Search Results (528)

Pulmonary fibrosis (PF) is a progressive lung condition characterized by irreversible scarring and high mortality, with limited effective treatments. Mitochondrial dysfunction has emerged as a critical factor in fibroblast activation in PF, although approaches to restore mitochondrial function remain underexplored. The present study investigated whether mitochondrial transfer from alveolar type II epithelial cells (A549) to patient-derived fibroblasts could restore mitochondrial function and bioenergetics. Histological analysis of fibrotic lungs reveals increased collagen deposition and elevated profibrotic markers, accompanied by reduced expression of mitochondrial biogenesis and respiratory proteins compared to non-fibrotic controls, indicating mitochondrial impairment. Freshly isolated donor mitochondria were functionally validated before mitoception using Seahorse analysis and patient-derived fibroblasts were confirmed by qRT-PCR using fibroblast-specific markers. In vitro transfer of mitochondria to diseased patient-derived fibroblasts exhibited a modest, dose and time-dependent increase in mitochondrial membrane potential compared to normal fibroblasts. Gene expression analysis revealed decreased fibrosis-associated markers and increased expression of mitochondrial and antioxidant genes following mitoception. Seahorse analysis after mitoception revealed enhanced ATP-linked respiration and improved selected mitochondrial bioenergetic parameters, whereas maximal respiration and spare respiratory capacity demonstrated variable responses. In contrast, normal fibroblasts displayed minimal changes. Collectively, these findings indicate that mitochondrial transfer modulates fibroblast bioenergetics and profibrotic signaling, supporting its potential as a therapeutic strategy for pulmonary fibrosis. Full article

Aging is the dominant risk factor for most chronic diseases, yet the mechanisms driving this relationship remain poorly integrated across biological scales. Existing frameworks have catalogued key hallmarks of aging but do not explain how these processes converge to produce organism-level decline and multimorbidity. A systems-level framework is introduced in which aging is conceptualized as progressive destabilization of interacting regulatory networks. Mitochondrial quality control, nutrient-sensing pathways, and chronic inflammatory signaling form a putative high-centrality network core: mitochondria coordinate redox balance, bioenergetics, and transcriptional adaptation, while NAD⁺-dependent signaling and NLRP3 inflammasome activation propagate perturbations across regulatory layers. This architecture provides a mechanistic basis for the convergence of neurodegenerative, cardiovascular, metabolic, and oncological phenotypes as emergent consequences of shared network instability. Reframing the hallmarks as coupled network nodes shifts the explanatory focus from isolated mechanisms to system-level resilience and non-linear dynamics. This narrative and conceptual review integrates evidence across mitochondrial biology, metabolic signaling, and inflammatory pathways to develop these arguments, with explicit acknowledgment that the proposed framework is hypothesis-generating rather than formally validated. Interventions targeting high-centrality nodes, including mTOR modulation, NAD⁺ restoration, mitophagy activation, and anti-inflammatory strategies, may exert system-wide effects by reconfiguring network dynamics rather than correcting individual pathways. This perspective suggests that biomarker-stratified, network-calibrated interventions may offer a broader systems-level therapeutic rationale than single-pathway approaches. Full article

Background/Objectives: Furan derivatives are commonly encountered in food and environmental matrices and may exert biological effects, but their acute cardiovascular actions and potential mitochondrial targets remain insufficiently characterised. This study examined the effects of two simple furan compounds, 2-acetylfuran (2AF) and 5-methylfurfural (5MFF), on arterial blood pressure in vivo and on oxidative phosphorylation in isolated rat liver mitochondria. Methods: Arterial blood pressure was recorded invasively in anaesthetised rats after intraperitoneal administration of 2AF or 5MFF (0.3 µL/g). Systolic, diastolic, and mean arterial pressures, as well as heart rate, were monitored over time. Mitochondrial respiration was assessed in isolated rat liver mitochondria using high-resolution respirometry. Results: Both 2AF and 5MFF induced a rapid hypotensive response, with significant reductions in systolic, diastolic, and mean arterial pressures within 10–15 min after administration. MAP was reduced to a similar extent by both compounds. However, their chronotropic and pulse pressure responses differed: 5MFF increased heart rate and pulse pressure, whereas 2AF induced a delayed bradycardic response without a significant change in pulse pressure. In isolated liver mitochondria, both compounds markedly reduced ADP-stimulated respiration and decreased the respiratory control index, indicating reduced coupling efficiency. Both compounds also increased the cytochrome c effect, suggesting partial impairment of outer mitochondrial membrane integrity. Conclusions: 2AF and 5MFF exert acute hypotensive effects in anaesthetised rats and impair oxidative phosphorylation in isolated rat liver mitochondria. This study provides the first in vivo evidence that 2AF and 5MFF exert hypotensive effects and identifies them as bioactive furan compounds with dual haemodynamic and bioenergetic actions. Full article

Background: Systemic arterial hypertension (SAH) induced by Nω-nitro-L-arginine methyl ester (L-NAME) is a well-established model characterized by nitric oxide (NO^●) synthase inhibition and vascular dysfunction. The transient receptor potential vanilloid 1 (TRPV1) regulates Ca²⁺ flux and may contribute to mitochondrial homeostasis. We hypothesized that TRPV1 activation modulates mitochondria function and attenuates cardiac damage during SAH. Methods: Hypertension was induced in Wistar rats by administration of L-NAME (200 mg/L) for 40 days. During the last four days, hypertensive animals received capsaicin (5 mg/kg/day), capsazepine (6 mg/kg/day), or their combination. Cardiac function was evaluated in isolated hearts using the Langendorff perfusion system. Myocardial tissue viability was assessed by triphenyltetrazolium chloride (TTC) staining, and mitochondrial function was evaluated by measuring respiratory control and apoptosis-related proteins. Results: Capsaicin treatment was associated with significant cardioprotective effects in hypertensive rats. Although the findings are consistent with a role of TRPV1 activation in mediating these effects, the partial protection observed with capsazepine suggests that TRPV1-independent mechanisms may also contribute. Conclusions: TRPV1 activation contributes to cardioprotection in SAH, likely through preservation of mitochondrial function and redox balance. However, additional mechanisms beyond TRPV1 modulation may also participate in the observed protective effects. Further studies—including direct assessment of mitochondrial Ca²⁺ flux and the use of more selective or genetic approaches—are currently underway to clarify the underlying mechanisms. Full article

Tauopathies are a group of neurodegenerative diseases characterized by the accumulation of abnormal tau protein, leading to mitochondrial dysfunction. Because of neurons’ high energy demands, such impairments significantly contribute to neuronal vulnerability. Recent evidence indicates that mitochondria can be transferred between cells to support energy-deficient cells through intercellular mitochondrial transfer (IMT). Given the impact of pathological tau on mitochondrial transport and cytoskeletal dynamics, we hypothesized that IMT is altered in tauopathies. We investigated IMT from astrocytes to neurons, as well as the influence of abnormal tau protein on this process, using co-cultures of SH-SY5Y cells (neuronal model) and A172 cells (astrocytic model). Key data were then confirmed in human iPSC-derived neurons and astrocytes. We show that IMT is enhanced in the presence of abnormal tau and occurs predominantly through contact-dependent mechanisms. Transferred mitochondria were either integrated into the host mitochondrial network, degraded in lysosomes, or remained isolated in the recipient cells’ cytosol. This transfer improved cellular respiration and was associated with increased bioenergetics in pathological cells. Together, our results highlight IMT as a link between tau pathology and neuronal metabolic adaptation, suggesting that this process reflects an endogenous metabolic adaptation holding therapeutic potential to mitigate energy deficits in neurodegenerative diseases. Full article

Background/Objectives: Methylxanthines, such as caffeine, exhibit neuroprotective properties in neurodegenerative conditions, partly linked to modulation of monoamine oxidase B (MAO-B) and oxidative stress pathways. This work aimed to design, synthesize and functionally characterize new caffeine-8-methylthioglycolic acid derivatives as selective MAO-B inhibitors with neuroprotective potential. Methods: A QSAR model was built on 94 studies of xanthine derivatives to guide the design of ten new semi- and thiosemicarbazides (Jas1–Jas10), followed by molecular docking to human MAO-B (PDB: 2V5Z) using Glide, GOLD and MM-GBSA binding free energy calculations. The target compounds were synthesized in relatively high yields, structurally confirmed by spectroscopic methods and tested in vitro for hMAO-A/B inhibition, as well as for neurotoxicity and neuroprotection in isolated mouse brain synaptosomes, mitochondria and microsomes under 6-hydroxydopamine (6-OHDA), tert-butyl hydroperoxide (t-BuOOH) and Fe/ascorbate (Fe²⁺/AA)-induced oxidative stress. Results: Docking and MM-GBSA identified Jas6 and Jas7 as the most stable MAO-B binders, with binding free energies approaching those of safinamide. All derivatives inhibited hMAO-A and hMAO-B in the submicromolar range, with Jas2 and Jas6 showing the highest MAO-B selectivity indices. At 100 µM, the series produced mild but significant pro-oxidant and cytotoxic effects when applied alone, yet under oxidative stress all compounds, especially Jas2 and Jas6, markedly preserved synaptosomal and mitochondrial viability, maintained glutathione levels, and reduced malondialdehyde production. Conclusions: The caffeine-based semi- and thiosemicarbazides, particularly Jas2 and Jas6, emerge as promising selective MAO-B inhibitors with pronounced antioxidant and neuroprotective activity, supporting their further optimization as multitarget candidates for neurodegenerative disorders such as Parkinson’s disease. Full article

Pregnancy-associated hemodynamic overload and hormonal changes induce hypertrophy and metabolic remodeling of the maternal heart. Mitochondrial motility, mediated by ras homolog family member T (RHOT) 1 and RHOT2, is essential for cardiac adaptation to increased workload, cardiomyocyte hypertrophy, and sarcomere maturation. To test the hypothesis that Rhot1/2 expression is required for pregnancy- and postpartum-associated adaptations of the maternal heart, female mice with tamoxifen-inducible, cardiomyocyte-selective deletion of Rhot1 and Rhot2 (iRhot1/2-KO) were mated. Following gene deletion in adult mice, cardiac tissue and function were analyzed after three to five successive pregnancies and postpartum nursing periods. Age-matched nulliparous iRhot1/2-KO mice and age-matched mice expressing Rhot1 and Rhot2 served as controls. Motility of mitochondria isolated from iRhot1/2-KO hearts was impaired, as determined by the number of mobile mitochondria in an in vitro motor protein-driven single mitochondrion motility assay performed on surface-immobilized microtubules. Despite loss of Rhot1/2 expression, contractile function assessed by transthoracic echocardiography, mRNA expression of peripartum-associated heart failure markers, cardiac structure, mitochondrial morphology, mitochondrial enzymatic activity, and mitochondrial DNA content were all comparable to controls expressing Rhot1/2 at the investigated time points. RNA sequencing-based gene profiling identified a transcriptional program through which RHOT proteins preserve cardiac energetic and contraction gene expression during pregnancy and postpartum. Together, cardiomyocyte-selective loss of Rhot1/2 expression in the adult heart does not cause peripartum-associated heart failure, despite reduced cardiac energetic and contraction gene expression. Full article

Background: The NLRP3 inflammasome drives pathological inflammation in various diseases. PINK1/Parkin-associated mitophagy serves as a critical negative regulator of NLRP3 activation, yet pharmacological enhancers remain scarce. Muscone, a natural macrocyclic ketone with blood–brain barrier permeability, exhibits potent anti-inflammatory properties; however, its mechanistic role within the NLRP3-mitophagy axis remains undefined. Methods: LPS/ATP-stimulated macrophages were employed to assess stage-specific effects of muscone on NLRP3 priming (NF-κB signaling, NLRP3, and pro-IL-1β expression) and activation (ASC oligomerization, ASC–pro-caspase 1 complex formation, and IL-1β secretion). RNA sequencing and bioinformatic analysis were performed for pathway enrichment. Mitophagy was characterized by MitoSOX Red staining for mt-ROS detection, electron microscopy, Western blotting of LC3B-II in isolated mitochondria and PINK1 and Parkin in whole-cell lysates, and live-cell mitochondria–lysosome tracking. In vivo protective efficacy was assessed in an LPS-induced endotoxemia mouse model. Results: Muscone dose-dependently suppressed both the priming and activation stages of the NLRP3 inflammasome, maximally reducing IL-1β secretion by ~60% at 50 μM. Mechanistically, muscone amplified PINK1/Parkin-associated mitophagy, scavenging excessive mt-ROS and attenuating NLRP3 activation. These effects were corroborated by RNA-seq and comprehensive functional assays. In vivo, muscone (30 mg/kg) significantly improved survival (3/8 mice alive at 98 h when all LPS controls had died; 2/8 survived to the 132-h endpoint), with concomitant enhancement of mitophagy markers in peritoneal macrophages. Conclusions: Muscone functions as a PINK1/Parkin-associated mitophagy enhancer that maintains mitochondrial quality control during NLRP3-driven inflammatory responses. Its unique macrocyclic structure and blood–brain barrier permeability provide a promising scaffold for developing therapeutics against inflammatory disorders associated with NLRP3 inflammasome activation. Full article

Herbicides, used worldwide to improve agricultural yields, are associated with pollution and significant health problems. Cardiac damage is a major concern, and the respective contributions of glyphosate (GP) and its commercial formulation, Roundup^® (RU), warrant investigation. We studied the specific effects of GP and RU on isolated rat cardiac mitochondria and on H9c2 cardiomyocytes cultured for 6 and 24 h to determine whether the potential cardiotoxicity of GP and/or RU are linked to impaired mitochondrial respiration and increased hydrogen peroxide (H₂O₂) production. To this end, we used various mitochondrial complex substrates and a high-resolution oxygraphy. Unlike the GP alone which demonstrated no significant effect, the RU decreased cardiac mitochondrial respiration (21.90 ± 2.99 vs. 41.23 ± 7.09 pmol/s/mL, −46.9%, p = 0.007) for OXPHOS CI in respectively the RU and the control groups. RU also impaired OXPHOS CI+II (−51.5%, p = 0.003), maximal mitochondrial respiration (ETS CI+II, −46.7%, p = 0.001) and coupling (−35.4%, p = 0.0003). Similarly, 24 h exposure to RU decreased H9c2 cell number (−48.59%, p = 0.0023) but increased their mitochondrial respiration (+38.2%, p = 0.03, +37.6%, p = 0.03, +43.2%, p = 0.03 for OXPHOS CI, OXPHOS CI+II and ETS CI+II respectively). We observed a similar trend (NS) after 24 h exposure to GP. In conclusion, these results support an enhanced cardiac toxicity of the Roundup^® as compared to the glyphosate. Both decreased mitochondrial respiration and increased hydrogen peroxide production were involved in isolated mitochondria impairment. After 24 h exposure to Roundup^®, a compensatory mechanism potentially counterbalanced the decreased H9c2 cell number. These data support future studies aiming to reduce Roundup^®-associated cardiac alterations not only by reducing its use but also by investigating the effectiveness of antioxidant and mitochondria-focused therapy. Full article

MAO-B-specific inhibition, either in knockout (KO) mice or pharmacologically, preserves left ventricular function and reduces cardiac fibrosis after myocardial infarction or pressure overload. We investigated whether stimulation of MAO-B in cardiac fibroblasts provokes ROS production and myofibroblast development. Fibroblast-specific MAO-B knockdown (KD) mice were created by crossing Col1a2CreERT mice with MAO-B^fl/fl mice. The KD was induced by tamoxifen injection. Fibroblasts of KD mice and wild types (WTs) were isolated and reduced MAO-B expression in KD fibroblasts was confirmed. In isolated mitochondria from the left ventricle of these mice, ROS production was reduced under stimulation with the specific MAO-B substrate β-phenylethylamine (PEA). Mitochondrial ROS production in fibroblasts, detected by MitoSox Red staining, increased under PEA (1000 µM) stimulation only in WT fibroblasts. mRNA of the marker genes for myofibroblast differentiation, Col1a1 and periostin, increased 2- or 3-fold, respectively, in WT but not in MAO-B KD fibroblasts. The enhanced migration potential under PEA was reduced in MAO-B KD fibroblasts. In conclusion, stimulation of MAO-B in cardiac fibroblasts leads to the formation of mitochondrial ROS, enhancement of myofibroblast marker gene expression and migration of the cells. Excessive fibrosis caused by elevated MAO-B activity in myocardial infarction can therefore contribute to cardiac dysfunction. Full article

Mitochondria are vital organelles for human cells with fundamental roles in major metabolic processes such as calcium homeostasis, ATP production, apoptosis and signal transduction. Defective morphology and activity of these organelles have been tightly associated with the pathological onset of severe human disorders, including cardiovascular diseases. Targeting mitochondrial dysfunction has been an area of extensive research encompassing several approaches ranging from pharmacological agents to mitochondrial replacement techniques. Among them, mitochondrial transplantation has been a rapidly evolving approach, especially in the field of cardiovascular dysfunction for the restoration of injured or damaged myocardial cells. Various methods including tunneling nanotubes, nanoblade and “mitopunch” ensure the effective mitochondrial transfer from the donor to the recipient cell, with the internalization of the organelles, via endocytosis, enabling functional restoration. Results of preclinical and clinical trials involving mitochondrial transfer support the application of this technique in improving the function of the myocardium after damage caused by ischemia reperfusion injury. Herein, we discuss the beneficial role of mitochondrial transplantation in cardiovascular diseases and the current technical challenges of mitochondrial isolation, preservation, and targeted delivery, as well as their role in advancing precision medicine, offering a patient tailored therapeutic approach. Full article

The ATP-dependent inhibition of cytochrome c oxidase (CytOx, complex IV of the electron transport chain) is the second mechanism of respiratory control adjusting mitochondrial respiration in order to prevent excessive electron flow and reactive oxygen species (ROS) production. Here, we investigate how tricarboxylic acid (TCA) cycle metabolites and the subsequent complex I or complex II activities influence this regulatory mechanism. Therefore, CytOx activity was assessed by the oxygen consumption rate after cytochrome c (Cyt c) titration to stimulate complex IV activity in isolated rat heart mitochondria (RHM) and permeabilized AC16 cells. Mitochondrial membrane potential (Δψm) and ROS formation were analysed by flow cytometry. Our results show that TCA cycle intermediates differed in their impact on CytOx activity and subsequent ROS formation. NADH-linked substrates such as α-ketoglutarate, glutamate and malate increased respiratory capacity, but preserved ATP-dependent control of CytOx, indicating that elevated electron supply alone does not necessarily abolish ATP sensitivity. In contrast, succinate, which feeds electrons directly into complex II, strongly increased respiration causing the loss of ATP-dependent respiratory control in both model systems. Despite this strong respiratory effect, succinate induced only modest changes in mitochondrial membrane potential in isolated mitochondria, whereas permeabilized cardiomyocytes exhibited reduced polarization accompanied by increased superoxide formation. Together, these findings demonstrate that the effectiveness of ATP-dependent CytOx inhibition is influenced by TCA cycle activity and depends on the site of electron entry into the respiratory chain. Thus, substrate-dependent modulation of respiratory control links metabolite availability to mitochondrial redox regulation in cardiac cells. Full article

Background/Objectives: Mitochondria are dynamic organelles that continuously undergo balanced cycles of fusion and division to maintain optimal function. Mitochondrial division is mediated by Dynamin-Related Protein 1 (DRP1), a cytosolic large GTPase whose phosphorylation at serine 616 (DRP1-S616Ⓟ) promotes its translocation to the outer mitochondrial membrane and organelle division. Dysregulated mitochondrial division disrupts cellular homeostasis and contributes to disease pathogenesis, including cancer. Our prior work demonstrated that the oncogene-induced mitogen-activated protein kinase (MAPK) pathway constitutively phosphorylates DRP1 at serine 616, which is essential to cellular transformation and correlates with oncogene status in patient tissues. Similarly, DRP1-S616Ⓟ is subject to pharmacologic control by targeted therapies against oncogenic MAPK signaling. Methods: Building upon this foundation, we developed and characterized a recombinant murine monoclonal antibody (referred to as 3G11) with high specificity for human DRP1-S616Ⓟ, raised against a peptide derived from the human DRP1 sequence. Results: Using diverse experimental platforms, we demonstrate the robust utility of 3G11 to detect DRP1-S616Ⓟ in melanoma cell extracts and isolated organelles. Immunofluorescence revealed that pharmacologic inhibition of oncogenic MAPK signaling reduces DRP1-S616Ⓟ levels, which correlates with mitochondrial hyperfusion, while immunohistochemistry showed that elevated DRP1-S616Ⓟ expression in human tissues correlates with BRAF^V600E disease. Conclusions: 3G11 is a new recombinant antibody for detecting DRP1-S616Ⓟ and supports studies of mitochondrial division in cancer. Together, these findings establish 3G11 as a specific, versatile, renewable, and cost-effective tool for studying mitochondrial division, with strong potential for clinical applications. Full article

(1) Background: Mild traumatic brain injury (mTBI), the most prevalent form of traumatic brain injury, often results from repetitive impacts to the head and is associated with long-term neurological impairment. The pathophysiology of mTBI is multifactorial and involves alterations in mitochondrial bioenergetics, a key determinant of neuronal function and survival. Although mitochondrial dysfunction is recognized as a hallmark of mTBI, its long-term effects on bioenergetics and the roles of regulatory cytosolic and mitochondrial proteins remain poorly understood. We hypothesized that repeated mTBI (rmTBI) induces sustained deficits in mitochondrial bioenergetics that are associated with long-term changes in key bioenergetic and other regulatory proteins. (2) Methods: Using the repeated CHIMERA injury model in adult male rats, randomly assigned to sham or rmTBI groups, we assessed mitochondrial respiration in isolated mitochondria and whole cerebral cortex homogenates using a Clark O₂ electrode and an Oroboros O₂k respirometer at time points ranging from 1 day to 2 months post-injury. Western blotting was performed for expression of regulatory proteins HKI, DRP1, MFN2, VDAC1, and ANT2. (3) Results: At 2 months post-rmTBI, respiration was faster and uncoupled, while ATP synthesis was significantly slowed compared with sham rats. This was accompanied by decreased expression of mitochondrial MFN2 and ANT2, by increased mitochondrial expression of DRP1, and by decreased translocation of HKI to mitochondria. There was no significant difference in VDAC1 expression. Earlier time points showed no significant differences in bioenergetics or protein expression, but neuro-inflammatory markers (GFAP and Iba1) were significantly elevated at these earlier time points of post-injury. (4) Conclusions: These findings indicate that rmTBI leads to a delayed long-term impairment of mitochondrial bioenergetics associated with alterations in proteins critical for bioenergetic regulation and mitochondrial control. This suggests a pathophysiologic mechanism for the persistent cognitive and behavioral deficits observed following rmTBI. Full article

Ischemic stroke induces complex molecular responses that disrupt subcellular organelles’ function and contribute to brain injury, yet the temporal changes of organelle-specific transcriptomic remodeling remain to be investigated. In this study, we performed in silico analysis of publicly available transcriptomic data from isolated brain microvessels of transient middle cerebral artery occlusion (tMCAO) mouse model. Using in silico approaches, we analyzed differential gene expression at 24 h (acute phase) and 7 d (intermediate phase) post-stroke, focusing on mitochondria, endoplasmic reticulum (ER), and Golgi apparatus. Functional enrichment (Gene Ontology, KEGG) and protein–protein interaction network analyses were performed. Our analysis of the data revealed that at 24 h post-stroke, all three organelles exhibited marked transcriptional remodeling, where mitochondrial pathways showed disrupted metabolic and redox regulation; ER pathways indicated activation of biosynthetic processes, stress signaling, and ferroptosis; and Golgi-related genes reflected altered vesicular trafficking and glycosylation. By 7 d, mitochondrial alterations subsided, whereas ER and Golgi pathways displayed downregulation of metabolic and neuronal signaling processes, indicating persistent dysfunction and incomplete microvascular recovery. Phase-specific drug–gene interaction analysis will be useful to understand temporal organelle-associated transcriptional organization and to guide future investigations of neurovascular remodeling after ischemic stroke. Full article