Disorder Of Mitochondrial Function Research Articles

A Two-Genome Portrayal of Mitochondrial Disorders: A Review with Clinical Presentations.

Disorders of mitochondrial function are responsible for many inherited neuromuscular and metabolic diseases. Their combination of high mortality, multi-systemic involvement, and economic burden cause devastating effects on patients and their families. Molecular diagnostic tools are becoming increasingly important in providing earlier diagnoses and guiding more precise therapeutic treatments for patients suffering from mitochondrial disorders. This review addresses fundamental molecular concepts relating to the pathogenesis of mitochondrial dysfunction and disorders. A series of short cases highlights the various clinical presentations, inheritance patterns, and pathogenic mutations in nuclear and mitochondrial genes that cause mitochondrial diseases. Graphical and tabular representations of the results are presented to guide the understanding of the important concepts related to mitochondrial molecular genetics and pathology. Emerging technology is incorporating preimplantation genetic testing for mtDNA disorders, while mitochondrial replacement shows promise in significantly decreasing the transfer of diseased mitochondrial DNA (mtDNA) to embryos. Medical professionals must maintain an in-depth understanding of the gene mutations and molecular mechanisms underlying mitochondrial disorders. Continued diagnostic advances and comprehensive management of patients with mitochondrial disorders are essential to achieve robust clinical impacts from comprehensive genomic testing. This is especially true when supported by non-genetic tests such as biochemical analysis, histochemical stains, and imaging studies. Such a multi-pronged investigation should improve the management of mitochondrial disorders by providing accurate and timely diagnoses to reduce disease burden and improve the lives of patients and their families.

Energy metabolism of the immune system : Consequences in chronic inflammation

Energy is the currency of life. The systemic and intracellular energy metabolism plays an essential role for the energy supply of the resting and activated immune system and this also applies to chronic inflammatory diseases. This presentation examines both components of the systemic and cellular energy metabolism in health and chronic inflammation. Aliterature search was conducted using PubMed, Embase and the Cochrane Library. The information is presented in the form of anarrative review. Achronically activated immune system acquires large amounts of energy-rich substrates that are lost for other functions of the body. In particular, the immune system and the brain are in competition. The consequences of this competition are many known diseases, such as fatigue, anxiety, depression, anorexia, sleep problems, sarcopenia, osteoporosis, insulin resistance, hypertension and others. The permanent change in the brain causes long-term alterations that stimulate disease sequelae even after disease remission. In the intracellular energy supply, chronic inflammation typically involves a conversion to glycolysis (to lactate, which has its own regulatory functions) and the pentose phosphate pathway in disorders of mitochondrial function. The chronic changes in immune cells of patients with rheumatoid arthritis (RA) lead to adisruption of the citric acid cycle (Krebs cycle). The hypoxic situation in the inflamed tissue stimulates many alterations. A differentiation is made between effector functions and regulatory functions of immune cells. Based on the energy changes mentioned, novel treatment suggestions can be made in addition to those already known in energy metabolism.

Tissue-specific differences in the assembly of mitochondrial Complex I are revealed by a novel ENU mutation in ECSIT.

Mitochondrial Complex I assembly (MCIA) is a multi-step process that necessitates the involvement of a variety of assembly factors and chaperones to ensure that the final active enzyme is correctly assembled. The role of the assembly factor evolutionarily conserved signalling intermediate in the toll (ECSIT) pathway was studied across various murine tissues to determine its role in this process and how this varied between tissues of varying energetic demands. We hypothesized that many of the known functions of ECSIT were unhindered by the introduction of an ENU-induced mutation, while its role in Complex I assembly was affected on a tissue-specific basis. Here, we describe a mutation in the MCIA factor ECSIT that reveals tissue-specific requirements for ECSIT in Complex I assembly. MCIA is a multi-step process dependent on assembly factors that organize and arrange the individual subunits, allowing for their incorporation into the complete enzyme complex. We have identified an ENU-induced mutation in ECSIT (N209I) that exhibits a profound effect on Complex I component expression and assembly in heart tissue, resulting in hypertrophic cardiomyopathy in the absence of other phenotypes. The dysfunction of Complex I appears to be cardiac specific, leading to a loss of mitochondrial output as measured by Seahorse extracellular flux and various biochemical assays in heart tissue, while mitochondria from other tissues were unaffected. These data suggest that the mechanisms underlying Complex I assembly and activity may have tissue-specific elements tailored to the specific demands of cells and tissues. Our data suggest that tissues with high-energy demands, such as the heart, may utilize assembly factors in different ways to low-energy tissues in order to improve mitochondrial output. These data have implications for the diagnosis and treatment of various disorders of mitochondrial function as well as cardiac hypertrophy with no identifiable underlying genetic cause.

3-MCPD exposure enhances ovarian fibrosis and reduces oocyte quality in mice

3-monochloro-1,2-propanediol (3-MCPD) is a food contaminant believed to be harmful to human health. Previous studies showed that 3-MCPD exerts toxic effects in multiple tissues, but whether 3-MCPD affects female reproductive function remained unknown. Here, using mouse gastric lavage models, we report that 3-MCPD exposure for four weeks affected body growth, decreased the ovary/body weight ratio, and increased atretic follicle numbers. Expression levels of follicular development-related factors decreased. Further studies found that ovaries from 3-MCPD exposed mice had activated the Transforming Growth Factor-β (TGF-β) signaling pathway and promoted ovarian fibrosis. Increased TNF-α, IL-1 and NF-κB expression also indicated the occurrence of ovarian inflammation. Exposure to 3-MCPD stimulated the caspase pathway and enhanced granulosa cell apoptosis. Consistent with disrupted ovarian homeostasis, 3-MCPD exposure interfered with mitochondrial function, generated more reactive oxygen species, increased ferrous ion and lipid peroxidation levels, and resulted in decreased oocyte development potential. Collectively, these findings indicated that 3-MCPD exposure induced ovarian inflammation and fibrosis, and caused disorders of mitochondrial function and ferrous ion homeostasis in oocytes, which consequently disturbed follicle maturation and reduced oocyte quality.

Regulatory roles of external cholesterol in human airway epithelial mitochondrial function through STARD3 signalling.

BackgroundHypercholesterolemia is found in patients with chronic lung inflammation, during which airway epithelial cells play important roles in maintenance of inflammatory responses to pathogens. The present study aims at molecular mechanisms by which cholesterol changes airway epithelial sensitivity in response to smoking.MethodsHuman bronchial epithelial cells (HBEs) were stimulated with cigarette smoke extract (CSE) and mice were exposed to CS/lipopolysaccharide (LPS) as models in vitro and in vivo. Severe COPD patients and healthy volunteers were also enrolled and the level of cholesterol in plasma was detected by metabolomics. Filipin III and elisa kits were used to stain free cholesterol. Mitochondrial function was detected by mitotracker green, mitotracker green, and Seahorse. Mitochondrial morphology was detected by high content screening and electron microscopy. The mRNA and protein levels of mitochondrial dynamics‐related proteins were detected by RT‐qPCR and Western blot,respectively. BODIPY 493/503 was used to stain lipid droplets. Lipidomics was used to detect intracellular lipid components. The mRNA level of interleukin (IL)‐6 and IL‐8 were detected by RT‐qPCR.ResultsWe found that the cholesterol overload was associated with chronic obstructive pulmonary disease (COPD) and airway epithelia‐driven inflammation, evidenced by hypercholesterolemia in patients with COPD and preclinical models, alteration of lipid metabolism‐associated genes in CSE‐induced airway epithelia and production of ILs. External cholesterol altered airway epithelial sensitivity of inflammation in response to CSE, through the regulation of STARD3‐MFN2 pathway, cholesterol re‐distribution, altered transport and accumulation of cholesterol, activities of lipid transport regulators and disorder of mitochondrial function and dynamics. MFN2 down‐regulation increased airway epithelial sensitivity and production of ILs after smoking, at least partially by injuring fatty acid oxidation and activating mTOR phosphorylation.ConclusionsOur data provide new insights for understanding molecular mechanisms of cholesterol‐altered airway epithelial inflammation and for developing diagnostic biomarkers and therapeutic targets to improve patient outcomes.

Metabolomics-Based Frailty Biomarkers in Older Chinese Adults.

Background/ObjectivesOwing to accelerated population aging, health in older adults is becoming increasingly important. Frailty can reflect the health status and disease risks of older adults; however, appropriate biomarkers for early screening of frailty have not been identified. Here, we applied metabolomics to identify frailty biomarkers and potential pathogenic mechanisms of frailty.MethodsSerum metabolic profiles from 25 frail and 49 non-frail (control) older adults were systematically investigated by liquid chromatography-mass spectrometry-based metabolomics.ResultsWe identified 349 metabolites of 46 classes, with four increased and seven decreased metabolites in frail older adults. Pearson correlation analysis identified 11 and 21 metabolites that were positively and negatively correlated with grip strength, and 7 and 76 metabolites that were positively and negatively correlated with gait speed, respectively. Pathway analysis identified 10 metabolite sets and 13 pathways significantly associated with one or more frailty phenotype criteria.ConclusionThese results revealed the metabolite characteristics of serum in frail older adults. Intermediates of carbohydrate metabolism (e.g., isocitrate, malate, fumarate, cis-aconitate, glucuronate, and pyruvate), saturated fatty acids (e.g., palmitic acid), unsaturated fatty acids (e.g., arachidonate and linoleic acid), and certain essential amino acids (e.g., tryptophan) may be candidate biomarkers for the early diagnosis of frailty. Mitochondrial function disorders, saturated fatty acid-mediated lipotoxicity, aberrant unsaturated fatty acid metabolism, and increased tryptophan degradation could be potential mechanisms of frailty.

PCSK9 mediates the oxidative low‑density lipoprotein‑induced pyroptosis of vascularendothelialcells via the UQCRC1/ROS pathway.

The present study aimed to explore the role and mechanisms of proprotein convertase subtilisin/kexin type 9 (PCSK9) in the oxidized low-density lipoprotein (oxLDL)-induced pyroptosis of vascular endothelial cells. For this purpose, human umbilical vein endothelial cells (HUVECs) were incubated with oxLDL (100 µg/ml) for 24 h to induce pyroptosis, which was detected using PI/hoechst33342 double staining. The expression of pyroptosis-associated molecules was measured by western blot analysis and RT-qPCR. Reactive oxygen species (ROS) and membrane potential were examined through ROS probe and JC-1 staining, respectively. PCSK9 and mitochondrial ubiquinol-cytochrome c reductase core protein 1 (UQCRC1) protein were knocked down by small interfering RNA (siRNA). PCSK9 was overexpressed by lentivirus. The results revealed that oxLDL induced HUVEC injury, pyroptosis and inflammatory factor release, and upregulated the expression of PCSK9 protein in the HUVECs in a concentration-dependent manner. The silencing of PCSK9 expression with siRNA suppressed the oxLDL-induced damage to HUVECs, the release of inflammatory substances and the occurrence of pyroptosis. In addition, oxLDL inhibited UQCRC1 expression, promoted mitochondrial membrane potential collapse and damaged mitochondrial function; however, these processes were reversed by the silencing of PCSK9. PCSK9 overexpression induced the pyroptosis of HUVECs, the generation of ROS and the disorder of mitochondrial function by inhibiting UQCRC1. Therefore, PCSK9 mediates the oxLDL-induced pyroptosis of vascular endothelial cells via the UQCRC1/ROS pathway.

The role of mitochondria in the ischemic-reperfusion damage of the brain in operations on aortal arch in conditions of experiment

Objective. To investigate in experiment a functional state of mitochondria from the brain cells after its acute ischemic injury and processes, which take place in the brain in conditions of experimental ischemia/reperfusion; to analyze a correlation of the states investigated with the ischemic-reperfusion changes in the brain of patients, having the aortal arch aneurism, operated in environment of hypothermia and artificial cerebral perfusion.
 Materials and metods. First stage of the investigation consisted of estimation of the cerebral cells mitochondria state in conditions of ischemia/reperfusion of the neurons primary culture, obtained from embryos of rats of the 17 - 18 gestation days, using mechanical and the enzyme dissociation in the brain cortex. The second stage of the investigation was conducted on 14 white male rats (Rattus norvegicus). In 7 animals ischemia/reperfusion of the brain was simulated, using temporary mechanical blockade of blood flow along vessels of aortal arch with its subsequent restoration, as following: thoracotomy was performed along lower edge of the third rib, making surgical access to aortal arch and main vessels with their mechanical occlusion during 30 min. Additionally in 7 experimental animals thoracotomy only without impact on cerebral blood flow was performed. After extraction of animals from the experiment the blood was collected from the heart cavities, the brain was extirpated from the skull in accordance to standard method. After finishing of the experiment MTT-test was accomplished for estimation of vital capacity of the brain neurons. The principal parameters of the mitochondria functioning and velocity of their respiration in States II and III were estimated in accordance to Chance and in a respiratory control - a ratio of respiration velocity in a State III towards respiration velocity in a State IV in accordance to Chance. Transmembranous mitochondrial potential in cellular suspension, consisting of cells of the brain. Activity of lactate dehydrogenase in the culture environment was determined. Immunohistochemical investigation of the neurons culture was conducted. Membranous mitochondrial potential was estimated in the neurons cultivated. Respiration of isolated mitochondria was measured. Energetic state of mitochondria was determined. The development degree of apoptotic processes was determined in accordance to the DNA fragmentation degree.
 Results. There was established, that index of respiratory control of mitochondria from the ischemized brain, measured in pH 7.4, was lowered more than by 40%, comparing with corresponding index of mitochondria, obtained from a control culture. Velocity of a completely stimulated respiration of mitochondria (State III), which were extracted from the brain in 24 h after ischemia/reperfusion, was significantly lower, than velocity of respiration in mitochondria of control cultures, witnessing presence of damage in respiratory chain and transition of electrons of cerebral cellular mitochondria in ischemia/reperfusion, as well as incapacity of such mitochondria to transit electrons effectively from oxidation substrate towards oxygen due to damage of some complexes of the electron-transporting chain. Thus, in conditions of ischemia several processes, impacting mitochondrial functioning and cellular bioenergetics, occur simultaneously. Due to simultaneous action of these processes the mitochondrial functioning fails so significantly, that becomes one of the key moments in cerebral cellular dysfunction and death. Statistically meaningful difference, taking into account the results, between two groups of investigated animals was not revealed.
 Conclusion. In environment of ischemia/reperfusion of the brain in its cells a significant disorders of mitochondrial function occur, which are demonstrated by raising of generation of the oxygen active forms, and by lowering of transmembranous mitochondrial potential and velocity of mitochondrial respiration by 40%, what is accompanied by development of lactate-acidosis. It is expedient to apply a mitochondria-directed antioxidants in complex of the treatment measures in the brain ischemia/reperfusion, as well as desintegrators of respiration and oxidated phosphorilation.

Mitochondrial dysfunction and endoplasmic reticulum stress involved in oocyte aging: an analysis using single-cell RNA-sequencing of mouse oocytes

ObjectTo explore the mechanisms of ovarian aging, we performed overall analysis on the age-related alterations of gene expression profiles in mouse germinal vesicle (GV) stage oocytes by means of single-cell RNA-sequencing method (scRNA-seq).MethodsTwo age groups (5-week-old and 32-week-old) female KM mice were used as young and old models. Subsequently, GV oocytes were collected for scRNA-seq. The bioinformatics was performed to analyze and compare the differences of gene expression profile between GV oocytes of young and old mice.ResultsThe analysis of scRNA-seq data showed that there were 624 differential expressed genes (DEGs) between two age groups of mouse GV stage oocytes. Four hundred forty-nine DEGs were up-regulated while 175 DEGs were down-regulated in the GV oocytes of the old group. KEGG pathway analysis revealed that the genes involved in mitochondrial function including oxidative phosphorylation and ATP production pathway were significantly down-regulated in GV oocytes of 32-week-old mice, especially the mitochondrial encoded NADH dehydrogenase (mt-Nd), including mt-Nd2, mt-Nd3, mt-Nd4, mt-Nd4L and mt-Nd5. Analysis of DEGs revealed that endoplasmic reticulum stress-related genes including AdipoR2, IRAK-1, RCAN1 and MsrB1 were significantly down-regulated in GV oocytes of 32-week-old mice. Also, analysis of DEGs demonstrated that anti-oxidation-related genes including Erbb3、Rcan1、Gsto2 and Msrb1 were significantly down-regulated in GV oocytes of old group.ConclusionThe disorder of mitochondrial function, endoplasmic reticulum stress and the reduced antioxidant capability might be involved in the progression of oocyte aging. Especially, the down regulation of mitochondrial encoded subunits of respiratory chain complexes might play critical roles in the relevant mechanisms.

Altered expression of mitochondrial DNA encoding ATP 6, 8 and encoding protein in energy metabolism during cold preservation and reperfusion injury in rat liver transplantation

To study the expression changes of mitochondrial (mt) DNA encoding ATP6, 8 of hepatocytes in rat liver transplantation with cold preservation-reperfusion and explore the relationship between the expression of F(0)F(1)-ATPase induced by changes of ATP 6, 8 genes. Orthotopic liver transplantation was performed in Wistar rats using the cuff technique as described by Kamada with modifications. Rats were randomly divided into four groups: group A (30 min cold preservation), group B (6 h cold preservation), group C (12 h cold preservation) and group D (sham operation). Samples were collected at 12 h, 24 h and on Day 3, 5 and 7 post-operation. The levels of mtDNA encoding ATPase 6, ATPase 8 mRNA were determined by RT-PCR. The expression changes of F(0)F(1)-ATPase protein were examined by Western blot. The ATP level of liver was observed in each group. The expression of ATPase 6, ATPase8 mRNA, F(0)F(1)-ATPase protein expression (ng/mg protein) and the level of ATP decreased (U/L) in groups A, B and C at 12 h post-operation. The lowest was in group C (0.81 +/- 0.08, 0.71 +/- 0.07, 0.47 +/- 0.07, 0.44 +/- 0.05) when compared with groups A (1.11 +/- 0.04, 1.07 +/- 0.07, 0.86 +/- 0.15, 0.70 +/- 0.11) and B (1.02 +/- 0.07, 0.90 +/- 0.10, 0.65 +/- 0.07, 0.55 +/- 0.08) (P < 0.05). The expression of ATPase 6, ATPase 8 mRNA increased after 24 h in groups A, B and C. The changes of F(0)F(1)-ATPase protein expression and ATP level were consistent with the expression of ATPase 6, ATPase 8 mRNA. Both ATPase 6 mRNA, ATPase 8 mRNA and F(0)F(1)-ATPase protein in group C increased slowly compared with groups A and B after 24 h. The mtDNA encoding ATPase 6, ATPase 8 changes of hepatocytes in rats liver transplantation with cold preservation-reperfusion suggested that mitochondrial function disorders are related to the changes of mitochondrial DNA encoding ATPase6, ATPase8. mtDNA encoding ATPase 6, ATPase 8 genes of hepatocytes decreased significantly and F(0)F(1)-ATPase protein expression decreased remarkably and resulted in the decrease of ATP levels. The energy deficiency led to liver injury during cold preservation-reperfusion in rats liver transplantation. The injury of liver was more obvious with the prolongation of cold preservation period. The abnormal expressions of mtDNA ATPase 6, ATPase 8 genes are important causes of the changes of F(0)F(1)-ATPase protein expression resulting in a disorder of mitochondrial energy metabolism.

Presentation and Diagnosis of Mitochondrial Disorders in Children

The first disorder of mitochondrial function was described by Luft in 1959. Over the ensuing decades, multiple cases of mitochondrial dysfunction were reported, and the term "mitochondrial disorder" arose to describe any defect in the mitochondrial electron transport chain. The sequence of the mitochondrial genome was elucidated in 1981 by Anderson et al., and during the next 20 years, >200 pathogenic point mutations, deletions, insertions, and rearrangements were described. Most of the original cases were adults, and the diagnosis of a mitochondrial disorder in an adult patient became relatively straightforward. Adults present with well-defined "mitochondrial syndromes" and generally carry mitochondrial DNA mutations that are easily identified. Children with mitochondrial disorders are much harder to define. Children are more likely to have a nuclear DNA mutation, whereas the "classic" syndromic findings tend to be absent. This review describes both the varying presentations of mitochondrial disorders and the common laboratory, imaging, and pathologic findings related to children.

Alpers syndrome: progressive neuronal degeneration of children with liver disease

Alpers syndrome was not clearly defined until the link between brain and liver disease was described. Alpers syndrome can now be clearly established as a disorder of oxidative metabolism related to mitochondrial dysfunction, and in most instances with an autosomal mode of inheritance. The symptoms and signs are discussed. The illness occurs in the first years of life with the sudden onset of intractable seizures associated with developmental delay, hypotonia, ataxia, cortical blindness, and hepatic failure, and death occurs within a short time. Treating the seizures with valproic acid can cause the rapid onset of liver failure and must be avoided. To establish a definite diagnosis, liver and muscle biopsies may be needed. The former shows bile duct proliferation with the evidence of cirrhosis, and the latter may support the involvement of the mitochondrial respiratory chain if there are ragged‐red fibres. Genetic studies can show an association with mitochondrial DNA depletion and mutations in the polymerase gene. Cytochrome c oxidase deficiency has been demonstrated in some patients. Useful diagnostic tests include liver function tests, lactic acid levels in the blood and cerebrospinal fluid, electroencephalograms, computed tomography, and magnetic resonance imaging. The differential diagnosis will be from other forms of neuronal degeneration and disorders of mitochondrial function. There is no specific treatment, which must await further research into causes.

Alpers syndrome: progressive neuronal degeneration of children with liver disease

Alpers syndrome was not clearly defined until the link between brain and liver disease was described. Alpers syndrome can now be clearly established as a disorder of oxidative metabolism related to mitochondrial dysfunction, and in most instances with an autosomal mode of inheritance. The symptoms and signs are discussed. The illness occurs in the first years of life with the sudden onset of intractable seizures associated with developmental delay, hypotonia, ataxia, cortical blindness, and hepatic failure, and death occurs within a short time. Treating the seizures with valproic acid can cause the rapid onset of liver failure and must be avoided. To establish a definite diagnosis, liver and muscle biopsies may be needed. The former shows bile duct proliferation with the evidence of cirrhosis, and the latter may support the involvement of the mitochondrial respiratory chain if there are ragged-red fibres. Genetic studies can show an association with mitochondrial DNA depletion and mutations in the polymerase gene. Cytochrome c oxidase deficiency has been demonstrated in some patients. Useful diagnostic tests include liver function tests, lactic acid levels in the blood and cerebrospinal fluid, electroencephalograms, computed tomography, and magnetic resonance imaging. The differential diagnosis will be from other forms of neuronal degeneration and disorders of mitochondrial function. There is no specific treatment, which must await further research into causes.

Mechanisms of Mitochondrial Disease and the Role of Exercise: A Symposium

Mechanisms of Mitochondrial Disease and the Role of Exercise: A Symposium

Targeting proteins to mitochondria using TAT

Disorders of mitochondrial function cause significant human disease and suffering. To date, correction of these mitochondria defects has depended on biochemical approaches and has not been achieved via gene therapies. Using previously described fusion proteins containing the transactivator of transcription (TAT) region from the HIV virus and green fluorescent protein (GFP), with and without a mitochondrial targeting sequence (MTS) from mitochondrial malate dehydrogenase (mMDH), we have investigated transduction across mitochondrial membranes. Both TAT-GFP and TAT-mMDH-GFP fusion proteins are protected from externally added protease when incubated with isolated mitochondria. Furthermore, both TAT fusion proteins rapidly enter cultured cells and transduce into mitochondria. However, the MTS allows processing of the fusion protein and is necessary for persistence in mitochondria over time. Neither degradation of import receptors nor disruption of the mitochondrial membrane potential or pH gradient inhibits protein transduction of either fusion protein. Furthermore, when injected into pregnant mice, TAT-mMDH-GFP is detectable throughout fetal and neonatal pups. These results indicate that TAT fusion proteins are able to traverse mitochondrial membranes through mechanisms that do not involve the regular import pathway, and that the addition of a MTS allows persistence of the fusion protein within mitochondria. TAT-MTS fusion proteins may represent a viable option as potential mitochondrial protein therapies.