Bisphenol A (BPA), a synthetic endocrine-disrupting chemical, has been implicated in several diseases, including cancer. However, its effects on mitochondrial function in cervical cancer (CC) remain poorly understood. We investigated the effects of BPA on mitochondrial structure and function in SiHa and CaSki CC cell lines. Confocal microscopy revealed that BPA (1 nM-1 µM) disrupted mitochondrial morphology, increased circularity (*p < 0.05), and reduced branch length and content. BPA increased mitochondrial ROS (mtROS) and Ca2⁺ levels, decreased mitochondrial membrane potential (MMP) and mitochondrial mass, induced mtDNA depletion, and impaired oxidative respiration (OCR). Notably, BPA increased glucose uptake and lipid peroxidation while reducing the antioxidant (GSH and CAT) activity. A dose-dependent increase in mitochondrial lipid droplets (p < 0.001) was accompanied by opposite ATP trends: ATP levels increased in SiHa cells but decreased in CaSki cells. Western blot and qRT‒PCR revealed the cell type-specific regulation of PGC1α, TFAM, and DRP1. Molecular docking revealed strong binding of BPA to IDH2 (docking score - 7.38), MAOB (- 6.29), and HK2 (- 3.208), which was further validated by stable MD simulation. Thus, BPA may induce mitochondrial dysfunction, metabolic changes, and oxidative stress in CC cells in vitro, suggesting a potential role in cervical carcinogenesis that warrants further in vivo investigation.
Mitochondrial transport and distribution are crucial for cellular homeostasis, yet whether and how they are regulated by endoplasmic reticulum (ER)-mitochondria contact sites remains unclear. Here, we demonstrate that the ER protein atlastin-2 (ATL2) orchestrates mitochondrial transport and distribution by promoting assembly of the transport machinery at ER-mitochondria contact sites. Mechanistically, ATL2 recruits the adaptor trafficking kinesin-binding protein 1 (TRAK1) to the ER membrane, strengthening the interaction of TRAK1 with the mitochondrial transport adaptor MIRO1 to promote anterograde mitochondrial transport. Loss of ATL2 disrupts this process, leading to perinuclear mitochondrial clustering. We further find that ATL2 stabilizes ER-mitochondria contact sites by interacting with MFN2, providing a platform for mitochondrial transport complex assembly. Moreover, in hypoxia, ATL2 is ubiquitinated at lysine 567 by the E3 ligase SYVN1, leading to its degradation and a resulting defect in mitochondrial distribution. Our findings elucidate a novel ER-mediated mechanism for mitochondrial transport.
Exercise and fasting are recognized for their ability to improve brain health and mitigate neurodegeneration. However, little is known about how these interventions acutely impact mitochondrial quality control mechanisms including mitophagy. We examined the effects of a single bout of fasting and exercise (FEx) on hippocampal mitochondrial function and proteomic remodeling in male and female mice. To assess in vivo autophagy dynamics, we combined proteomics with chloroquine (CQ) inhibition of autophagic flux. Mice were assigned to sedentary (Sed), fasting (F), exercise (Ex), or combined FEx groups and received unilateral intrahippocampal injections of CQ or PBS following treatments. Four hours later, hippocampi were collected for analysis. LC3-II levels significantly increased in the FEx group only following CQ treatment, indicating enhanced autophagic flux. Proteomic profiling showed sedentary males failed to mount a robust response to FEx however females exhibited upregulation of proteins involved in the TCA cycle, glutathione metabolism, and oxidative phosphorylation, suggesting greater mitochondrial adaptability. Functional assays supported these findings, females showed increased complex IV activity post-FEx. The mitochondrial DNA / nuclear DNA ratio increased after FEx regardless of sex, and upstream regulator analysis predicted activation of mitochondrial biogenesis. Together, these data reveal sex-specific mitochondrial remodeling in response to acute fasting and exercise. Defining these normative responses is critical for understanding how mitochondrial adaptability shapes resilience or vulnerability to neurological challenges.
Pentavalent antimony [Sb(V)] contamination is an emerging threat in the human environment, yet its mechanistic basis for developmental toxicity remains poorly defined. This study investigates whether mitochondrial dysfunction mediates Sb(V)-induced developmental toxicity in human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and peripheral blood mononuclear cells (PBMCs), using an integrated suite of bioenergetic, imaging, ultrastructural, and proteomic endpoints. Cells exposed to Sb(V) at 0, 25, 50, 75, and 100 μmol/L for 72 h exhibited dose-dependent mitochondrial impairment. Seahorse extracellular flux analysis revealed significant reductions in basal oxygen consumption rate (p < 0.01 at ≥ 50 μmol/L), adenosine triphosphate (ATP)-linked respiration, and maximal respiratory capacity, alongside increased proton leak. Real-time ATP imaging with the FRET-based AT1.03 sensor showed progressive, compartment-specific energy depletion, most pronounced in cardiomyocytes (up to 33% reduction at 100 μmol/L; p < 0.01). Tetramethylrhodamine ethyl ester (TMRE) staining confirmed mitochondrial membrane depolarization, and MitoSOX staining demonstrated elevated mitochondrial superoxide generation. Electron transport chain (ETC) Complex I activity was reduced by 54.6% at 100 μmol/L (p < 0.01), with lesser but significant reductions in Complexes II, III, and IV. Transmission electron microscopy (TEM) revealed cristae disorganization, matrix swelling, and outer membrane disruption in a concentration-dependent pattern. Quantitative proteomics identified 286 differentially expressed proteins enriched in oxidative phosphorylation and tricarboxylic acid (TCA) cycle pathways. These mitochondrial perturbations correlated strongly with contractile dysfunction (Pearson r = 0.89-0.91, p < 0.001) and reduced cell viability/growth (r = 0.82-0.87, p < 0.001). The findings establish an adverse outcome pathway: Sb(V) exposure causes ETC inhibition, which drives ATP depletion and oxidative stress, leading to mitochondrial structural damage, cardiac cell dysfunction, and impaired cell growth. Mitochondrial endpoints such as basal oxygen consumption rate, ATP production capacity, and Complex I activity offer sensitive and quantifiable biomarkers for sublethal antimony monitoring in human health risk assessment, particularly for children in communities affected by mining, smelting, or antimony-contaminated drinking water.
Bcl-xL, an anti-apoptotic Bcl-2 family protein, engages laterally with Bak/Bax in the outer mitochondrial membrane (OMM) to inhibit apoptosis and interacts with the IP3 receptor Ca2+ channels (IP3Rs) in the endoplasmic reticulum (ER) membrane to control Ca2+ release. It is unknown if OMM-localized Bcl-xL can also interact in trans with IP3Rs at ER-mitochondrial contacts to form a tethering complex that supports IP3R-mediated local Ca2+ transfer from ER to mitochondria. We establish that IP3R-mitochondria Ca2+ signal propagation depends on Bcl-xL. By targeting Bcl-xL specifically to different subcellular compartments, we find that OMM-localized Bcl-xL increases the efficacy of ER-mitochondrial Ca2+ transfer without changing ER Ca2+ release, despite attenuating mitochondrial Ca2+ uptake. We find interaction between Bcl-xL and each IP3R isoform occurring at the mitochondria and a complex formed by OMM-localized Bcl-xL and IP3Rs. OMM Bcl-xL interacts with IP3Rs in trans at ER-mitochondrial contacts to optimize local Ca2+ signal propagation into the mitochondria.
Third-generation EGFR tyrosine kinase inhibitors (EGFR-TKIs), including osimertinib, show robust clinical efficacy in EGFR-mutant (EGFRm) non-small cell lung cancer (NSCLC), yet acquired resistance remains inevitable. Here, we demonstrate that osimertinib and other EGFR-TKIs suppress PPARGC1B expression and its regulated mitochondrial biogenesis in EGFRm NSCLC cells through a previously unrecognized FOSL1/AP-1-mediated transactivation mechanism. Upon acquisition of osimertinib resistance, PPARGC1B expression and its encoded protein PGC1β rebound and become refractory to osimertinib-mediated suppression. Enforced overexpression of PPARGC1B confers resistance to osimertinib in sensitive EGFRm NSCLC cells, whereas PPARGC1B knockdown restores drug sensitivity in resistant cells. Moreover, combining osimertinib with the mitochondria-targeting agent CPI-613 synergistically suppresses mitochondrial biogenesis, induces apoptosis, and inhibits the growth of osimertinib-resistant cells and tumors. Collectively, these findings identify PGC1β-dependent mitochondrial biogenesis as a critical determinant of therapeutic response to osimertinib and suggest co-targeting mitochondrial metabolism as a potential strategy to overcome acquired resistance in EGFRm NSCLC.
Myoferlin, a type 2 transmembrane protein in the ferlin family, is traditionally known for its role in membrane fusion during muscle development and repair. Recent research identifies myoferlin as a potential biomarker and a critical driver of cancer progression, particularly in breast cancer and pancreatic ductal adenocarcinoma. While its lack of specificity limits its use as a biomarker, its multifaceted role in cellular membrane dynamics makes it a promising therapeutic target. In cancer cells, myoferlin regulates the recycling and stability of receptor tyrosine kinases, thereby promoting invasion and metastasis. Beyond the plasma membrane, it maintains mitochondrial homeostasis by interacting with the machinery for mitochondrial fusion and calcium exchange at the endoplasmic reticulum-mitochondria interface. Depletion of myoferlin disrupts these processes, leading to mitochondrial fragmentation, reduced ATP production, and iron-dependent cell death. Furthermore, myoferlin influences the tumour microenvironment by regulating pancreatic cancer-associated fibroblasts. It interacts with SEC24 to facilitate the coat protein complex II-mediated transport of the transforming growth factor-beta 1 receptor, driving the desmoplastic reaction and matrix protein deposition. The 'one punch-two hits' strategy-simultaneously targeting the metabolic and signalling pathways of both malignant cells and the stroma-offers a novel therapeutic perspective. The development of small molecules targeting myoferlin's C2 domains confirms its potential to reduce tumour growth and metastatic dissemination.
The development and functional maintenance of CD8+ T cells are metabolically regulated processes in which mitochondria serve as the central hub. Here, we identify glucose-regulated protein 75 (GRP75) as a critical mitochondrial regulator controlling these processes. Using T cell-specific Hspa9 (encodes GRP75) knockout mice, we demonstrate that GRP75 deficiency disrupts CD8+ T cell fate, leading to defective T cell homeostasis and impaired memory differentiation. Mechanistically, impaired mitochondrial function in GRP75-deficient CD8+ T cells leads to perturbation of IL-7R signaling and aberrant expression of effector-associated molecules. Further studies reveal that GRP75 deficiency leads to upregulation of interferon regulatory factor 4 (IRF4), a critical transcription factor for effector versus memory fate, which in turn suppresses memory CD8+ T cell differentiation. Our findings establish GRP75 as a pivotal mitochondrial checkpoint that coordinates metabolic state and functional fate in CD8+ T cells.
The evolution of mitochondria provides crucial insights into the diversification of eukaryotes, with complex events of gene losses revealed through comparative analyses of mitochondrial genomes (mitogenomes) across eukaryotic lineages. However, the mitogenomes of many microbial eukaryotes remain underexplored due to challenges in their isolation and cultivation. Particularly understudied are Foraminifera (Rhizaria, SAR), unicellular calcifiers that are widely distributed across global oceans and important paleoenvironmental proxies. Using single-cell genomic sequencing, we report a 22-kb mitogenome from a planktonic foraminiferan in tropical seawater, the smallest known to date among sequenced mitogenomes of Rhizaria, a major lineage of eukaryotes. It contains only six protein-coding genes (including reduced versions of nad1 and cox2) and fragmented ribosomal RNA genes, and has lost most genes in oxidative phosphorylation and all genes encoding mitochondrial ribosomal proteins. Such genome reduction is associated with accelerated evolutionary rates and a lower GC content than that of benthic foraminiferan and other rhizarian mitogenomes. These findings highlight a unique trajectory of mitogenome reduction during rhizarian evolution and the use of single-cell approaches to recover microbial eukaryotic genomes and expand our understanding of mitochondrial evolution.
The chick chorioallantoic membrane (CAM) assay is an in vivo metastasis model, yet traditional Alu-qPCR quantification suffers from non-specific background noise in avian tissue. To improve analytical specificity, we developed a species-specific digital PCR (dPCR) assay targeting the human mitochondrial locus MT-ND4. While Alu-qPCR generated false positives in chicken-only DNA, MT-ND4 dPCR provided high species specificity and robust relative quantification of human mitochondrial DNA across various embryonic organs. As a proof of concept in five embryos, organ-dependent differences in metastatic burden were detected (p < 0.001) with high technical precision. This MT-ND4 dPCR assay avoids false-positive overestimations, providing a specific tool for quantifying human metastatic burden in the CAM model.
Cardiovascular diseases are the leading cause of death worldwide, with arterial calcification being a risk factor, especially in patients with diabetes and kidney disease. Arterial calcification involves hydroxyapatite deposition and the transformation of vascular smooth muscle cells (vSMCs) into osteoblast-like cells, processes mediated in part by tissue-nonspecific alkaline phosphatase (TNAP, encoded by ALPL), a key regulator of mineralization. Previous work showed increased activity of electron transport chain complexes I and IV during vSMC calcification. This study examined the role of mitochondrial complex I in vSMC calcification. vSMCs were calcified using osteogenic medium. Rotenone was used as a complex I inhibitor. Metabolomic profiling, extracellular pH measurements, and TNAP activity assays were performed. Rotenone dose-dependently reduced matrix mineralization, with near-complete inhibition at 50 nM, without affecting cell viability. Metabolomic analysis showed that rotenone increased both intracellular and extracellular lactic acid while decreasing pyruvic acid, indicating a shift toward glycolysis. Exogenous lactic acid reduced mineralization by 24%; sodium lactate reduced mineralization without altering extracellular pH; and extracellular acidification independently reduced mineralization by 23%, indicating that both lactate and proton-mediated acidification partially contribute to the anti-calcific mechanism. Additionally, rotenone decreased TNAP activity by 59% without affecting ALPL transcript levels. Inhibition of mitochondrial complex I causes metabolic reprogramming in calcifying vSMCs, promoting lactic acid accumulation that partially inhibits matrix mineralization through lactate-mediated and acidification-dependent mechanisms. The near-complete inhibition of calcification by rotenone suggests additional uncharacterized mechanisms beyond the lactic acid-pH axis contribute to its full anti-calcific effect, warranting further investigation.
Heme biosynthesis is tightly coordinated to support essential functions without accumulating toxic porphyrins and depleting cellular iron. Heme induces degradation of the heme biosynthetic enzyme, 5-aminolevulinate synthase (ALAS), by the mitochondrial caseinolytic protease complex CLPX-CLPP (CLPXP), but the mechanism for heme-triggered degradation had not been elucidated. We found that polymerase delta-interacting protein 2 (POLDIP2) is a heme-sensing adaptor protein sufficient to reconstitute negative feedback degradation of ALAS by CLPXP. POLDIP2 was necessary to support ALAS turnover in cells and regulate heme production during erythropoiesis. POLDIP2 directly recognized and recruited heme-bound ALAS to CLPXP. Degradation initiation required a carboxyl-terminal element of ALAS, truncations of which cause an erythropoietic protoporphyria. Our findings establish a mechanism for conditional degradation by CLPXP that underlies erythropoietic protoporphyrias linked to CLPX and ALAS.
暂无摘要（点击查看详情）
暂无摘要（点击查看详情）
Type 2 diabetes mellitus (T2DM) is increasingly recognized as a major risk factor for cognitive impairment and dementia, extending beyond its traditional metabolic complications. Emerging evidence suggests that cognitive decline in T2DM arises from a complex interplay among gut dysbiosis, chronic inflammation, impaired insulin signaling, blood-brain barrier dysfunction, and mitochondrial abnormalities. The gut-brain-mitochondria (GBM) axis has recently emerged as an integrated mechanistic framework linking peripheral metabolic disturbances to central neurodegenerative processes. Within this axis, alterations in gut microbial composition and function promote intestinal barrier disruption, endotoxemia, and aberrant production of microbial metabolites, which collectively contribute to systemic inflammation, neuroinflammation, and neuronal insulin resistance. Mitochondria occupy a central position in this network by regulating cellular bioenergetics, redox homeostasis, calcium signaling, and neuronal survival. Mitochondrial dysfunction amplifies oxidative stress, impairs synaptic plasticity, disrupts adult hippocampal neurogenesis, and accelerates neurodegenerative signaling pathways associated with cognitive decline. Furthermore, bidirectional communication between gut-derived metabolites and mitochondrial pathways influences neuronal resilience and disease progression. This review synthesizes current evidence describing the mechanistic convergence of gut dysbiosis, mitochondrial dysfunction, and neuroinflammation in T2DM-associated cognitive impairment. We also discuss emerging biomarkers, including mitochondrial DNA copy number, circulating lactate, lipopolysaccharide, zonulin, and microbial metabolite signatures, that may facilitate early disease detection and patient stratification. Finally, we critically evaluate therapeutic strategies targeting the GBM axis, including microbiota-directed interventions, mitochondrial-targeted antioxidants, metabolic modulators, and lifestyle-based approaches. Understanding the integrated role of the GBM axis may provide novel opportunities for precision-based interventions to prevent or slow diabetes-associated cognitive decline.
Total parenteral nutrition (TPN) is essential for patients who cannot tolerate enteral nutrition, but prolonged exposure to soybean oil-derived lipid emulsions (SOLEs) has been associated with parenteral nutrition-associated liver disease (PNALD). Sole contains phytosterols, including stigmasterol (ST), which may contribute to hepatocellular oxidative stress and apoptosis. This study investigated whether ST and SOLE promote oxidative injury, mitochondrial dysfunction, and apoptosis-related signaling in human L02 liver cells. Cells were exposed to ST, 0.5% SOLE, or their combination for 24 or 48 h. Cell viability, intracellular reactive oxygen species (ROS), mitochondrial membrane potential, apoptosis-related proteins, Nrf2/Keap1 signaling, and pathway-level molecular interactions were assessed using MTT and trypan blue assays, DCFH-DA fluorescence, JC-1 staining, Western blotting, immunofluorescence, and enrichment/network analyses. ST reduced L02 cell viability in a dose- and time-dependent manner and increased intracellular ROS accumulation. SOLE exposure further enhanced oxidative stress, whereas NAC partially attenuated ROS accumulation, supporting the contribution of oxidative stress to ST-induced cytotoxicity. ST also disrupted mitochondrial membrane potential and altered apoptosis-related markers, including caspase-3, PARP1, BAX/BCL2-associated signaling, and Nrf2/Keap1 stress-response regulation. Pathway enrichment and interaction network analyses further supported convergence of oxidative stress, mitochondrial dysfunction, DNA damage response, and apoptosis-related pathways. These findings provide mechanistic in vitro evidence that phytosterol-derived soybean oil emulsion can promote hepatocellular oxidative injury and mitochondrial apoptosis, supporting further investigation of phytosterol-reduced lipid formulations to improve hepatic safety during long-term PN.
Salidroside (SAL), the primary active compound extracted from Rhodiola rosea, has demonstrated potential efficacy against MASH-induced fibrosis; however, its precise pharmacological mechanisms and molecular targets remain incompletely elucidated. The aim of this study was to explore the protective effects of against MASH-induced liver fibrosis and its underlying pharmacological mechanisms. In this work, a MASH-induced fibrosis model was established using the high-fat diet (HFD) and choline-deficient, L-amino acid-defined (CDAHFD) diet-fed in vivo and palmitic acid/oleic acid-stimulated hepatocytes in vitro. By comprehensively conducting biochemical index measurements, pathological analysis, Western blot, PCR detection, RNA sequencing, 4D-DIA proteomic analyses, molecular docking, and validation experiments, the study revealed the critical association of PI3K/AKT/mTOR-mediated autophagy-associated mitochondrial quality control with the treatment of MASH-related liver fibrosis. Sal treatment significantly ameliorates hepatic steatosis, inflammatory infiltration, and liver fibrosis induced by CDAHFD and HFD. Integrated RNA sequencing and 4D-DIA proteomics analysis revealed that the anti-fibrotic effect of Sal on MASH-induced liver fibrosis may be associated with regulation of the PI3K/AKT/mTOR signaling pathway, thereby enhancing autophagy-associated mitochondrial quality control in hepatocytes, preventing cytoplasmic accumulation of mitochondrial DNA (mtDNA), and ultimately blocking the activation of the cGAS-STING pathway and subsequent production of pro-inflammatory cytokines, including TNF-α and IL-1β. Further in vivo and in vitro andvalidation experiments demonstrated that the anti-fibrotic effects of Sal could be reversed by combined treatment with PI3K or mTOR agonists, specifically manifested as activation of the PI3K/AKT/mTOR signaling pathway, reduced hepatocyte mitochondrial autophagy, restoration of cGAS-STING pathway activity, and increased production of pro-inflammatory cytokines. In summary, Our findings demonstrate that SAL mitigates MASH-induced liver fibrosis by targeting hepatocyte autophagy-associated mitochondrial quality control to suppress the mtDNA-dependent cGAS-STING inflammatory pathway, thereby revealing a novel therapeutic strategy.
Infection rates involving bacterial and viral pathogens have increased precipitously after the COVID-19 pandemic, though underlying causes remain elusive. Potential causes ranging from increased hospitalizations during the pandemic or greater use of antibiotics have been proposed, but precisely why rates remain high today remains unknown. Here, we demonstrate that decreased mitochondrial function in antigen-specific T cells post-COVID may contribute to higher infection susceptibility by metabolically immobilizing T cell responses. Using donor-matched peripheral blood samples from 31 COVID-naïve individuals who subsequently contracted COVID-19, we tracked how influenza A (IAV), Staphylococcus aureus (SA), and Varicella-zoster virus (VZV)-stimulated T cell responses were impacted by SARS-CoV-2 infection. Post-COVID CD4 memory T cells exhibited decreased activation- and increased mitochondrial redox-related gene expression. Despite this, mitochondrial flux and reactive oxygen species production were functionally limited in post-COVID antigen-specific T cells after stimulation with IAV, SA, and VZV. Post-COVID plasma was depleted in carnitine and TCA cycle species important for activating fatty acid oxidation, and this correlated with a disordered relationship between memory T cell mobilization of glycolysis, fatty acid metabolism, and oxidative phosphorylation pathways. Metabolic perturbations ultimately resulted in diminished use of catabolic, energy-generating pathways including glycolysis and fatty acid oxidation in antigen-specific T cells. Activating mitochondrial function with metformin and ubiquinol partially rescued the post-COVID decline in T cell catabolism. Collectively, these findings indicate that COVID-19 infection may inhibit T cell metabolism upon exposure to commonly encountered pathogens, which can be partly corrected with commonly available medications that activate mitochondrial metabolism. Our findings may have significant implications for the clinical care of immunologically vulnerable populations in the post-pandemic era.
Pontocerebellar hypoplasia type 6 (PCH6) is caused by biallelic pathogenic variants in RARS2, encoding mitochondrial arginyl-tRNA synthetase. Although mitochondrial dysfunction is a recognised feature, how RARS2 deficiency disrupts neural lineage development remains unclear. We generated rars2-deficient zebrafish using the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system and performed single-cell RNA sequencing (scRNA-seq) at 48 hours postfertilisation, complemented by immunofluorescence, in situ hybridisation, behavioural assays and ultrastructural analyses. Neural lineage composition, developmental trajectories, intercellular communication and transcriptional programmes were systematically examined. rars2 -/- zebrafish displayed impaired survival, locomotor deficits, early mitochondrial ultrastructural damage and marked disruption of neurogenesis. scRNA-seq revealed reduced neuronal populations and expansion of neural progenitor and glial-like cells. Key neurogenic regulators (neurod4, her6 and pou3f1) were downregulated, whereas glial and stress-associated markers (hmgb1a, fabp7a and foxp1b) were upregulated. Developmental pathways including Notch and non-canonical Wnt were attenuated while extracellular matrix (ECM), adhesion and inflammatory programmes were activated. Additional trajectory-based analyses supported dysregulated lineage progression characterised by glial programme activation and impaired maintenance of neurogenic differentiation. RARS2 deficiency disrupts mitochondrial integrity and reprograms neural lineage development through coordinated suppression of neurogenic transcriptional networks and activation of glial/ECM programmes. These findings provide mechanistic insight into loss-of-function RARS2 deficiency and highlight candidate molecular pathways for future therapeutic investigation.
Coenzyme Q (CoQ) is an essential electron carrier and lipophilic antioxidant whose biological functions depend on its redox state. However, accurate measurement of reduced and oxidized CoQ is hindered by rapid ex vivo oxidation during sample preparation and analysis. Here, we develop a dual-isotope-based oxidation-correction LC-MS/MS platform that enables accurate quantification of CoQ redox status across diverse biological matrices. Coupled with rapid subcellular fractionation, this approach resolves cytosolic and mitochondrial CoQ pools and reveals that the CoQ10H2/CoQ10 ratio is substantially higher in the cytosol than in mitochondria. During ferroptosis, compartment-specific remodeling of CoQ redox status is observed, consistent with distinct roles for CoQ in plasma membrane and mitochondrial biology. The platform also enables robust analysis of plasma CoQ redox status in mouse and human samples. These advances provide a broadly applicable approach for studying redox biology, ferroptosis, mitochondrial metabolism, and biomarker discovery.