Mitochondrial Cristae Research Articles

MCAUSE: Prioritizing mitochondrial targets that alleviate pancreatic cancer cell phenotypes

Substantial changes in energy metabolism are a hallmark of pancreatic cancer. To adapt to hypoxic and nutrient-deprived microenvironments, pancreatic cancer cells remodel their bioenergetics from oxidative phosphorylation to glycolysis. This bioenergetic shift makes mitochondria an Achilles' heel. Since mitochondrial function remains essential for pancreatic cancer cells, further depleting mitochondrial energy production is an appealing treatment target. However, identifying effective mitochondrial targets for treatment is challenging. Here, we developed an approach, mitochondria-targeted cancer analysis using survival and expression (mCAUSE), to prioritize target proteins from the entire mitochondrial proteome. Selected proteins were further tested for their impact on pancreatic cancer cell phenotypes. We discovered that targeting a dynamin-related GTPase, OPA1, which controls mitochondrial fusion and cristae, effectively suppresses pancreatic cancer activities. Remarkably, when combined with a mutation-specific KRAS inhibitor, OPA1 inhibition showed a synergistic effect. Our findings offer a therapeutic strategy against pancreatic cancer by simultaneously targeting mitochondria dynamics and KRAS signaling.

Apoptosis and Oxidative Stress in Human Intestinal Epithelial Caco-2 Cells Caused by Marine Phycotoxin Azaspiracid-2.

When humans consume seafood contaminated by lipophilic polyether phycotoxins, such as azaspiracids (AZAs), the toxins are mainly leached and absorbed in the small intestine, potentially causing intestinal damage. In this study, human intestinal epithelial Caco-2 cells were used to investigate the adverse effects of azaspiracid-2 (AZA-2) on human intestinal epithelial cells. Cell viability, apoptosis, oxidative damage and mitochondrial ultrastructure were investigated, and ribonucleic acid sequence (RNA-seq) analysis was applied to explore the potential mechanisms of AZA-2 toxicity to Caco-2 cells. Results showed that AZA-2 significantly reduced the proliferation of Caco-2 cells in a concentration-dependent response, and the 48 h EC50 of AZA-2 was 12.65 nmol L-1. AZA-2 can induce apoptosis in Caco-2 cells in a dose-dependent manner. Visible mitochondrial swelling, cristae disintegration, membrane rupture and autophagy were observed in Caco-2 cells exposed to AZA-2. Reactive oxygen species (ROS) and malondialdehyde (MDA) content were significantly increased in Caco-2 cells after 48 h of exposure to 1 and 10 nmol L-1 of AZA-2. Transcriptome analysis showed that KEGG pathways related to cellular oxidative damage and lipid metabolism were affected, mainly including mitophagy, oxidative phosphorylation, cholesterol metabolism, vitamin digestion and absorption, bile secretion and the peroxisome proliferator-activated receptor signaling pathway. The cytotoxic effects of AZA-2 on Caco-2 cells may be associated with ROS-mediated autophagy and apoptosis in mitochondrial cells. Results of this study improve understanding of the cytotoxicity and molecular mechanisms of AZA-2 on Caco-2 cells, which is significant for protecting human health.

CHCHD2 mutant mice display mitochondrial protein accumulation and disrupted energy metabolism.

Mutations in the mitochondrial cristae protein CHCHD2 lead to a late-onset autosomal dominant form of Parkinson's disease (PD) which closely resembles idiopathic PD, providing the opportunity to gain new insights into the mechanisms of mitochondrial dysfunction contributing to PD. To begin to address this, we used CRISPR genome-editing to generate CHCHD2 T61I point mutant mice. CHCHD2 T61I mice had normal viability, and had only subtle motor deficits with no signs of premature dopaminergic (DA) neuron degeneration. Nonetheless, CHCHD2 T61I mice exhibited robust molecular changes in the brain including increased CHCHD2 insolubility, accumulation of CHCHD2 protein preferentially in the substantia nigra (SN), and elevated levels of α-synuclein. Metabolic analyses revealed an increase in glucose metabolism through glycolysis relative to the TCA cycle with increased respiratory exchange ratio, and immune-electron microscopy revelated disrupted mitochondria in DA neurons. Moreover, spatial genomics revealed decreased expression of mitochondrial complex I and III respiratory chain proteins, while proteomics revealed increased respiratory chain and other mitochondrial protein-protein interactions. As such, the CHCHD2 T61I point-mutation mice exhibit robust mitochondrial disruption and a consequent metabolic shift towards glycolysis. These findings thus establish CHCHD2 T61I mice as a new model for mitochondrial-based PD, and implicate disrupted respiratory chain function as a likely causative driver.

Mitochondrial Physiology of Cellular Redox Regulations.

Mitochondria (mt) represent the vital hub of the molecular physiology of the cell, being decision-makers in cell life/death and information signaling, including major redox regulations and redox signaling. Now we review recent advances in understanding mitochondrial redox homeostasis, including superoxide sources and H2O2 consumers, i.e., antioxidant mechanisms, as well as exemplar situations of physiological redox signaling, including the intramitochondrial one and mt-to-cytosol redox signals, which may be classified as acute and long-term signals. This review exemplifies the acute redox signals in hypoxic cell adaptation and upon insulin secretion in pancreatic beta-cells. We also show how metabolic changes under these circumstances are linked to mitochondrial cristae narrowing at higher intensity of ATP synthesis. Also, we will discuss major redox buffers, namely the peroxiredoxin system, which may also promote redox signaling. We will point out that pathological thresholds exist, specific for each cell type, above which the superoxide sources exceed regular antioxidant capacity and the concomitant harmful processes of oxidative stress subsequently initiate etiology of numerous diseases. The redox signaling may be impaired when sunk in such excessive pro-oxidative state.

Effects on Iron Metabolism and System Xc- /GPX4 Pathway from Hydroquinone Suggest Ferroptosis of Jurkat Cells.

Prolonged exposure to hydroquinone (HQ), a metabolite of benzene, can cause severe haematologic disorders in humans. However, the mechanism is still unclear. In the present study, we investigated whether HQ can induce haematological diseases through ferroptosis, which is another form of cell death apart from apoptosis. The results showed that HQ inhibited the viability of Jurkat cells in a dose-dependent and time-dependent manner. The half inhibitory concentrations (IC50s) of HQ-treated Jurkat cells for 12 h, 24 h and 48 h were 107.16 μmol/L, 33.29 μmol/L, and 14.78 μmol/L. The exposure of Jurkat cells to HQ increased intracellular Fe2+, malondialdehyde (MDA) and lipid reactive oxygen species (ROS) levels and down-regulated glutathione (GSH) levels. We used erastin-treated cells as a positive control and cells treated with HQ combined with deferoxamine mesylate (DFO) and ferrostain-1 (Fer-1)-treated cells as the negative controls. DFO and Fer-1 partially restored the degradation of cell viability and GSH content and the accumulation of Fe2+, MDA and lipid ROS caused by HQ. In addition, we found that cellular mitochondria in the HQ-treated group showed a decrease in volume, an increase in the density of the bilayer membrane and a decrease or disappearance of mitochondrial cristae. Changes in the erastin-treated group were similar to those in the HQ-treated group. We inferred that HQ induces ferroptosis in Jurkat cells. Subsequently, we found that HQ up-regulated the levels of transferrin receptor 1 (TFRC) mRNA and protein expression and down-regulated FTH1, SLC7A11 and synthetic substrate of antioxidant enzyme 4 (GPX4) mRNA levels and protein expression levels. However, the exposure of Jurkat cells to HQ with DFO and Fer-1 alleviated these changes. Notably, the activation of TFRC and the inhibition of FTH1 and System Xc- (cystine-glutamate reverse transporter protein) /GPX4 were associated with HQ-induced ferroptosis. These results provide novel insights into how HQ exacerbates haematopoietic cytotoxicity and provide potential targets for the prevention of HQ-induced diseases.

Electroacupuncture pretreatment maintains mitochondrial quality control via HO-1/MIC60 signaling pathway to alleviate endotoxin-induced acute lung injury

Electroacupuncture has been demonstrated to mitigate endotoxin-induced acute lung injury by enhancing mitochondrial function. This study investigates whether electroacupuncture confers lung protection through the regulation of mitochondrial quality control mediated by heme oxygenase-1 (HO-1) and the mitochondrial inner membrane protein MIC60. HO-1, an inducible stress protein, is crucial for maintaining mitochondrial homeostasis and protecting against lung injury. MIC60, a key component of the mitochondrial contact site and cristae organizing system, supports mitochondrial integrity.We employed genetic knockout/silencing and cell transfection techniques to model lipopolysaccharide (LPS)-induced lung injury, assessing changes in mitochondrial structure, reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP), and the expression of proteins essential for mitochondrial quality control. Our findings reveal that electroacupuncture alleviates endotoxin-induced acute lung injury and associated mitochondrial dysfunction, as evidenced by reductions in lung injury scores, decreased ROS production, and suppressed expression of proteins involved in mitochondrial fission and mitophagy. Additionally, electroacupuncture enhanced MMP and upregulated proteins that facilitate mitochondrial fusion and biogenesis. Importantly, the protective effects of electroacupuncture were reduced in models with Hmox1 knockout or Mic60 silencing, and in macrophages transfected with Hmox1-siRNA or Mic60-siRNA. Moreover, HO-1 was found to influence MIC60 expression during electroacupuncture preconditioning and LPS challenge, demonstrating that these proteins not only co-localize but also interact directly.In conclusion, electroacupuncture effectively modulates mitochondrial quality control through the HO-1/MIC60 signaling pathway, offering an adjunctive therapeutic strategy to ameliorate endotoxin-induced acute lung injury in both in vivo and in vitro settings.

Quercetin alleviates depressive-like behavior by modulating acetyl-H3K9 mediated ferroptosis pathway in hypothalamus of perimenopausal depression rat model

Perimenopausal depression is a subtype of depression and is prevalent among perimenopausal women, which has brought a heavy burden to family and society. The pathogenesis of perimenopausal depression is still unclear, which affects the prevention and treatment of perimenopausal depression to a certain extent. Quercetin is a flavonoid compound, and has estrogenic activity and pharmacological effects such as antioxidant, anti-inflammatory, and neuroprotective effects. This study investigated whether quercetin improved perimenopausal depression-like behaviors and potential mechanism. The results demonstrated that quercetin could alleviate the depression-like behaviors in perimenopausal depression rat model, inhibit astrocyte activation, improve ferroptosis-associated mitochondrial damage (such as mitochondrial pyknosis and mitochondrial cristae reduction) in hypothalamus, increase the expressions of histone 3 lysine 9 acetylation (acetyl-H3K9), ferroptosis-associated protein including glutathione peroxidase 4 (GPX4) and Xc- antiporter (SLC7A11), and reduce the expressions of endoplasmic reticulum stress-related proteins including inositol-requiring enzyme 1 (IRE1α), phosphorylated IRE1α (p-IRE1α), X-box binding protein 1 (XBP1) and glucose-regulated protein 78 (GRP78) in hypothalamus of perimenopausal depression rat model. Furtherly, in vitro study indicated that quercetin could restore histone acetylase (HAT)/histone deacetylase (HDAC) homeostasis through binding to estrogen receptors and increase the expression of acetyl-H3K9, inhibiting ferroptosis through IRE1α/XBP1 pathway in astrocytes of hypothalamus. Our findings demonstrated that acetyl-H3K9 is a crucial target in development of perimenopausal depression, and quercetin exhibited antidepressant effects through modulating acetyl-H3K9 mediated ferroptosis in perimenopausal depression. Quercetin might be the prevention and adjuvant treatment strategy of perimenopausal depression.

Chinese acupuncture: A potential treatment for autism rat model via improving synaptic function

PurposeAutistic symptom improvement can be observed in children treated with acupuncture, but the mechanism is still being explored. In the present study, we used scalp acupuncture to treat autism rat model, and then their improvement in the abnormal behaviors and specific mechanisms behind were revealed by detecting animal behaviors, analyzing the RNA sequencing of the prefrontal cortex (PFC), and observing the ultrastructure of PFC neurons under the transmission electron microscope. MethodsOn gestational day 12.5, Wistar rats were given valproic acid (VPA) by intraperitoneal injection, and their offspring were considered to be reliable rat models of autism. They were randomized to VPA or VPA-acupuncture group (n = 8). Offspring of Wistar pregnant rats that were simultaneously injected with saline were randomly selected as the wild-type group (WT). VPA_acupuncture group rats received acupuncture intervention at 23 days of age for 4 weeks, and the other two groups followed without intervention. After the intervention, all experimental rats underwent behavioral tests. Immediately afterward, they were euthanized by cervical dislocation, and their prefrontal cortex was isolated for RNA sequencing and transmission electron microscopy. ResultsThe main results are as follows: 1. Animal behavioural tests: VPA group rats showed more anxiety-like behaviour and repetitive, stereotyped behaviour than WT group rats. While VPA group rats showed less spatial exploration ability, activity level, social interaction, and social novelty preference than WT group rats. It was gratifying to observe that acupuncture indeed improved these abnormal behaviors of autism rat model. 2. RNA-sequencing: The three groups of rats differed in the expression and enrichment pathways of multiple genes related to synaptic function, neural signal transduction, immune-inflammatory responses and circadian rhythm regulation. Our experiments indicated that acupuncture can alleviate the major symptoms of ASD by improving these neurological abnormalities. 3. Under the transmission electron microscopy, several lysosomes and mitochondrial structural abnormalities were observed in the prefrontal neurons of VPA group rats, which were manifested as atrophy of the mitochondrial membrane, blurring or disappearance of the mitochondrial cristae, and even vacuolization. Moreover, the number of synapses and synaptic vesicles was relatively small. Conversely, the mitochondrial structure of rats in the WT group and VPA_acupuncture was normal, and the number of synapses and synaptic vesicles was relatively large. ConclusionAcupuncture effectively improved the abnormal behaviors of autism rat model and the ultrastructure of the PFC neurons, which might worked by improving their abnormal synaptic function, synaptic plasticity promoting neuronal signal transduction and regulating immune-inflammatory responses.

FAM210A: An emerging regulator of mitochondrial homeostasis.

Mitochondrial homeostasis serves as a cornerstone of cellular function, orchestrating a delicate balance between energy production, redox status, and cellular signaling transduction. This equilibrium involves a myriad of interconnected processes, including mitochondrial dynamics, quality control mechanisms, and biogenesis and degradation. Perturbations in mitochondrial homeostasis have been implicated in a wide range of diseases, including neurodegenerative diseases, metabolic syndromes, and aging-related disorders. In the past decades, the discovery of numerous mitochondrial proteins and signaling has led to a more complete understanding of the intricate mechanisms underlying mitochondrial homeostasis. Recent studies have revealed that Family with sequence similarity 210 member A (FAM210A) is a novel nuclear-encoded mitochondrial protein involved in multiple aspects of mitochondrial homeostasis, including mitochondrial quality control, dynamics, cristae remodeling, metabolism, and proteostasis. Here, we review the function and physiological role of FAM210A in cellular and organismal health. This review discusses how FAM210A acts as a regulator on mitochondrial inner membrane to coordinate mitochondrial dynamics and metabolism.

PRRSV hijacks DDX3X protein and induces ferroptosis to facilitate viral replication

Porcine reproductive and respiratory syndrome virus (PRRSV) is a severe disease with substantial economic consequences for the swine industry. The DEAD-box helicase 3 (DDX3X) is an RNA helicase that plays a crucial role in regulating RNA metabolism, immunological response, and even RNA virus infection. However, it is unclear whether it contributes to PRRSV infection. Recent studies have found that the expression of DDX3X considerably increases in Marc-145 cells when infected with live PRRSV strains Ch-1R and SD16; however, it was observed that inactivated viruses did not lead to any changes. By using the RK-33 inhibitor or DDX3X-specific siRNAs to reduce DDX3X expression, there was a significant decrease in the production of PRRSV progenies. In contrast, the overexpression of DDX3X in host cells substantially increased the proliferation of PRRSV. A combination of transcriptomics and metabolomics investigations revealed that in PRRSV-infected cells, DDX3X gene silencing severely affected biological processes such as ferroptosis, the FoxO signalling pathway, and glutathione metabolism. The subsequent transmission electron microscopy (TEM) imaging displayed the typical ferroptosis features in PRRSV-infected cells, such as mitochondrial shrinkage, reduction or disappearance of mitochondrial cristae, and cytoplasmic membrane rupture. Conversely, the mitochondrial morphology was unchanged in DDX3X-inhibited cells. Furthermore, silencing of the DDX3X gene changed the expression of ferroptosis-related genes and inhibited the virus proliferation, while the drug-induced ferroptosis inversely promoted PRRSV replication. In summary, these results present an updated perspective of how PRRSV infection uses DDX3X for self-replication, potentially leading to ferroptosis via various mechanisms that promote PRRSV replication.

Very virulent infectious bursal disease virus infection triggered microscopic changes, apoptosis, and inflammatory cytokines imbalance in chicken spleen and thymus

ABSTRACT Infectious bursal disease virus (IBDV) can cause a highly contagious disease, resulting in severe damage to the immune system that causes immunosuppression in young chickens. Both spleen and thymus are important immune organs, which play a key role in eliciting protective immune responses. However, the effects of very virulent IBDV (vvIBDV) strain LJ-5 infection on chicken spleen and thymus are still unknown. In the present study, 3-week-old specific pathogen-free chickens were infected with vvIBDV for 1–5 days. The vvIBDV infection significantly increased the spleen index and decreased the thymus index. Microscopic analysis indicated necrosis, depletion of the lymphoid cells, and complete loss of structural integrity in spleen and thymus. Ultrastructural analysis displayed mitochondrial and nuclear damage, including mitochondrial cristae breaks, and deformation of nuclear membrane in vvIBDV-infected spleen and thymus tissues. Cytokine levels increased in the spleen and thymus after IBDV infection, promoting inflammation and causing an inflammatory imbalance. Moreover, the mRNA expression of apoptosis-related genes was significantly upregulated in the vvIBDV-infected group compared to the control group. Meanwhile, the mRNA expression of mitochondrial dynamics was altered in the spleen and thymus of vvIBDV-infected chickens. These results suggested that vvIBDV infection triggers an imbalance of inflammatory cytokines, and apoptosis in the spleen and thymus, resulting in immune injury in chickens. This study provides basic data for the further study of vvIBDV pathogenesis.

MIRO1 controls energy production and proliferation of smooth muscle cells.

The outer mitochondrial Rho GTPase 1, MIRO1, mediates mitochondrial motility within cells, but implications for vascular smooth muscle cell (VSMC) physiology and its roles invascular diseases, such as neointima formation following vascular injury are widely unknown. An in vivo model of selective Miro1 deletion in VSMCs was generated, and the animals were subjected to carotid artery ligation. The molecular mechanisms relevant to VSMC proliferation were then explored in explanted VSMCs by imaging mitochondrial positioning and cristae structure and assessing the effects on ATP production, metabolic function and interactions with components of the electron transport chain (ETC). MIRO1 was robustly expressed in VSMCs within human atherosclerotic plaques and promoted VSMC proliferation and neointima formation in mice by blocking cell-cycle progression at G1/S, mitochondrial positioning, and PDGF-induced ATP production and respiration; overexpression of a MIRO1 mutant lacking the EF hands that are required for mitochondrial mobility did not fully rescue these effects. At the ultrastructural level, Miro1 deletion distorted the mitochondrial cristae and reduced the formation of super complexes and the activity of ETC complex I. Mitochondrial motility is essential for VSMC proliferation and relies on MIRO1. The EF-hands of MIRO1 regulate the intracellular positioning of mitochondria. Additionally, the absence of MIRO1 leads to distorted mitochondrial cristae and reduced ATP generation. Our findings demonstrate that motility is linked to mitochondrial ATP production. We elucidated two unrecognized mechanisms through which MIRO1 influences cell proliferation by modulating mitochondria: first, by managing mitochondrial placement via Ca2+-dependent EF hands, and second, by affecting cristae structure and ATP synthesis.

The Toxoplasma gondii homolog of ATPase inhibitory factor 1 is critical for mitochondrial cristae maintenance and stress response.

The production of energy in the form of ATP by the mitochondrial ATP synthase must be tightly controlled. One well-conserved form of regulation is mediated via ATPase inhibitory factor 1 (IF1), which governs ATP synthase activity and gene expression patterns through a cytoprotective process known as mitohormesis. In apicomplexans, the processes regulating ATP synthase activity are not fully elucidated. Using the model apicomplexan Toxoplasma gondii, we found that knockout and overexpression of TgIF1, the structural homolog of IF1, significantly affected gene expression. Additionally, TgIF1 overexpression resulted in the formation of a stable TgIF1 oligomer that increased the presence of higher order ATP synthase oligomers. We also show that parasites lacking TgIF1 exhibit reduced mitochondrial cristae density, and that while TgIF1 levels do not affect growth in conventional culture conditions, they are crucial for parasite survival under hypoxia. Interestingly, TgIF1 overexpression enhances recovery from oxidative stress, suggesting a mitohormetic function. In summary, while TgIF1 does not appear to play a role in metabolic regulation under conventional growth conditions, our work highlights its importance for adapting to stressors faced by T. gondii and other apicomplexans throughout their intricate life cycles.

Mitochondrial permeability transition dictates mitochondrial maturation upon switch in cellular identity of hematopoietic precursors

The mitochondrial permeability transition pore (mPTP) is a supramolecular channel that regulates exchange of solutes across cristae membranes, with executive roles in mitochondrial function and cell death. The contribution of the mPTP to normal physiology remains debated, although evidence implicates the mPTP in mitochondrial inner membrane remodeling in differentiating progenitor cells. Here, we demonstrate that strict control over mPTP conductance shapes metabolic machinery as cells transit toward hematopoietic identity. Cells undergoing the endothelial-to-hematopoietic transition (EHT) tightly control chief regulatory elements of the mPTP. During EHT, maturing arterial endothelium restricts mPTP activity just prior to hematopoietic commitment. After transition in cellular identity, mPTP conductance is restored. In utero treatment with NIM811, a molecule that blocks sensitization of the mPTP to opening by Cyclophilin D (CypD), amplifies oxidative phosphorylation (OXPHOS) in hematopoietic precursors and increases hematopoiesis in the embryo. Additionally, differentiating pluripotent stem cells (PSCs) acquire greater organization of mitochondrial cristae and hematopoietic activity following knockdown of the CypD gene, Ppif. Conversely, knockdown of Opa1, a GTPase critical for proper cristae architecture, induces cristae irregularity and impairs hematopoiesis. These data elucidate a mechanism that regulates mitochondrial maturation in hematopoietic precursors and underscore a role for the mPTP in the acquisition of hematopoietic fate.

Protective effects of mesenchymal stem cells-derived extracellular vesicles against ischemia-reperfusion injury of hearts donated after circulatory death: Preliminary study in a pig model

IntroductionInsufficient supply of cardiac grafts represents a severe obstacle in heart transplantation. Donation after Circulatory Death (DCD), in addition to conventional donation after brain death, is one promising option to overcome the organ shortage. However, DCD organs undergo an inevitable more extended period of warm unprotected ischemia between circulatory arrest and graft procurement. Mesenchymal stromal cell-derived extracellular vesicles (MSC-EVs) have shown remarkable protective effects against ischemia-reperfusion injury. Thus, we aimed to enhance grafts preservation from DCD donors, through treatment with MSC-EVs. MethodsFemale pigs were euthanized by barbiturate overdose and after 20 min of a flat EKG, the chest was opened, the heart harvested and subsequently connected to an extracorporeal perfusion machine. MSC-EVs, isolated by ion exchange chromatography, were added to the perfusion solution (1×1011 particles) and the heart was perfused for 2 h. Then, heart tissue biopsies were taken to assess histological changes, mitochondrial morphology, antioxidant enzyme activity and inflammation mediators’ expression. Biochemical parameters of myocardial viability were assessed in the perfusate. ResultsThe treatment with MSC-EVs significantly prevented mitochondria swelling, mitochondrial cristae loss and oxidative stress in cardiac tissue. The protective effect of MSC-EVs was confirmed by the delayed increase of the cardiac-specific enzymes CK and TnC in the perfusate and the reduction of caspase-3+ cells in tissue sections. ConclusionMSC-EVs improve graft quality by preserving the mitochondrial ultrastructure protecting the myocardium against oxidative stress, reducing apoptosis of cardiac cells and preventing the increase of pro-inflammatory cytokines.

Abstract Tu107: Decreased Mitochondrial Protein Expression Accompanies Atrial Hypertrophy but Precedes Atrial Fibrillation Onset in a Mouse Model of Spontaneous Atrial Fibrillation

Introduction: Atrial fibrillation (AF), characterized by episodes of rapid and irregular heart rate, increases left atrial (LA) demand for ATP. While the heart is metabolically flexible, able to use fatty acids, lactate, glucose, ketones, and amino acids to generate ATP, fatty acids are the primary fuel source. AF is associated with altered mitochondrial structure and function, but the role of energetic pathways in early stages of AF progression is not clear. Hypothesis: Decreased expression of mitochondrial proteins contributes to AF progression. Aim: Evaluate expression of proteins that influence mitochondrial structure and energetic capacity in AF onset and progression. Methods: AF burden was assessed weekly by electrocardiography (ECG) in male wild-type (WT) and cAMP responsive element modulator IbΔC-X heterozygous mice (CREM) (n=3). LA (4 - 9 mg) was obtained at 7 weeks and 16 weeks. Global proteomics was conducted, and 139,316 peptides were identified that mapped to 6000 robustly quantified proteins. Results: LA hypertrophy was evident in 7-week and 16-week CREM mice (Figure 1). We detected 2408 proteins differentially expressed in CREM compared to WT. At 7 weeks, Hallmark pathways adipogenesis, fatty acid metabolism, oxidative phosphorylation, and myogenesis were decreased in CREM (Table 1). These changes persisted at 16 weeks. We identified 740 differentially expressed mitochondrial proteins using MitoCarta 3.0 (552 decreased, 188 increased). Mitochondrial pathways associated with energy production, including branched chain amino acid metabolism, fatty acid transport and oxidation, carbohydrate metabolism, and tricarboxylic acid cycle were decreased (Table 1). Mitochondrial cristae organizing system (MICOS) and mitophagy, pathways that influence mitochondrial structure and content, were decreased. Terminal ECGs confirmed AF in 16-week CREM mice only. The coefficient of variation of RR intervals (CVRR), a measure of heart rate variability, was increased in 16-week CREM (Figure 2). These data suggest that decreased expression of mitochondrial proteins accompanies LA hypertrophy but precedes onset of AF. Conclusions: Decreased expression of proteins that regulate mitochondrial structure and energy production prior to AF onset suggests that these proteins play an important role in early LA changes that promote AF onset and progression. Future studies will examine how these changes in protein expression relate to alterations in atrial energy homeostasis.

External pressure induces the dysfunction of spermatogonia via triggering the intrinsic pathway of apoptosis.

Cryptorchidism, the failure of testes to descend into the scrotum, exposes the testes to higher temperature and external pressure. Scholars from Razi University found through research conducted at different pressure gradients (0, 25, 50, and 100 mmHg) and time gradients (2 and 4 h) that high hydrostatic pressure may lead to sperm apoptosis. In this work, we investigated the effect of external pressure on spermatogonia, exploring a new mechanism of male infertility caused by cryptorchidism. Various pressure gradients (0, 25, 50, and 100 mmHg) were applied to spermatogonia for different durations (0, 2, and 4 h) in the Cell Counting Kit-8 (CCK8) experiment. Morphological changes, cell ultrastructure, apoptosis rates, and the expression of apoptosis-related proteins (bax, bcl-2, caspase-3, and caspase-9) were assessed through immunofluorescence, electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay, flow cytometry, immunohistochemistry, real-time quantitative polymerase chain reaction (qPCR), and western blot. The cell viability assay showed that higher external pressure had a greater negative time-dependent impact on cell viability. Immunofluorescence results indicated that external pressure stimuli altered the morphology of spermatogonia. The results of TUNEL assay and flow cytometry demonstrated that external pressure stimuli induced apoptosis in spermatogonia. Transmission electron microscopy (TEM) observations showed the generation of apoptotic bodies, mitochondrial swelling, vacuolization, and mitochondrial cristae fusion. The results of immunohistochemistry indicated that pressure induced the expression of caspase-3 and caspase-9 proteins. qPCR and western blot analyses revealed an increased ratio of bax/bcl-2 and expression of caspase-3 and caspase-9. Methazolamide (cytochrome C inhibitor) blocked the pressure-induced cell apoptosis and inhibited the activation of caspase-3 while Z-IETD-FMK (caspase-8 inhibitor) did not. External pressure promotes spermatogonia apoptosis through the intrinsic apoptosis pathway, which may be one of the mechanisms of male infertility induced by cryptorchidism.

Inducible RPE-specific GPX4 knockout causes oxidative stress and retinal degeneration with features of age-related macular degeneration

Age-related macular degeneration (AMD) is one of the leading causes of vision loss in the elderly. This disease involves oxidative stress burden in the retina leading to death of retinal pigment epithelial (RPE) cells and photoreceptors. The retina is susceptible to oxidative stress, in part due to high metabolic activity and high concentration of polyunsaturated fatty acids that undergo lipid peroxidation chain reactions. Antioxidant enzymes exist in the retina to combat this stress, including glutathione peroxidase 4 (GPX4). GPX4 specifically reduces oxidized lipids, protecting against lipid peroxidation-induced oxidative stress, which is noted in dry AMD. We hypothesize that Gpx4 knockout within the RPE will result in an environment of chronic oxidative stress yielding degeneration akin to AMD. C57BL/6J mice with a floxed Gpx4 gene were mated with Rpe65Cre/ER mice. Offspring containing Rpe65Cre ± alleles and either Gpx4 WT or Gpx4 fl/fl alleles were administered tamoxifen to induce Gpx4 knockout in Gpx4 fl/fl mice. At sequential timepoints, retinal phenotypes were assessed via in vivo imaging utilizing confocal scanning laser ophthalmoscopy and optical coherence tomography (OCT), and visual function was probed by electroretinography. Retinas were studied post-mortem by immunohistochemical analyses, electron microscopy, plastic sectioning, and quantitative polymerase chain reaction and Western analyses. The RPE-specific Gpx4 knockout model was validated via Western analysis indicating diminished GPX4 protein only within the RPE and not the neural retina. Following Gpx4 knockout, RPE cells became dysfunctional and died, with significant cell loss occurring 2 weeks post-knockout. Progressive thinning of the photoreceptor layer followed RPE degeneration and was accompanied by loss of visual function. OCT and light microscopy showed hyperreflective foci and enlarged, pigmented cells in and above the RPE layer. Electron microscopy revealed decreased mitochondrial cristae and loss of basal and apical RPE ultrastructure. Finally, there was increased carboxyethylpyrrole staining, indicating oxidation of docosahexaenoic acid, and increased levels of mRNAs encoding oxidative stress-associated genes in the RPE and photoreceptors. Overall, we show that RPE-localized GPX4 is necessary for the health of the RPE and outer retina, and that knockout recapitulates phenotypes of dry AMD.

The evolving process of ferroptosis in thyroid cancer: Novel mechanisms and opportunities.

Thyroid cancer (TC) is a prevalent endocrine malignancy, with a significant increase in incidence worldwide. Ferroptosis is a novel form of programmed cell death, primarily caused by iron overload and reactive oxygen species (ROS)-dependent accumulation of lipid peroxides. The main manifestations of cellular ferroptosis are rupture of the outer membrane, crumpling of the mitochondria and shrinkage or disappearance of the mitochondrial cristae, thus leading to cell death. Ferroptosis is an important phenomenon in tumour progression, with crosstalk with tumour-associated signalling pathways profoundly affecting tumour progression, immune effects and treatment outcomes. The functions and mechanisms of ferroptosis in TC have also attracted increasing attention, mainly in terms of influencing tumour proliferation, invasion, migration, immune response, therapeutic susceptibility and genetic susceptibility. However, at present, the tumour biology of the morphological, biological and mechanism pathways of ferroptosis is much less deep in TC than in other malignancies. Hence, in this review, we highlighted the emerging role of ferroptosis in TC progression, including the novel mechanisms and potential opportunities for diagnosis and treatment, as well as discussed the limitations and prospects. Ferroptosis-based diagnostic and therapeutic strategies can potentially provide complementary management of TCs.

Preoperative Increases in T2-Weighted Fat-Suppressed Magnetic Resonance Imaging Signal Intensities Associated With Advanced Tissue Degeneration and Mitochondrial Dysfunction in Rotator Cuff Tears

PurposeThe purpose of this study was to investigate the relationship between magnetic resonance imaging (MRI) signal intensities and mitochondrial function in patients undergoing arthroscopic rotator cuff repair, assessed through histological and genetic profiling of tendon tissue. MethodsThis study, conducted between April 2022 and January 2023, included 20 patients undergoing rotator cuff repair for atraumatic/degenerative tears. Rotator cuff tendon edge samples were obtained during arthroscopic rotator cuff repair. Patients were classified based on signal intensity from preoperative T2-weighted fat suppressed MRI. Specifically, they were categorized as having either high or low signal intensity at the rotator cuff tendon edge, with the deltoid muscle serving as a reference. Comparative analyses specifically compared the histological features and genetic profiles of the tendon tissue at the rotator cuff tendon edge. Histological evaluation of harvested tendon specimens during arthroscopic rotator cuff repair employed the modified Bonar score. Real-time polymerase chain reaction was used to assess expression of various mitochondrial and apoptosis-related genes. The mitochondrial morphology of the rotator cuff torn site was examined using electron microscopy. ResultsThe higher signal intensity group showed significantly higher modified Bonar scores (p=0.0068), decreased mitochondrial gene expression, increased TUNEL-positive cells (p=0.032), lower SOD activity (p=0.011), reduced ATP5A (p=0.031), and increased cleaved caspase-9 activity (p=0.026) when compared to the lower signal intensity group. Electron microscopy revealed fewer mitochondrial cristae in the higher signal intensity group. ConclusionOur results suggest correlations between high MRI signal intensities and the presence of degeneration, mitochondrial dysfunction, and increased apoptosis in rotator cuff tissues. This underscores the utility of MRI signal intensity as an indicator of tissue condition. Clinical RelevanceBuilding on the established premise that elevated preoperative MRI signal intensities may indicate higher rates of postoperative rotator cuff re-tears, the current study substantiates these findings from a mitochondrial function perspective.