, respectively. For mutation analysis of mitochondrial DNA 12S rRNA, 1494C> T and 1555A> G were homogeneous mutations in 4 and 42 neonates, respectively. In addition, 6 complexity mutations were detected, which demonstrated that one newborn had heterozygous mutations at GJB2 235delC and SLC26A4, IVS7-2A> G, one had heterozygous mutation GJB2 235delC and 12S rRNA homogeneous mutation, 1555 A> G, one heterozygous mutations at GJB2, 299delAT, and GJB3, 538C> T, one at GJB2, 299delAT and 12S rRNA, 1555 A> G, two at GJB2, 299delAT, and SLC26A4, IVS7-2A> G. All mutations as above were confirmed by DNA sequencing.

CONCLUSIONThe total mutation carrier rate of the 4 deafness genes is 3.19% in healthy newborns at Chengdu. Mutations of GJB2 and SLAC26A4 are major ones (86.5% of total). The mutation rate of mitochondrial DNA 12S rRNA is 2.71‰, which may have deafness induced by aminoglycoside antibiotics. Newborn screening for mutation of genes related to hereditary deafness plays an important role in the early detection and proper management for neonatal deafness as well as genetic counseling for premarital, prenatal and postnatal diagnosis.

Asian Continental Ancestry Group ; genetics ; Base Sequence ; China ; Connexin 26 ; Connexins ; genetics ; DNA Mutational Analysis ; DNA, Mitochondrial ; chemistry ; genetics ; Deafness ; diagnosis ; ethnology ; genetics ; Dried Blood Spot Testing ; Genetic Predisposition to Disease ; ethnology ; genetics ; Genetic Testing ; methods ; Humans ; Infant, Newborn ; Membrane Transport Proteins ; genetics ; Microarray Analysis ; methods ; Mutation ; Neonatal Screening ; methods ; RNA, Ribosomal ; genetics

OBJECTIVETo delineate the characteristics of the clinical manifestation, pathology of skeletal muscle and gene mutations of mitochondrial encephalomyopathy with lactic acidosis and stroke-like episode (MELAS) in children.

METHODThe clinical manifestation, laboratorial data, brain images, muscle pathology and mitochondrial gene mutations were analyzed in 24 patients with MELAS who were diagnosed in Department of Pediatrics, Peking University First Hospital. Their prognosis was evaluated by following up.

RESULTSymptoms of central nervous system such as stroke-like episodes, vomiting, convulsion and headache were present in all the 24 cases. Nine cases had the symptoms of myopathy. Twenty cases had developmental delay. Short stature, being thin and hairy was very common in these cases. Serum lactate level increased in all the cases, pyruvate increased in 17 cases. Elevated CSF lactate was found in 2 cases. Magnetic resonance imaging (MRI) was performed on 24 cases, out of them 23 were abnormal. The lesions mainly involved cerebral lobes. Occipital lobe was the most common site of lesions. Computed tomography (CT) was performed on 13 cases, low density lesions were present in 10 cases, basal ganglia calcifications in 5 cases. Muscle biopsy was performed on 8 cases, ragged-red fibers (RRF) were found in 4/8 cases, and abnormal accumulation of mitochondria were found in 3/8 cases. The mtDNA gene mutational analysis showed A3243G mutation in these patients. The mutation rates varied from 11.6% to 75.0%. The same mutation were found in 4/5 mothers who had the genetic tests, and the mutation rates of the mothers varied from 15.0% to 23.6%. The clinical information of 11 cases was available through recent following up. Three cases died, the others had some degrees of mental retardation.

CONCLUSIONChildren with MELAS had various clinical manifestations. Central nervous system and skeletal muscle were usually involved. Short stature and hypertrichosis were common signs. The prognosis of this disease was disappointing. mtDNA A3243G was the most common mutation in MELAS. Fully understanding the characteristics of its clinical manifestation, laboratory tests, brain image, muscle pathology and molecular features can be helpful to the early diagnosis and treatment.

Acidosis, Lactic ; blood ; Adolescent ; Brain ; diagnostic imaging ; pathology ; Child ; Child, Preschool ; DNA Mutational Analysis ; DNA, Mitochondrial ; genetics ; Electroencephalography ; Female ; Follow-Up Studies ; Humans ; Infant ; MELAS Syndrome ; diagnosis ; genetics ; pathology ; Magnetic Resonance Imaging ; Male ; Muscle, Skeletal ; diagnostic imaging ; pathology ; Point Mutation ; Pyruvic Acid ; blood ; Stroke ; diagnostic imaging ; genetics ; pathology ; Syndrome ; Tomography, X-Ray Computed

OBJECTIVEMitochondrial disease is a group of energy metabolic disorders, characterized by involvement of multisystem with high energy requirements. Encephalomyopathies are common clinical findings of the mitochondrial diseases. However, mitochondrial cardiac damage is not rare. In this study, the clinical, biological, and genetic analyses were performed in three patients with mitochondrial cardiac damage, in order to understand the characteristics of mitochondrial diseases.

METHODThree girls presented with arrhythmia and cardiac enlargement from the age of 3, 4 and 8 years respectively. They were admitted into the Peking University First Hospital. Infection, autoimmune diseases, aminoacidopathies, organic acidurias, mitochondrial-fatty acid oxidation defects, and lysosomal storage disease were excluded by routine laboratory examinations and metabolic analysis for blood amino acids, acylcarnitines, urinary organic acids, and lysosome activity assay. Peripheral leukocytes mitochondrial respiratory chain enzyme I to V activities were measured by spectrophotometry. The entire sequence of the mitochondrial DNA was analyzed.

RESULTIn two patients (case 1 and case 3), hypertrophic cardiomyopathy and grade I to grade II of cardiac function were found. One patient (case 2) was diagnosed with arrhythmia and grade I of cardiac function. Increased creatine phosphokinase and creatine kinase isoenzyme MB were observed. Mitochondrial respiratory chain complex deficiencies were indentified in the three patients. Patient 1 had combined deficiencies of complex III and V. The activity of complex I+III was 18.7 nmol/(min·mg mitochondrial protein) (control 84.4 ± 28.5). The activity of complex V was 20.4 nmol/(min·mg mitochondrial protein) (control 103.7 ± 29.2). In her mitochondrial gene, A14693G on tRNA(Glu) and T16519C on D-loop were found. Patient 2 had an isolated complex I deficiency. The activity was 22.0 nmol/(min·mg mitochondrial protein) (control 44.0 ± 5.4). A16183C, T16189C and G15043A mutations on D-loop were found. Patient 3 had a combined deficiency of complex IV and V. The activity of complex IV was 21.0 nmol/(min·mg mitochondrial protein) (control 54.1 ± 12.3). The activity of complex V was 23.2 nmol/(min·mg mitochondrial protein) (control 103.7 ± 29.2). C253T and C16187T mutations on D-loop were detected. Haplotype analysis showed that three patients belong to H2a2a. Improvement was observed after the treatment with L-carnitine, coenzyme Q10, vitamin C and E. At present, the patients are 7, 5 and 8 years old. Although excise intolerance still persists, they had a good general condition with normal school life.

CONCLUSIONThe mitochondrial diseases with cardiac damage show cardiomyopathy, arrhythmia and exercise intolerance. Many kinds of mitochondrial respiratory chain deficiency were observed. A14693G in mitochondrial tRNA(Glu) gene is probably one of the causes in China.

Arrhythmias, Cardiac ; diagnosis ; genetics ; metabolism ; Biomarkers ; blood ; urine ; Cardiomyopathy, Hypertrophic ; diagnosis ; genetics ; metabolism ; Child ; Child, Preschool ; DNA Mutational Analysis ; DNA, Mitochondrial ; genetics ; Electron Transport Chain Complex Proteins ; deficiency ; genetics ; metabolism ; Female ; Humans ; Male ; Mitochondria, Heart ; enzymology ; pathology ; Mitochondrial Diseases ; diagnosis ; genetics ; metabolism ; Mutation