Mitochondria serve as the energy provider and enable life activities, and they are the only organelles containing extra-chromosomal DNA. Each mitochondrion contains multiple copies of its genome, which is usually referred to as mitochondrial DNA (mtDNA). mtDNA encodes necessary electron transport chain complex subunits, as well as the essential RNAs for their translation within the organelle. Therefore, the precondition for intact mitochondrial function and cardiomyocyte survival is the integrity of mtDNA. Accumulating evidence suggests that the disruption of mtDNA integrity is involved in ischemia/reperfusion-induced mitochondrial dysfunction and cardiomyocyte injury. Here, we review the current opinions about the pathways of mtDNA integrity maintenance and discuss the role of mtDNA integrity in cardiomyocyte injury reacting to ischemia/reperfusion. We also discuss the mechanisms by which mtDNA mediates ischemia/reperfusion-induced cardiomyocyte injury, together with therapeutic strategies by targeting mtDNA.

Mutations that negatively impact mitochondrial function are highly prevalent in humans and lead to disorders with a wide spectrum of disease phenotypes, including deficiencies in immune cell development and/or function. Previous analyses of mice with a hepatocyte-specific cytochrome c oxidase (COX) deficiency revealed an unexpected peripheral blood leukopenia associated with splenic and thymic atrophy. Here, we use mice with a hepatocyte-specific deletion of the COX assembly factor Sco1 to show that metabolic defects extrinsic to the hematopoietic compartment lead to a pan-lymphopenia represented by severe losses in both B and T cells. We further demonstrate that immune defects in these mice are associated with the loss of bone marrow lymphoid progenitors common to both lineages and early signs of autoantibody-mediated autoimmunity. Our findings collectively identify hepatocyte dysfunction as a potential instigator of immunodeficiency in patients with congenital mitochondrial defects who suffer from chronic or recurrent infections.

Mammalian mitochondrial DNA (mtDNA) is replicated by DNA polymerase γ (POLγ), a heterotrimeric complex consisting of a catalytic POLγA subunit and two accessory POLγB subunits1. More than 300 mutations in POLG, the gene encoding the catalytic subunit, have been linked to severe, progressive conditions with high rates of morbidity and mortality, for which no treatment exists2. Here we report on the discovery and characterization of PZL-A, a first-in-class small-molecule activator of mtDNA synthesis that is capable of restoring function to the most common mutant variants of POLγ. PZL-A binds to an allosteric site at the interface between the catalytic POLγA subunit and the proximal POLγB subunit, a region that is unaffected by nearly all disease-causing mutations. The compound restores wild-type-like activity to mutant forms of POLγ in vitro and activates mtDNA synthesis in cells from paediatric patients with lethal POLG disease, thereby enhancing biogenesis of the oxidative phosphorylation machinery and cellular respiration. Our work demonstrates that a small molecule can restore function to mutant DNA polymerases, offering a promising avenue for treating POLG disorders and other severe conditions linked to depletion of mtDNA.

The house mouse, Mus musculus, is a widely used animal model in biomedical research, with classical laboratory strains (CLS) being the most frequently employed. However, the limited genetic variability in CLS hinders their applicability in evolutionary studies. Wild-derived strains (WDS), on the other hand, provide a suitable resource for such investigations. This study quantifies genetic and phenotypic data of 101 WDS representing 5 species, 3 subspecies, and 8 natural Y consomic strains and compares them with CLS. Genetic variability was estimated using whole mtDNA sequences, the Prdm9 gene, and copy number variation at two sex chromosome-linked genes. WDS exhibit a large natural variation with up to 2173 polymorphic sites in mitogenomes, whereas CLS display 92 sites. Moreover, while CLS have two Prdm9 alleles, WDS harbour 46 different alleles. Although CLS resemble M. m. domesticus and M. m. musculus WDS, they differ from them in 10 and 14 out of 16 phenotypic traits, respectively. The results suggest that WDS can be a useful tool in evolutionary and biomedical studies with great potential for medical applications.

Mitochondrial dysfunction has been associated with neurodegenerative diseases (NDDs). This study aimed to explore the association between blood mitochondrial DNA copy number (mtDNA-CN) and development of NDDs. This study was based on two-sample Mendelian randomization (MR) analysis. The genome wide association study (GWAS) data of NDDs including Alzheimer's disease (AD), amyotrophic lateral sclerosis (ALS), age-related macular degeneration (AMD), multiple sclerosis (MS), Parkinson's disease (PD), primary open-angle glaucoma (POAG), and vascular dementia (VD) was obtained from FinnGen consortium. Inverse-variance weighted (IVW) was applied as the primary approach for MR estimation. MR results revealed that blood mtDNA-CN exhibited a significant relationship with the incidence of AD (IVW-P = 0.011, odds ratio [OR] = 0.65) and AMD (IVW-P = 0.038, OR = 0.64). However, there was no significant association observed between blood mtDNA-CN and other NDDs (IVW-P > 0.05). Our findings supported the relationship between mitochondrial dysfunction and development of AD and AMD, and that blood mtDNA-CN may serve as a potential biomarker for the incidence of these two NDDs.

Prostate cancer (PCa) is a significant healthcare challenge, associated with considerable mortality and morbidity among men, particularly in developed countries. PCa mortality and morbidity are primarily related to its most advanced form, metastatic castration-resistant PCa (mCRPC), for which there is presently no cure. Therefore, novel therapeutic approaches to increase mCRPC survival are critically needed. Due to PCa tumor heterogeneity and a complex tumor microenvironment, the efficacy of single-target radiopharmaceuticals, such as the Food and Drug Administration-approved [177Lu]Lu-PSMA-617, is currently under reassessment. The design and development of PCa dual-target radiopharmaceuticals have garnered considerable attention, due to their benefits over single-target counterparts, namely increased therapeutic specificity and efficacy, as well as the ability to overcome the challenge of inconsistent tumor visualization caused by variable receptor expression across diverse lesions, thereby enabling more comprehensive imaging. Several PCa biomarkers are currently being investigated as potential targets for dual-target radiopharmaceuticals, including prostate-specific membrane antigen, gastrin-releasing peptide receptor, integrin αvβ3 receptor, fibroblast activation protein, sigma-1 receptor, as well as albumin, the radiosensitive cell nucleus, and mitochondria. This review explores recent advancements in heterobivalent metal-based radiopharmaceuticals for dual targeting in PCa, highlighting their significance in theranostic and personalized medicine.

Gene therapy approaches for mitochondrial DNA (mtDNA)-associated damage/diseases have thus far been limited, and despite advancements in single gene therapy for mtDNA mutations and progress in mitochondrial transplantation, no method exists for restoring the entire mtDNA molecule in a clinically translatable manner. Here, we present for the first time a strategy to deliver an exogenous, fully intact, and healthy mtDNA template into cells to correct endogenous mtDNA mutations and deletions, with the potential to be developed into an efficient pan-therapy for inherited and/or acquired mtDNA disorders. More specifically, the novel therapeutic nanoparticle complex used in our study was generated by combining a cell-penetrating peptide (CPP) with purified mtDNA, in conjunction with a mitochondrial targeting reagent. The generated nanoparticle complexes were found to be taken up by cells and localized to mitochondria, with exogenous mtDNA retention/maintenance, along with mitochondrial RNA and protein production, observed in mitochondria-depleted ARPE-19 cells at least 4 weeks following a single treatment. These data demonstrate the feasibility of restoring mtDNA in cells via a CPP carrier, with the therapeutic potential to correct mtDNA damage independent of the number of gene mutations found within the mtDNA.

Tartrazine (TRZ), a synthetic red azo dye derived from coal tar, is widely used as a food colorant in various food products, pharmaceuticals, and cosmetics. This study aims to investigate the impact of TRZ on the expression levels of DNA methyltransferases (DNMT1, DNMT3a, and DNMT3b) and histone deacetylases (HDAC5 and HDAC6). Additionally, we evaluate genomic DNA stability using the alkaline comet assay in three human cell lines: immortalized human keratinocyte (HaCaT), human hepatocellular carcinoma (HepG2), and human lung adenocarcinoma (A549). The research question focuses on whether TRZ exposure alters epigenetic regulation and DNA integrity, potentially implicating its role in carcinogenesis.

The selected human cell lines were exposed to different concentrations of TRZ (20 µM, 40 µM, and 80 µM), with DMBA serving as a positive control. After treatment, we quantified the expression levels of DNMT1, DNMT3a, DNMT3b, HDAC5, and HDAC6 using quantitative real-time PCR. Additionally, we assessed DNA fragmentation via the alkaline comet assay to determine the extent of DNA damage resulting from TRZ exposure.

Our findings indicate that TRZ significantly upregulates the expression of HDAC5, HDAC6, DNMT1, DNMT3a, and DNMT3b in comparison to the control group. Furthermore, TRZ exposure leads to a notable increase in DNA damage, as evidenced by elevated tail moments across all examined human cell lines.

These results suggest that TRZ may play a role in carcinogenesis and epigenetic modifications. The observed upregulation of DNMTs and HDACs, coupled with increased DNA damage, highlights the potential risks associated with TRZ exposure. Further research is necessary to explore these mechanisms and assess their implications for human health.

Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled precise base substitutions and the efficient elimination of genomes carrying pathogenic mutations. However, reconstituting mtDNA deletions linked to mitochondrial myopathies remains challenging. Here, we engineered mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. Using mitochondrial EJ (mito-EJ) and mito-ScaI, we generated a panel of clonal cell lines harboring a ∼3.5 kb mtDNA deletion across the full spectrum of heteroplasmy. Investigating these cells revealed a critical threshold of ∼75% deleted genomes, beyond which oxidative phosphorylation (OXPHOS) protein depletion, metabolic disruption, and impaired growth in galactose-containing media were observed. Single-cell multiomic profiling identified two distinct nuclear gene deregulation responses: one triggered at the deletion threshold and another progressively responding to heteroplasmy. Ultimately, we show that our method enables the modeling of disease-associated mtDNA deletions across cell types and could inform the development of targeted therapies.

Selenocysteine (Sec), the 21st proteogenic amino acid, is a key component of selenoproteins, where it performs critical roles in redox reactions. Sec incorporation during translation is unique and highly sensitive to selenium levels. Encoded by the UGA codon, typically a termination signal, its insertion necessitates the presence of a selenocysteine insertion sequence (SECIS) within the 3' untranslated region (UTR) of selenoprotein mRNAs. This SECIS element orchestrates the recruitment of specialized molecular factors, including SECISBP2, the unique tRNA[Ser]Sec, and its dedicated elongation factor, EEFSEC. The extended variable arm of tRNA[Ser]Sec permits its specific recognition by EEFSEC. While the structure of the ribosome-bound complex is known, the precise mechanism by which EEFSEC-tRNA[Ser]Sec recodes UGA in the presence of SECIS and SECISBP2 remains unclear. tRNA[Ser]Sec has relatively few epitranscriptomic modifications, but those at the anticodon loop are crucial. Key modifications include N6-isopentenyladenosine (i6A) at position 37 and two forms of 5-methoxycarbonylmethyluridine (mcm5U and mcm5Um) at position 34. The ratio of these isoforms varies with tissue type and selenium levels, influencing mRNA-specific Sec recoding. A C65G mutation in the acceptor stem, identified in patients, disrupts these modifications at position 34, impairing selenoprotein synthesis. This highlights the essential role of wobble position modifications in anticodon function. tRNA[Ser]Sec exemplifies the complex regulation of UGA codon recoding and underscores the interplay of structural and epitranscriptomic factors in selenoprotein translation.

Ovarian aging has become a focal point in current research on female aging and refers to the gradual decline in ovarian function as women age. Numerous factors influence ovarian aging, among which mitochondrial function is one because it plays a crucial role by affecting oocytes and granulosa cells. Mitochondrial deterioration not only leads to a decrease in oocyte quality but also hinders follicle development, further impacting women's reproductive health and fertility.

This review summarizes and integrates research on the impact of mitochondrial function on ovarian aging, outlining the mechanisms by which mitochondria regulate the functions of oocytes and granulosa cells. This study aims to provide potential therapeutic directions to mitigate mitochondrial decline and support female reproductive health.

According to a 2023 study published in Cell, factors such as oxidative stress, mitochondrial dysfunction, chronic inflammation, and telomere shortening collectively drive ovarian aging, directly affecting female fertility. Among these factors, mitochondrial dysfunction plays a key role. This study reviewed literature from databases such as PubMed, Google Scholar, and CNKI, using keywords such as "mitochondrial dysfunction", "decline in oocyte quality and quantity", and "ovarian aging", aiming to summarize current research on the mechanisms of the impact of mitochondrial dysfunction on ovarian aging and provide theoretical support for future exploration of related therapeutic strategies.

The main characteristics of ovarian aging include a decline in oocyte quantity and quality, fluctuations in hormone levels, and a reduction in granulosa cell function. Studies have shown that mitochondria affect fertility by regulating cellular energy metabolism, exacerbating oxidative stress, causing mitochondrial DNA (mtDNA) damage, and impacting the physiological function of granulosa cells within the ovary, gradually diminishing the ovarian reserve.

This review focuses on analyzing the effects of mitochondrial decline on energy production in oocytes and granulosa cells, the accumulation of reactive oxygen species (ROS), and the calcium ion (Ca2+) concentration, which all contribute to the ovarian aging process, and understanding them will provide new insights into the mechanisms of ovarian aging.

Therapeutic interventions targeting mitochondrial dysfunction may help delay ovarian aging and improve female reproductive health.

The inherited genetic disorder β-thalassemia affects the hematopoietic system and is caused by the low production or absence of adult hemoglobin (HbA). Ineffective erythropoiesis is the hallmark of β-thalassemia pathophysiology and is characterized by an erythropoietin-driven substantial increase in erythroblast proliferation, coupled with an increase in late-stage precursor apoptosis, which results in low levels of circulating mature red blood cells (RBCs) and chronic anemia. Mitochondrial dysfunction commonly occurs in these cells because of the increased demand for energy production and the need to manage abnormal hemoglobin chain synthesis. Moreover, several studies have highlighted the importance of gradual mitochondrial clearance for mature erythroid cell production. This review offers an overview of the mitochondrial role in essential cellular processes, particularly those crucial for maintaining RBC health and function. Additionally, recent evidence regarding the contribution of mitochondrial dysfunction to the pathophysiology and severity of β-thalassemia is discussed, along with updated insights into indirect mitochondria-targeting treatments, which present potential pharmacological targets.

Mitochondrial DNA (mtDNA) mutagenesis remains poorly understood despite its crucial role in disease, aging, and evolutionary tracing. In this study, we reconstructed a comprehensive 192-component mtDNA mutational spectrum for chordates by analyzing 118,397 synonymous mutations in the CytB gene across 1,697 species and five classes. This analysis revealed three primary forces shaping mtDNA mutagenesis: (i) symmetrical, replication-driven errors by mitochondrial polymerase (POLG), resulting in C > T and A > G mutations that are highly conserved across classes; (ii) asymmetrical, damage-driven C > T mutations on the single-stranded heavy strand with clock-like dynamics; and (iii) asymmetrical A > G mutations on the heavy strand, with dynamics suggesting sensitivity to oxidative damage. The third component, sensitive to oxidative damage, positions mtDNA mutagenesis as a promising marker for metabolic and physiological processes across various classes, species, organisms, tissues, and cells. The deconvolution of the mutational spectra into mutational signatures uncovered deficiencies in both base excision repair (BER) and mismatch repair (MMR) pathways. Further analysis of mutation hotspots, abasic sites, and mutational asymmetries underscores the critical role of single-stranded DNA damage (components ii and iii), which, uncorrected due to BER and MMR deficiencies, contributes roughly as many mutations as POLG-induced errors (component i).

Mitochondria are organelles in most human cells which release the energy required for cells to function. Oxidative phosphorylation (OXPHOS) is a key biochemical process within mitochondria required for energy production and requires a range of proteins and protein complexes. Mitochondria contain multiple copies of their own genome (mtDNA), which codes for some of the proteins and ribonucleic acids required for mitochondrial function and assembly. Pathology arises from genetic defects in mtDNA and can reduce cellular abundance of OXPHOS proteins, affecting mitochondrial function. Due to the continuous turn-over of mtDNA, pathology is random and neighbouring cells can possess different OXPHOS protein abundance. Estimating the proportion of cells where OXPHOS protein abundance is too low to maintain normal function is critical to understanding disease severity and predicting disease progression. Currently, one method to classify single cells as being OXPHOS deficient is prevalent in the literature. The method compares a patient's OXPHOS protein abundance to that of a small number of healthy control subjects. If the patient's cell displays an abundance which differs from the abundance of the controls then it is deemed deficient. However, due to the natural variation between subjects and the low number of control subjects typically available, this method is inflexible and often results in a large proportion of patient cells being misclassified. These misclassifications have significant consequences for the clinical interpretation of these data. We propose a single-cell classification method using a Bayesian hierarchical mixture model, which allows for inter-subject OXPHOS protein abundance variation. The model accurately classifies an example dataset of OXPHOS protein abundances in skeletal muscle fibres (myofibres). When comparing the proposed and existing model classifications to manual classifications performed by experts, the proposed model results in estimates of the proportion of deficient myofibres that are consistent with expert manual classifications.

Human mitochondrial 12S ribosomal RNA (rRNA) 1555A>G mutation has been associated with aminoglycoside-induced and nonsyndromic deafness in many families worldwide. Our previous investigation revealed that the m.1555A>G mutation impaired mitochondrial translation and oxidative phosphorylation (OXPHOS). However, the mechanisms by which mitochondrial dysfunctions induced by m.1555A>G mutation regulate intracellular signaling for mitochondrial and cellular integrity remain poorly understood. Here, we demonstrated that the m.1555A>G mutation downregulated the expression of nucleus-encoded subunits of complexes I and IV but upregulated the expression of assemble factors for OXPHOS complexes, using cybrids derived from one hearing-impaired Chinese subject bearing the m.1555A>G mutation and from one hearing normal control lacking the mutation. These alterations resulted in the aberrant assembly, instability, and reduced activities of respiratory chain enzyme complexes I, IV, and V, rate of oxygen consumption, and diminished ATP production. Furthermore, the mutant cell lines carrying the m.1555A>G mutation exhibited decreased membrane potential and increased the production of reactive oxygen species. The aberrant assembly and biogenesis of OXPHOS impacted mitochondrial quality controls, including the imbalance of mitochondrial dynamics via increasing fission with abnormal mitochondrial morphology and impaired mitophagy. Strikingly, the cells bearing the m.1555A>G mutation revealed the upregulation of both ubiquitin-dependent and independent mitophagy pathways, evidenced by increasing levels of Parkin, Pink, BNIP3 and NIX, respectively. The m.1555A>G mutation-induced deficiencies ameliorate the cell homeostasis via elevating the autophagy process and upregulating apoptotic pathways. Our findings provide new insights into pathophysiology of mitochondrial deafness arising from reshaping mitochondrial and cellular homeostasis due to 12S rRNA 1555A>G mutation.

Genetic diseases constitute the majority of rare human diseases, resulting from abnormalities in an individual's genetic composition. Traditional treatments offer limited relief for these challenging conditions. In contrast, the rapid advancement of gene therapy presents significant advantages by directly addressing the underlying causes of genetic diseases, thereby providing the potential for precision treatment and the possibility of curing these disorders. This review aims to delineate the mechanisms and outcomes of current gene therapy approaches in clinical applications across various genetic diseases affecting different body systems. Additionally, genetic muscular disorders will be examined as a case study to investigate innovative strategies of novel therapeutic approaches, including gene replacement, gene suppression, gene supplementation, and gene editing, along with their associated advantages and limitations at both clinical and preclinical levels. Finally, this review emphasizes the existing challenges of gene therapy, such as vector packaging limitations, immunotoxicity, therapy specificity, and the subcellular localization and immunogenicity of therapeutic cargos, while discussing potential optimization directions for future research. Achieving delivery specificity, as well as long-term effectiveness and safety, will be crucial for the future development of gene therapies targeting genetic diseases.

Mitochondrial dysfunction is critical for the development and progression of cardiovascular diseases (CVDs). Complex-1 (CI) is an essential component of the mitochondrial electron transport chain that participates in oxidative phosphorylation and energy production. CI is the largest multisubunit complex (~ 1 Mda) and comprises 45 protein subunits encoded by seven mt-DNA genes and 38 nuclear genes. These subunits function as the enzyme nicotinamide adenine dinucleotide  hydrogen (NADH): ubiquinone oxidoreductase. CI dysregulation has been implicated in various CVDs, including heart failure, ischemic heart disease, pressure overload, hypertrophy, and cardiomyopathy. Several studies demonstrated that impaired CI function contributes to increased oxidative stress, altered calcium homeostasis, and mitochondrial DNA damage in cardiac cells, leading to cardiomyocyte dysfunction and apoptosis. CI dysfunction has been associated with endothelial dysfunction, inflammation, and vascular remodeling, critical processes in developing atherosclerosis and hypertension. Although CI is crucial in physiological and pathological conditions, no potential therapeutics targeting CI are available to treat CVDs. We believe that a lack of understanding of CI's precise mechanisms and contributions to CVDs limits the development of therapeutic strategies. In this review, we comprehensively analyze the role of CI in cardiovascular health and disease to shed light on its potential therapeutic target role in CVDs.

Mitochondrial genomic integrity is a key element of physiological processes and health. Changes in the half-life of the mitochondrial genome are implicated in the generation and accumulation of age-induced mitochondrial DNA (mtDNA) mutations, which are implicated in skeletal muscle aging and sarcopenia. There are conflicting data on the half-life of mtDNA, and there is limited information on how aging affects half-life in skeletal muscle. We hypothesized that skeletal muscle mtDNA synthesis rates would decrease with age in both female and male rats concomitant with changes in mtDNA integrity reflected in mtDNA copy number and mutation frequency. We measured mitochondrial genome half-life using stable isotope labeling over a period of 14 days and assessed mtDNA copy number and deletion mutation frequency using digital PCR in the quadriceps muscle of 9-month-old and 26-month-old male and female OKC-HET rats. We found a significant age-related increase in mtDNA half-life, from 132 days at 9 months to 216 days at 26 months of age in OKC-HET quadriceps. Concomitant with the increase in mtDNA half-life, we found an age-related increase in mtDNA deletion mutation frequency in both male and female rats. Notably, 26-month-old female rats had a lower mutation frequency than male rats, and there were no changes in mtDNA copy number with sex, age, or mitochondrial genotype. These data reveal several key findings: (1) mtDNA turnover in rat skeletal muscle decreases with age, (2) mtDNA half-lives in skeletal muscle are approximately an order of magnitude longer than what is reported for other tissues, and (3) muscle mtDNA turnover differs significantly from the turnover of other mitochondrial macromolecules including components of the mitochondrial nucleoid. These findings provide insight into the factors driving age-induced mtDNA mutation accumulation, which contribute to losses of mitochondrial genomic integrity and may play a role in skeletal muscle dysfunction.

Human mitochondrial DNA (mtDNA) mutations are causally implicated in maternally inherited mitochondrial respiratory disorders; however, the role of somatic mtDNA mutations in both late-onset chronic diseases and cancer remains less clear. Although recent advances in mtDNA base editing technology have the potential to model and characterize many of these mutations, current editing approaches are complicated by the potential for multiple unintentional edits (bystanders) that are only identifiable through empirical 'trial and error', thereby sacrificing valuable time and effort towards suboptimal construct development.

We developed MitoEdit, a novel tool that incorporates empirical base editor patterns to facilitate identification of optimal target windows and potential bystander edits. MitoEdit allows users to input DNA sequences in a text-based format, specifying the target base position and its desired modification. The program generates a list of candidate target windows with a predicted number of bystander edits and their functional impact, along with flanking nucleotide sequences designed to bind TALE (transcription activator-like effectors) array proteins. In silico evaluations indicate that MitoEdit can predict the majority of bystander edits, thereby reducing the number of constructs that need to be tested empirically. To the best of our knowledge, MitoEdit is the first tool to automate prediction of base edits.

MitoEdit is freely available at Kundu Lab GitHub ( https://github.com/Kundu-Lab/mitoedit ).

Corresponding email: Gang.Wu@stjude.org ; Mondira.Kundu@stjude.org.

Supplementary data are available at Bioinformatics online.

Mitochondrial genomic stability is critical to prevent various human inflammatory diseases. Bacterial infection significantly increases oxidative stress, driving mitochondrial genomic instability and initiating inflammatory human disease. Oxidative DNA base damage is predominantly repaired by base excision repair (BER) in the nucleus (nBER) as well as in the mitochondria (mtBER). In this review, we summarize the molecular mechanisms of spontaneous and H. pylori infection-associated oxidative mtDNA damage, mtDNA replication stress, and its impact on innate immune signaling. Additionally, we discuss how mutations located on mitochondria targeting sequence (MTS) of BER genes may contribute to mtDNA genome instability and innate immune signaling activation. Overall, the review summarizes evidence to understand the dynamics of mitochondria genome and the impact of mtBER in innate immune response during H. pylori-associated pathological outcomes.

Human mitochondrial DNA (mtDNA) harbors essential mutations linked to aging, neurodegenerative diseases, and complex muscle disorders. Due to its uniparental and haploid inheritance, mtDNA captures matrilineal evolutionary trajectories, playing a crucial role in population and medical genetics. However, critical questions about the genomic diversity patterns, inheritance models, and evolutionary and medical functions of mtDNA remain unresolved or underexplored, particularly in the transition from traditional genotyping to large-scale genomic analyses. This review summarizes recent advancements in data-driven genomic research and technological innovations that address these questions and clarify the biological impact of nuclear-mitochondrial segments (NUMTs) and mtDNA variants on human health, disease, and evolution. We propose a streamlined pipeline to comprehensively identify mtDNA and NUMT genomic diversity using advanced sequencing and computational technologies. Haplotype-resolved mtDNA sequencing and assembly can distinguish authentic mtDNA variants from NUMTs, reduce diagnostic inaccuracies, and provide clearer insights into heteroplasmy patterns and the authenticity of paternal inheritance. This review emphasizes the need for integrative multi-omics approaches and emerging long-read sequencing technologies to gain new insights into mutation mechanisms, the influence of heteroplasmy and paternal inheritance on mtDNA diversity and disease susceptibility, and the detailed functions of NUMTs.

While 5-fluorouracil (5FU) plays a central role in chemotherapy for colorectal cancer (CRC), resistance to 5FU remains a major challenge in CRC treatment, and its underlying mechanisms remain unclear. In this study, we investigated the relationship between 5FU resistance acquisition, stemness, and energy metabolism. Among the two CRC cell lines, HT29 cells exhibited glycolytic and quiescent properties, while CT26 cells relied on oxidative phosphorylation (OXPHOS) for energy. In contrast, the 5FU-resistant sublines (HT29R and CT26R), developed through continuous exposure to low concentrations of 5FU, demonstrated enhanced stemness. This was associated with glycolytic dominance, low proliferation, and reduced reactive oxygen species (ROS) production. However, treatment with the medium-chain fatty acid lauric acid shifted the cells to OXPHOS, reducing stemness, increasing ROS levels, and inducing cell death, therefore reversing 5FU resistance. These findings suggest that an enhancement in stemness and the reprogramming of energy metabolism play key roles in acquiring 5FU resistance in CRC. While lauric acid reversed 5FU resistance, further clinical studies are required.

Biosensing plays a vital role in healthcare monitoring, disease detection, and treatment planning. In recent years, nanofluidic technology has been increasingly explored to be developed into lab-on-a-chip biosensing systems. Given now the possibility of fabricating geometrically defined nanometric channels that are commensurate with the size of many biomolecules, nanofluidic-based devices are likely to become a key technology for the analysis of various clinical biomarkers, including DNA (deoxyribonucleic acid) and proteins in liquid biopsies. This review summarizes the fundamentals and technological advances of nanofluidics from the purview of single-molecule analysis, detection of low-abundance molecules, and single-cell analysis at the subcellular level. The extreme confinement and dominant surface charge effects in nanochannels provide unique advantages to nanofluidic devices for the manipulation and transport of target biomarkers. When coupled to a microfluidic network to facilitate sample introduction, integrated micro-nanofluidic biosensing devices are proving to be more sensitive and specific in molecular analysis compared to conventional assays in many cases. Based on recent progress in nanofluidics and current clinical trends, the review concludes with a discussion of near-term challenges and future directions for the development of nanofluidic-based biosensing systems toward enabling a new wave of lab-on-a-chip technology for personalized and preventive medicine.

Low leukocyte mitochondrial DNA (mtDNA) abundance has been associated with a higher risk of atherosclerotic cardiovascular disease, but through unclear mechanisms. We aimed to investigate whether low mtDNA abundance is associated with worse metabolomic profiling, as being potential intermediate phenotypes, using cross-sectional and genetic studies.

Among 61,186 unrelated European participants from UK Biobank, we performed multivariable-adjusted linear regression analyses to examine the associations between mtDNA abundance and 168 NMR-based circulating metabolomic measures and nine metabolomic principal components (PCs) that collectively covered 91.5% of the total variation of individual metabolomic measures. Subsequently, we conducted Mendelian randomization (MR) to approximate the causal effects of mtDNA abundance on the individual metabolomic measures and their metabolomic PCs.

After correction for multiple testing, low mtDNA abundance was associated with 130 metabolomic measures, predominantly lower concentrations of some amino acids and higher concentrations of lipids, lipoproteins and fatty acids; moreover, mtDNA abundance was associated with seven out of the nine metabolomic PCs. Using MR, genetically-predicted low mtDNA abundance was associated with lower lactate (standardized beta and 95% confidence interval: -0.17; -0.26, -0.08), and higher acetate (0.15; 0.07,0.23), and unsaturation degree (0.14; 0.08,0.20). Similarly, genetically-predicted low mtDNA abundance was associated with lower metabolomic PC2 (related to lower concentrations of lipids and fatty acids), and higher metabolomic PC9 (related to lower concentrations of glycolysis-related metabolites).

Low mtDNA abundance is associated with metabolomic perturbations, particularly reflecting a pro-atherogenic metabolomic profile, which potentially could link low mtDNA abundance to higher atherosclerosis risk.

Mitochondrial translation is essential to maintain mitochondrial function and energy production. Mutations in genes associated with mitochondrial translation cause several developmental disorders, and immune dysfunction is observed in many such patients. Besides genetic mutations, several antibiotics targeting bacterial ribosomes are well-established to inhibit mitochondrial translation. However, the effect of such antibiotics on different immune cells is not fully understood. Here, we addressed the differential effect of mitochondrial translation inhibition on different subsets of helper T cells (Th) of mice and humans. Inhibition of mitochondrial translation reduced the levels of mitochondrially encoded electron transport chain subunits without affecting their nuclear-encoded counterparts. As a result, mitochondrial oxygen consumption reduced dramatically, but mitochondrial mass was unaffected. Most importantly, we show that inhibition of mitochondrial translation induced apoptosis, specifically in Th2 cells. This increase in apoptosis was associated with higher expression of Bim and Puma, two activators of the intrinsic pathway of apoptosis. We propose that this difference in the sensitivity of Th1 and Th2 cells to mitochondrial translation inhibition reflects the intrinsic metabolic demands of these subtypes. Though Th1 and Th2 cells exhibit similar levels of oxidative phosphorylation, Th1 cells exhibit higher levels of aerobic glycolysis than Th2 cells. Moreover, Th1 cells are more sensitive to the inhibition of glycolysis, while higher concentrations of glycolysis inhibitor 2-deoxyglucose are required to induce cell death in the Th2 lineage. These observations reveal that selection of metabolic pathways for substrate utilization during differentiation of Th1 and Th2 lineages is a fundamental process conserved across species.

Retinitis pigmentosa (RP) is a progressive inherited retinal dystrophy, characterized by the degeneration of photoreceptors, presenting as a rod-cone dystrophy. Approximately 20-30% of patients with RP also exhibit extra-ocular manifestations in the context of a syndrome. This manuscript discusses the broad spectrum of syndromes associated with RP, pathogenic mechanisms, clinical manifestations, differential diagnoses, clinical management approaches, and future perspectives. Given the diverse clinical and genetic landscape of syndromic RP, the diagnosis may be challenging. However, an accurate and timely diagnosis is essential for optimal clinical management, prognostication, and potential treatment. Broadly, the syndromes associated with RP can be categorized into ciliopathies, inherited metabolic disorders, mitochondrial disorders, and miscellaneous syndromes. Among the ciliopathies associated with RP, Usher syndrome and Bardet-Biedl syndrome are the most well-known. Less common ciliopathies include Cohen syndrome, Joubert syndrome, cranioectodermal dysplasia, asphyxiating thoracic dystrophy, Mainzer-Saldino syndrome, and RHYNS syndrome. Several inherited metabolic disorders can present with RP including Zellweger spectrum disorders, adult Refsum disease, α-methylacyl-CoA racemase deficiency, certain mucopolysaccharidoses, ataxia with vitamin E deficiency, abetalipoproteinemia, several neuronal ceroid lipofuscinoses, mevalonic aciduria, PKAN/HARP syndrome, PHARC syndrome, and methylmalonic acidaemia with homocystinuria type cobalamin (cbl) C disease. Due to the mitochondria's essential role in supplying continuous energy to the retina, disruption of mitochondrial function can lead to RP, as seen in Kearns-Sayre syndrome, NARP syndrome, primary coenzyme Q10 deficiency, SSBP1-associated disease, and long chain 3-hydroxyacyl-CoA dehydrogenase deficiency. Lastly, Cockayne syndrome and PERCHING syndrome can present with RP, but they do not fit the abovementioned hierarchy and are thus categorized as 'Miscellaneous'. Several first-in-human clinical trials are underway or in preparation for some of these syndromic forms of RP.

Mitochondrial biogenesis requires the expression of genes encoded by both the nuclear and mitochondrial genomes. However, aside from a handful transcription factors regulating specific subsets of mitochondrial genes, the overall architecture of the transcriptional control of mitochondrial biogenesis remains to be elucidated. The mechanisms coordinating these two genomes are largely unknown. We performed a targeted RNAi screen in developing eyes with reduced mitochondrial DNA content, anticipating a synergistic disruption of tissue development due to impaired mitochondrial biogenesis and mitochondrial DNA (mtDNA) deficiency. Among 638 transcription factors annotated in the Drosophila genome, 77 were identified as potential regulators of mitochondrial biogenesis. Utilizing published ChIP-seq data of positive hits, we constructed a regulatory network revealing the logic of the transcription regulation of mitochondrial biogenesis. Multiple transcription factors in core layers had extensive connections, collectively governing the expression of nearly all mitochondrial genes, whereas factors sitting on the top layer may respond to cellular cues to modulate mitochondrial biogenesis through the underlying network. CG1603, a core component of the network, was found to be indispensable for the expression of most nuclear mitochondrial genes, including those required for mtDNA maintenance and gene expression, thus coordinating nuclear genome and mtDNA activities in mitochondrial biogenesis. Additional genetic analyses validated YL-1, a transcription factor upstream of CG1603 in the network, as a regulator controlling CG1603 expression and mitochondrial biogenesis.

Increased reactive oxygen species (ROS) are involved in the pathological process of dry eye disease. Our previous results suggested that norepinephrine (NE) has a protective effect on dry eye.

This study explored the potential therapeutic role and underlying mechanisms of NE in benzalkonium chloride (BAC)-induced dry eye disease.

BAC-pretreated human corneal epithelial cells (HCEpiC) were cultured with various concentrations of NE. A BAC-induced dry eye mice model was established to explore the role of NE. Alterations in mice corneal tissues, ROS levels, mitochondrial function, and mitophagy levels were analyzed.

In vitro, our results revealed that BAC-exposed HCEpiC led to mitochondrial malfunction, which involved excessive ROS production, decreased mitochondrial membrane potential (MMP), and promoted mitochondrial fragmentation through increased DRP1 and fission protein 1 (Fis1) expression and reduced mitofusin 2 (Mfn2) expression. Moreover, topical BAC application induced excessive mitophagy. These effects were reversed by NE. Additionally, the increased expression of LC3B, SQSTM1/p62, PINK1, and Parkin, which control mitophagy, in BAC-exposed HCEpiC was suppressed by NE. In BAC-induced C57BL/6J mice, NE resulted in lower fluorescein staining scores, decreased TUNEL-positive cells, and decreased mitochondrial fragmentation.

In conclusion, our findings showed that NE therapy prevented HCEpiC following BAC application by regulating mitochondrial quality control, which is controlled by PINK1/Parkin-dependent mitophagy. Our research suggests a potential targeted treatment for dry eye disease.

Mitochondrial DNA encodes essential components of the respiratory chain complexes, serving as the foundation of mitochondrial respiratory function. Mutations in mtDNA primarily impair energy metabolism, exerting far-reaching effects on cellular physiology, particularly in the context of aging. The intrinsic vulnerability of mtDNA is increasingly recognized as a key driver in the initiation of aging and the progression of its related diseases. In the field of aging research, it is critical to unravel the intricate mechanisms underpinning mtDNA mutations in living organisms and to elucidate the pathological consequences they trigger. Interestingly, certain effects, such as oxidative stress and apoptosis, may not universally accelerate aging as traditionally perceived. These phenomena demand deeper investigation and a more nuanced reinterpretation of current findings to address persistent scientific uncertainties. By synthesizing recent insights, this review seeks to clarify how pathogenic mtDNA mutations drive cellular senescence and systemic health deterioration, while also exploring the complex dynamics of mtDNA inheritance that may propagate these mutations. Such a comprehensive understanding could ultimately inform the development of innovative therapeutic strategies to counteract mitochondrial dysfunctions associated with aging.

Mitochondrial dysfunction is well recognized as a critical component of the complicated pathogenesis of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. This review investigates the influence of mitochondrial DNA single nucleotide polymorphisms on mitochondrial function, as well as their role in the onset and progression of these neurodegenerative diseases. Furthermore, the contemporary approaches to mitochondrial regulation in these disorders are discussed. Our objective is to uncover early diagnostic targets and formulate precision medicine strategies for neurodegenerative diseases, thereby offering new paths for preventing and treating these conditions.

Mitochondria, cellular powerhouses, harbor DNA (mtDNA) inherited from the mothers. MtDNA mutations can cause diseases, yet whether they increase with age in human germline cells-oocytes-remains understudied. Here, using highly accurate duplex sequencing of full-length mtDNA, we detected de novo mutations in single oocytes, blood, and saliva in women between 20 and 42 years of age. We found that, with age, mutations increased in blood and saliva but not in oocytes. In oocytes, mutations with high allele frequencies (≥1%) were less prevalent in coding than non-coding regions, whereas mutations with low allele frequencies (<1%) were more uniformly distributed along mtDNA, suggesting frequency-dependent purifying selection. In somatic tissues, mutations caused elevated amino acid changes in protein-coding regions, suggesting positive or destructive selection. Thus, mtDNA in human oocytes is protected against accumulation of mutations having functional consequences and with aging. These findings are particularly timely as humans tend to reproduce later in life.

The relationship between mitochondrial DNA (mtDNA) heteroplasmy and nuclear DNA (nDNA) methylation (CpGs) remains to be studied. We conducted an epigenome-wide association analysis of heteroplasmy burden scores across 10,986 participants (mean age 77, 63% women, and 54% non-White races/ethnicities) from seven population-based observational cohorts. We identified 412 CpGs (FDR p < 0.05) associated with mtDNA heteroplasmy. Higher levels of heteroplasmy burden were associated with lower nDNA methylation levels at most significant CpGs. Functional inference analyses of genes annotated to heteroplasmy-associated CpGs emphasized mitochondrial functions and showed enrichment in cardiometabolic conditions and traits. We developed CpG-scores based on heteroplasmy-count associated CpGs (MHC-CpG scores) using elastic net Cox regression in a training cohort. A one-unit higher level of the standardized MHC-CpG scores were associated with 1.26-fold higher hazard of all-cause mortality (95% CI: 1.14, 1.39) and 1.09-fold higher hazard of CVD (95% CI: 1.01-1.17) in the meta-analysis of testing cohorts, adjusting for age, sex, and smoking. These findings shed light on the relationship between mtDNA heteroplasmy and DNA methylation, and the role of heteroplasmy-associated CpGs as biomarkers in predicting all-cause mortality and cardiovascular disease.

Mitochondrial diseases are distinct types of metabolic and/or neurologic abnormalities that occur as a consequence of dysfunction in oxidative phosphorylation, affecting several systems in the body. There is no effective treatment modality for mitochondrial disorders so far, emphasizing the clinical significance of preventing the inheritance of these disorders. Various reproductive options are available to reduce the probability of inheriting mitochondrial disorders, including in vitro fertilization (IVF) using donated oocytes, preimplantation genetic testing (PGT), and prenatal diagnosis (PND), among which PGT not only makes it possible for families to have genetically-owned children but also PGT has the advantage that couples do not have to decide to terminate the pregnancy if a mutation is detected in the fetus. PGT for mitochondrial diseases originating from nuclear DNA includes analyzing the nuclear genome for the presence or absence of corresponding mutations. However, PGT for mitochondrial disorders arising from mutations in mitochondrial DNA (mtDNA) is more intricate, due to the specific characteristics of mtDNA such as multicopy nature, heteroplasmy phenomenon, and exclusive maternal inheritance. Therefore, the present review aims to discuss the utility and challenges of PGT as a preventive approach to inherited mitochondrial diseases caused by mtDNA mutations.

Mitochondrial DNA (mtDNA) variations affect the efficiency of the electron transport chain and production of reactive oxygen species, contributing to carcinogenesis. The D-loop region of mtDNA has emerged as a variation hotspot region in human neoplasia; however, the potential contribution of these variations in breast cancer risk prediction remains unknown. We investigated the relationship between germline single nucleotide polymorphisms (SNPs) in the entire D-loop region and breast cancer risk in Chinese women. Peripheral blood-isolated mtDNA from 2329 patients with breast cancer and 2328 cancer-free controls was examined for SNPs. In the combined cohort, we used traditional risk factors, susceptibility germline polymorphisms, and logistic regression analysis to evaluate the predictive value of susceptibility variants for breast cancer risk. We calculated the area under the receiver operating characteristic curve (AUC) as a measure. We also measured the content of 8-hydroxy-2'-deoxyguanosine (8-OHdG). Individual polymorphisms SNP573 were significantly associated with breast cancer risk in both the discovery and validation cohorts. In the combined cohort, the AUC of the traditional risk factors was 64.3%; after adding susceptibility variants, the AUC was 64.9% (DeLong test, p = 0.007). 8-OHdG levels were significantly higher in patients with breast cancer than in controls and higher in individuals with SNP573 than in those negative for this variation. Overall, oxidative stress might be associated with the risk of breast cancer, and SNP573 might be associated with oxidative stress. Our results indicate the risk potential of polymorphisms in the D-loop region in breast cancer in Southern China.

This narrative review summarizes current literature on the relationship of mitochondrial biomarkers with obesity-related characteristics, including body mass index and body composition.

Mitochondria, as cellular powerhouses, play a pivotal role in energy production and the regulation of metabolic process. Altered mitochondrial functions contribute to obesity, yet evidence of the intricate relationship between mitochondrial dynamics and obesity-related outcomes in human population studies is scarce and warrants further attention. We discuss emerging evidence linking obesity and related health outcomes to impaired oxidative phosphorylation pathways, oxidative stress and mtDNA variants, copy number and methylation, all hallmark of suboptimal mitochondrial function. We also explore the influence of dietary interventions and metabolic and bariatric surgery procedures on restoring mitochondrial attributes of individuals with obesity. Finally, we report on the potential knowledge gaps in the mitochondrial dynamics for human health for future study.

Mitochondria, pivotal to cellular metabolism, serve as the primary sources of biological energy and are key regulators of intracellular calcium ion storage, crucial for maintaining cellular calcium homeostasis. Dysfunction in these organelles impairs ATP synthesis, diminishing cellular functionality. Emerging evidence implicates mitochondrial dysfunction in the etiology and progression of diverse diseases. Environmental factors that induce mitochondrial dysregulation raise significant public health concerns, necessitating a nuanced comprehension and classification of mitochondrial-related hazards. This review systematically adopts a toxicological perspective to illuminate the biological functions of mitochondria, offering a comprehensive exploration of how toxicants instigate mitochondrial dysfunction. It delves into the disruption of energy metabolism, the initiation of mitochondrial fragility and autophagy, and the induction of mutations in mitochondrial DNA by mutagens. The overarching objective is to enhance our understanding of the repercussions of mitochondrial damage on human health.

Primary liver cancer (PLC), which includes hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (iCCA), remains a leading cause of cancer-related death worldwide. Chronic liver diseases, such as hepatitis B and C infections and metabolic dysfunction-associated steatotic liver disease (MASLD), are key risk factors for PLC. Metabolic reprogramming, a defining feature of cancer, enables liver cancer cells to adapt to the demands of rapid proliferation and the challenging tumor microenvironment (TME). This manuscript examines the pivotal role of metabolic reprogramming in PLC, with an emphasis on the alterations in glucose, lipid, and amino acid metabolism that drive tumor progression. The Warburg effect, marked by increased glycolysis, facilitates rapid energy production and biosynthesis of cellular components in HCC. Changes in lipid metabolism, including elevated de novo fatty acid synthesis and lipid oxidation, support membrane formation and energy storage essential for cancer cell survival. Amino acid metabolism, particularly glutamine utilization, supplies critical carbon and nitrogen for nucleotide synthesis and maintains redox homeostasis. These metabolic adaptations not only enhance tumor growth and invasion but also reshape the TME, promoting immune escape. Targeting these metabolic pathways presents promising therapeutic opportunities for PLC. This review underscores the interaction between metabolic reprogramming and tumor immunity, suggesting potential metabolic targets for innovative therapeutic strategies. A comprehensive understanding of PLC's intricate metabolic landscape may lead to more effective treatments and better patient outcomes. Integrating metabolomics, genomics, and proteomics in future research will be vital for identifying precise therapeutic targets and advancing personalized therapies for liver cancer.

The inference of body fluids and tissues is critical in reconstructing crime scenes and inferring criminal behaviors. Nevertheless, present methods are incompatible with conventional DNA genotyping, and additional testing might result in excessive consumption of forensic scene materials. This study aims to investigate the feasibility of distinguishing common body fluids/tissues through the difference in mitochondrial DNA copy number (mtDNAcn). Four types of body fluids/tissues were analyzed in this study - hair, saliva, semen, and skeletal muscle. MtDNAcn was estimated by dividing the read counts of mitochondrial DNA to that of nuclear DNA (RRmt/nu). Results indicated that there were significant differences in RRmt/nu between different body fluids/tissues. Specifically, hair samples exhibited the highest RRmt/nu (log10RRmt/nu: 4.3 ± 0.28), while semen samples showed the lowest RRmt/nu (log10RRmt/nu: -0.1 ± 0.28). RRmt/nu values for DNA samples without extraction were notably higher (approximately 2.9 times) than those obtained after extraction. However, no significant difference in RRmt/nu was observed between various age and gender groups. Hierarchical clustering and Kmeans clustering analyses showed that body fluids/tissues of the same type clustered closely to each other and could be inferred with high accuracy. In conclusion, this study demonstrated that the simultaneous detection of nuclear and mitochondrial DNA made it possible to perform conventional DNA analyses and body fluid/tissue inference at the same time, thus killing two birds with one stone. Furthermore, mtDNAcn has the potential to serve as a novel and promising biomarker for the identification of body fluids/tissues.