Mitochondrial Morphology Research Articles

Inhibition of calpain reduces oxidative stress and attenuates pyroptosis and ferroptosis in Clostridium perfringens Beta-1 toxin-induced macrophages

Clostridium perfringens Beta-1 toxin (CPB1) is a lethal toxin, which can lead to necrotic enteritis, but the pathological mechanism has not been elucidated. We investigated whether reactive oxygen species (ROS) participated in CPB1-induced pyroptosis and ferroptosis, and investigated the effects of calpain on CPB1-induced oxidative stress and inflammation. Scavenging ROS by N-acetyl cysteine (NAC) led to the reduction of ROS, inhibited the death of macrophages, cytoplasmic swelling and membrane rupture, the expression of pyroptosis-related proteins and proinflammatory factor, while increased the expression of anti-inflammatory factors in cells treated with rCPB1. Adenosine triphosphate (ATP) synthase, H+ transporting, mitochondrial F1 complex, alpha subunit 1 (ATP5A1) was identified specifically interact with rCPB1. Silencing ATP5A1 inhibited accumulation of ATP and ROS, leaded to less cytoplasmic swelling and membrane rupture, attenuated pyroptosis and inflammation in rCPB1-treated cells. We also found that rCPB1 induces ferroptosis in macrophages, and the level of ferroptosis was similar with H2O2. Of note, H2O2 is a major ROS source, indicated that ROS production may play a major role in the regulation of ferroptosis in macrophages treated with rCPB1. This finding was further corroborated in rCPB1- induced human acute monocytic leukemia cells, which were treated with NAC. In addition, the inhibition of ferroptosis using liproxstatin-1 inhibited the shriveled mitochondrial morphology, increased the expression of glutathione peroxidase 4, nicotinamide adenine dinucleotide (phosphate) hydrogen: quinone oxidoreductase 1 and cysteine/glutamic acid reverse transport solute carrier family 7 members 11, decreased the expression of heme oxygenase 1, nuclear receptor coactivator 4 and transferrin receptor proteins, reduced malondialdehyde and lipid peroxidation levels, and increased intracellular L-glutathione levels in cells treated with rCPB1. Furthermore, calpain inhibitor PD151746 was used to investigate how pyroptosis and ferroptosis were involved simultaneously in rCPB1-treated macrophages. We showed that PD151746 inhibited ATP and ROS production, reversed the representative pyroptosis/ferroptosis indicators and subsequently reduced inflammation. The above findings indicate that rCPB1 might lead to macrophage pyroptosis and ferroptosis through the large and sustained increase in intracellular calpain and oxidative stress, further lead to inflammation.

SENP1 prevents high fat diet-induced non-alcoholic fatty liver diseases by regulating mitochondrial dynamics

Mitochondrial dynamics plays a crucial role in the occurrence and development of non-alcoholic fatty liver diseases (NAFLD). SENP1, a SUMO-specific protease, catalyzes protein de-SUMOylation and involves in various physiological and pathological processes. However, the exact role of SENP1 in NAFLD remains unclear. Therefore, we investigated the regulatory role of SENP1 in mitochondrial dynamics during the progression of NAFLD. In the study, the NAFLD in vivo model induced by high fat diet (HFD) and in vitro model induced by free fatty acids (FFA) were established to investigate the role and underlying mechanism of SENP1 through detecting mitochondrial morphology and dynamics. Our results showed that the down-regulation of SENP1 expression and the mitochondrial dynamics dysregulation occurred in the NAFLD, evidenced as mitochondrial fragmentation, up-regulation of p-Drp1 ser616 and down-regulation of MFN2, OPA1. However, over-expression of SENP1 significantly alleviated the NAFLD, rectified the mitochondrial dynamics disorder, reduced Cyt-c release and ROS levels induced by FFA or HFD; moreover, the over-expression of SENP1 also reduced the SUMOylation levels of Drp1 and prevented the Drp1 translocation to mitochondria. Our findings suggest that the possible mechanisms of SENP1 were through rectifying the mitochondrial dynamics disorder, reducing Cyt-c release and ROS-mediated oxidative stress. The findings would provide a novel target for the prevention and treatment of NALFD.

Effects of glial-derived neurotrophic factor on remodeling and mitochondrial function in human airway smooth muscle cells.

Airway smooth muscle (ASM) cells play important roles in airway remodeling of asthma. Our previous studies show that in vivo administration of glial-derived neurotrophic factor (GDNF) in mice induces thickening and collagen deposition in bronchial airways, whereas chelation of GDNF by GFRα1-Fc attenuates airway remodeling in the context of allergen exposure. To determine whether GDNF has direct effects on ASM, in this study, we examined GDNF in ASM cells from normal versus asthmatic humans. We found that GDNF treatment of human ASM cells had only minor effects on cell proliferation and migration, intracellular expression or extracellular deposition of collagen I (COL1), collagen III (COL3), and fibronectin. Endoplasmic reticulum (ER) stress response and mitochondrial function have been implicated in asthma. We investigated whether GDNF regulates these aspects in human ASM. We found that GDNF treatment did not affect ER stress protein expression in normal or asthmatic cells. However, GDNF treatment impaired mitochondrial morphology in ASM but without significant effects on mitochondrial respiration. Thus, it is likely that in vivo effects of GDNF on airway remodeling per se involve cell types other than those on ASM, and thus ASM may serve more as a source of GDNF rather than a target.NEW & NOTEWORTHY Our previous study suggests that glial-derived neurotrophic factor (GDNF) is involved in allergen-induced airway hyperreactivity and remodeling in vivo. Here, we show that GDNF has no direct effects in remodeling of human airway smooth muscle (ASM) but GDNF dysregulates mitochondrial morphology in human ASM in the context of asthma.

Effects of normal mitochondrial transplantation on proliferation, apoptosis and stemness of triple-negative breast cancer cells

Objectives: To observe the mitochondrial morphology of normal and triple-negative breast cancer cells, extract mitochondria from normal cells, and investigate the effects of mitochondrial transplantation on proliferation, apoptosis, and stemness of triple-negative breast cancer cells. Methods: The morphology of mitochondria was observed by transmission electron microscope. Mitochondria were extracted by mitochondrial extraction kit, mitochondrial protein was identified by western blot, and mitochondrial activity was detected by mitochondrial membrane potential detection kit. MitoTracker Green or MitoTracker Deep Red fluorescent probes were used to label the mitochondria of living cells, and the degree of mitochondria entering LTT cells was observed by confocal laser microscopy at 12, 24, and 96 hours. The effects of mitochondrial transplantation on proliferation, apoptosis, and stemness of breast cancer cells were examined by CCK8, colony formation assay, flow cytometry, and sphere formation assay after 24 hours of mitochondrial transplantation. Results: The mitochondria of normal cells were rod-shaped or elongated, while the mitochondria of triple-negative breast cancer cells were swollen and vacuolated. Western blot results showed that cytochrome c oxidase subunit I (MT-CO1) protein encoded by mitochondria was present in the isolated mitochondria. The content of heat shock protein 60 (HSP60) was higher in mitochondria than that in cytoplasm. The result of the multi-mode microplate reader showed that the content of mitochondrial J-aggregates/monomer was 1.67±0.06, which was significantly higher than 0.35±0.04 of the control group (P＜0.001). Exogenous mitochondria were observed in LTT cells at 12, 24, and 96 hours after mitochondrial transplantation. The results of the CCK8 experiment showed that OD450 of LTT cells was 0.27±0.13 after 48 hours transplantation, which was lower than 0.62±0.36 of the control group (P=0.023). The OD450 of MDA-MB-468 cells was 0.30±0.03, which was lower than 0.65±0.10 of the control group (P=0.004). After 120 hours of mitochondrial transplantation, OD450 in both groups was still significantly lower than that in the control group (P＜0.01). The number of clones formed by mitochondrial transplantation of LTT cells was 21.33±7.31, which was lower than 35.22±13.59 of the control group (P=0.016). Flow cytometry showed that the early apoptosis rate of LTT cells was (30.07±2.15)% after 24 hours of mitochondrial transplantation, which was higher than 2.07±1.58 of the control group (P＜0.001). The proportion of early apoptosis in MDA-MB-468 cells was 24.47%±5.22%, which was higher than (7.83±2.06)% in the control group (P=0.007). In addition, the number of mitochondria transplanted LTT cells into the cell sphere was 46.25±5.40, which was significantly lower than 62.58±6.43 of the control group (P＜0.001). Conclusion: Normal mitochondria can enter triple-negative breast cancer cells by co-culture, inhibit the proliferation and stemness of triple-negative breast cancer cells, and promote the apoptosis of triple-negative breast cancer cells.

Mitochondrial Dynamics and mRNA Translation: A Local Synaptic Tale.

Mitochondria are dynamic organelles that can adjust and respond to different stimuli within a cell. This plastic ability allows them to effectively coordinate several cellular functions in cells and becomes particularly relevant in highly complex cells such as neurons. An imbalance in mitochondrial dynamics can disrupt mitochondrial function, leading to abnormal cellular function and ultimately to a range of diseases, including neurodegenerative disorders. Regulation of mRNA transport and local translation inside neurons is crucial for maintaining the proteome of distal mitochondria, which is vital for energy production and synaptic function. A significant portion of the axonal transcriptome is dedicated to mRNAs for mitochondrial proteins, emphasizing the importance of local translation in sustaining mitochondrial function in areas far from the cell body. In neurons, local translation and the regulation of mRNAs encoding mitochondrial-shaping proteins could be essential for synaptic plasticity and neuronal health. The dynamics of these mRNAs, including their transport and local translation, may influence the morphology and function of mitochondria, thereby affecting the overall energy status and responsiveness of synapses. Comprehending the mitochondria-related mRNA regulation and local translation, as well as its influence on mitochondrial morphology near the synapses will help to better understand neuronal physiology and neurological diseases where mitochondrial dysfunction and impaired synaptic plasticity play a central role.

6-Gingerol improves lipid metabolism disorders in skeletal muscle by regulating AdipoR1/AMPK signaling pathway

BackgroundTo delve into the precise mechanisms by which 6-gingerol ameliorates lipid metabolism disorders in skeletal muscle. MethodsThe level of triglycerides (TG) was used to evaluate lipid deposition. In skeletal muscle, transmission electron microscopy (TEM) was employed to observe mitochondrial morphology. Additionally, PCR was applied to detect mitochondrial biogenesis, and levels of malondialdehyde (MDA), catalase (CAT), glutathione, r-glutamyl cysteingl+glycine (GSH) and nicotinamide adenine dinucleotide (NADH) were measured to assess mitochondrial oxidative stress levels. In vivo, flow cytometry and immunofluorescence assays were conducted to quantify reactive oxygen species (ROS) and mitochondrial membrane potential (MMP). Furthermore, the Seahorse XF assays was utilized to assess mitochondrial respiratory capacity. Fluorescence confocal microscopy and molecular docking were applied to analyze the binding of 6-gingerol and adiponectin receptor 1 (AdipoR1). The expression of AdipoR1, AMPK, PGC-1α and SIRT1 were detected by Western Blot. ResultsIn vivo, 6-gingerol could reduce body weight in mice induced by a high-fat diet, enhance metabolic profiles in plasma, decrease lipid accumulation in skeletal muscle and liver, and elevate adiponectin levels. In skeletal muscle, it could restore mitochondrial morphology, boost mitochondrial copy number and biogenesis, and mitigate oxidative stress. In vitro, 6-gingerol may directly interact with AdipoR1 to upregulate the expression of downstream proteins p-AMPK, SIRT1, and PGC-1α, leading to a reduction in lipid deposition, a decrease in ROS production, an increase in mitochondrial membrane potential, and an enhancement of mitochondrial respiratory capacity in C2C12 myotubes. Conclusion6-Gingerol ameliorated lipid metabolism in skeletal muscle by regulating the AdipoR1/AMPK signaling pathway.

LRRK2G2019S Gene Mutation Causes Skeletal Muscle Impairment in Animal Model of Parkinson's Disease

ABSTRACTBackgroundWhile the gradually aggravated motor and non‐motor disorders of Parkinson's disease (PD) lead to progressive disability and frequent falling, skeletal muscle impairment may contribute to this condition. The leucine‐rich repeat kinase2 (LRRK2) is a common disease‐causing gene in PD. Little is known about its role in skeletal muscle impairment and its underlying mechanisms.MethodsTo investigate whether the mutation in LRRK2 causes skeletal muscle impairment, we used 3‐month‐old (3mo) and 14‐month‐old (14mo) LRRK2G2019S transgenic (TG) mice as a model of PD, compared with the age‐matched littermate wild‐type (WT) controls. We measured the muscle mass and strength, ultrastructure, inflammatory infiltration, mitochondrial morphology and dynamics dysfunction through behavioural analysis, electromyography (EMG), immunostaining, transmission electron microscopy (TEM) and other molecular biology techniques.ResultsThe 3mo‐TG mice display mild skeletal muscle impairment with spontaneous potentials in EMG (increased by 130%, p < 0.05), myofibre necrosis (p < 0.05) and myosin heavy chain‐II changes (reduced by 19%, p < 0.01). The inflammatory cells and macrophage infiltration are significantly increased (CD8a+ and CD68+ cells up 1060% and 579%, respectively, both p < 0.0001) compared with the WT mice. All of the above pathogenic processes are aggravated by aging. The 14mo‐TG mice EMG examinations show a reduced duration (by 31%, p < 0.01) and increased polyphasic waves of motor unit action potentials (by 28%, p < 0.05). The 14mo‐TG mice present motor behavioural deficits (p < 0.05), muscle strength and mass reduction by 37% and 8% (p < 0.05 and p < 0.01, respectively). A remarkable increase in inflammatory infiltration is accompanied by pro‐inflammatory cytokines in the skeletal muscles. TEM analysis shows muscle fibre regeneration with the reduced length of sarcomeres (by 6%;p < 0.05). The muscle regeneration is activated as Pax7+ cells increased by 106% (p < 0.0001), andmyoblast determination protein elevated by 71% (p < 0.01). We also document the morphological changes and dynamics dysfunction of mitochondria with the increase of mitofusin1 by 43% (p < 0.05) and voltage‐dependent anion channel 1 by 115% (p < 0.001) in the skeletal muscles of 14mo‐TG mice.ConclusionsTaken together, these findings may provide new insights into the clinical and pathogenic involvement of LRRK2G2019 mutation in muscles, suggesting that the diseases may affect not only midbrain dopaminergic neurons, but also other tissues, and it may help overall clinical management of this devastating disease.

Ferroptosis Integrates Mitochondrial Derangements and Pathological Inflammation to Promote Pulmonary Hypertension.

Mitochondrial dysfunction, characterized by impaired lipid metabolism and heightened reactive oxygen species (ROS) generation, results in lipid peroxidation and ferroptosis. Ferroptosis is an inflammatory mode of cell death that promotes complement activation and macrophage recruitment. In pulmonary arterial hypertension (PAH), pulmonary arterial endothelial cells (PAEC) exhibit cellular phenotypes that promote ferroptosis. Moreover, there is ectopic complement deposition and inflammatory macrophage accumulation in the pulmonary vasculature. However, the effects of ferroptosis inhibition on these pathogenic mechanisms and the cellular landscape of the pulmonary vasculature are incompletely defined. Multi-omics and physiological analyses evaluated how ferroptosis inhibition modulated preclinical PAH. The impact of AAV1-mediated expression of the pro-ferroptotic protein ACSL4 on PAH was determined, and a genetic association study in humans further probed the relationship between ferroptosis and pulmonary hypertension (PH). Ferrostatin-1, a small-molecule ferroptosis inhibitor, mitigated PAH severity in monocrotaline rats. RNA-seq and proteomics analyses demonstrated ferroptosis was associated with PAH severity. RNA-seq, proteomics, and confocal microscopy revealed complement activation and pro-inflammatory cytokines/chemokines were suppressed by ferrostatin-1. Additionally, ferrostatin-1 combatted changes in endothelial, smooth muscle, and interstitial macrophage abundance and gene activation patterns as revealed by deconvolution RNA-seq. Ferroptotic PAEC damage associated molecular patterns restructured the transcriptomic signature, mitochondrial morphology, and promoted proliferation of pulmonary artery smooth muscle cells, and created a pro-inflammatory phenotype in monocytes in vitro. AAV1-Acsl4 induced an inflammatory PAH phenotype in rats. Finally, single-nucleotide polymorphisms in six ferroptosis genes identified a potential link between ferroptosis and PH severity in the Vanderbilt BioVU repository. Ferroptosis promotes PAH through metabolic and inflammatory mechanisms in the pulmonary vasculature.

Mitochondrial membrane potential and oxidative stress interact to regulate Oma1-dependent processing of Opa1 and mitochondrial dynamics.

Mitochondrial form and function are regulated by the opposing forces of mitochondrial dynamics: fission and fusion. Mitochondrial dynamics are highly active and consequential during neuronal ischemia/reperfusion (I/R) injury. Mitochondrial fusion is executed at the mitochondrial inner membrane by Opa1. The balance of long (L-Opa1) and proteolytically cleaved short (S-Opa1) isoforms is critical for efficient fusion. Oma1 is the predominant stress-responsive protease for Opa1 processing. In neuronal cell models, we assessed Oma1 and Opa1 regulation during mitochondrial stress. In an immortalized mouse hippocampal neuron line (HT22), Oma1 was sensitive to mitochondrial membrane potential depolarization (rotenone, FCCP) and hyperpolarization (oligomycin). Further, oxidative stress was sufficient to increase Oma1 activity and necessary for depolarization-induced proteolysis. We generated Oma1 knockout (KO) HT22 cells that displayed normal mitochondrial morphology and fusion capabilities. FCCP-induced mitochondrial fragmentation was exacerbated in Oma1 KO cells. However, Oma1 KO cells were better equipped to perform restorative fusion after fragmentation, presumably due to preserved L-Opa1. We extended our investigations to a combinatorial stress of neuronal oxygen-glucose deprivation and reoxygenation (OGD/R), where we found that Opa1 processing and Oma1 activation were initiated during OGD in an ROS-dependent manner. These findings highlight a novel dependence of Oma1 on oxidative stress in response to depolarization. Further, we demonstrate contrasting fission/fusion roles for Oma1 in the acute response and recovery stages of mitochondrial stress. Collectively, our results add intersectionality and nuance to the previously proposed models of Oma1 activity.

Hederagenin inhibits mitochondrial damage in Parkinson’s disease via mitophagy induction

BackgroundParkinson's disease (PD) is a neurodegenerative disorder marked by the loss of dopaminergic neurons and the formation of α-synuclein aggregates. Mitochondrial dysfunction and oxidative stress are pivotal in PD pathogenesis, with impaired mitophagy contributing to the accumulation of mitochondrial damage. Hederagenin (Hed), a natural triterpenoid, has shown potential neuroprotective effects; however, its mechanisms of action in PD models are not fully understood. MethodWe investigated the effects of Hed on 6-hydroxydopamine (6-OHDA)-induced cytotoxicity in SH-SY5Y cells by assessing cell viability, mitochondrial function, and oxidative stress markers. Mitophagy induction was evaluated using autophagy and mitophagy inhibitors and fluorescent staining techniques. Additionally, transgenic Caenorhabditis elegans (C. elegans) models of PD were used to validate the neuroprotective effects of Hed in vivo by focusing on α-synuclein aggregation, mobility, and dopaminergic neuron integrity. ResultsHed significantly enhanced cell viability in 6-OHDA-treated SH-SY5Y cells by inhibiting cell death and reducing oxidative stress. It ameliorated mitochondrial damage, evidenced by decreased mitochondrial superoxide production, restored membrane potential, and improved mitochondrial morphology. Hed also induced mitophagy, as shown by increased autophagosome formation and reduced oxidative stress; these effects were diminished by autophagy and mitophagy inhibitors. In C. elegans models, Hed activated mitophagy and reduced α-synuclein aggregation, improved mobility, and mitigated the loss of dopaminergic neurons. RNA interference targeting the mitophagy-related genes pdr-1 and pink-1 partially reversed these benefits, underscoring the role of mitophagy in Hed's neuroprotective actions. ConclusionHed exhibits significant neuroprotective effects in both in vitro and in vivo PD models by enhancing mitophagy, reducing oxidative stress, and mitigating mitochondrial dysfunction. These findings suggest that Hed holds promise as a therapeutic agent for PD, offering new avenues for future research and potential drug development.

Exercise Alleviates Cardiovascular Diseases by Improving Mitochondrial Homeostasis.

Engaging in regular exercise and physical activity contributes to delaying the onset of cardiovascular diseases (CVDs). However, the physiological mechanisms underlying the benefits of regular exercise or physical activity in CVDs remain unclear. The disruption of mitochondrial homeostasis is implicated in the pathological process of CVDs. Exercise training effectively delays the onset and progression of CVDs by significantly ameliorating the disruption of mitochondrial homeostasis. This includes improving mitochondrial biogenesis, increasing mitochondrial fusion, decreasing mitochondrial fission, promoting mitophagy, and mitigating mitochondrial morphology and function. This review provides a comprehensive overview of the benefits of physical exercise in the context of CVDs, establishing a connection between the disruption of mitochondrial homeostasis and the onset of these conditions. Through a detailed examination of the underlying molecular mechanisms within mitochondria, the study illuminates how exercise can provide innovative perspectives for future therapies for CVDs.

4-Octyl itaconate attenuates renal tubular injury in db/db mice by activating Nrf2 and promoting PGC-1α-mediated mitochondrial biogenesis

Objectives: The aim of this study was to investigate the mechanism of itaconate’s potential effect in diabetic kidney disease. Methods: Renal immune responsive gene 1 (IRG1) levels were measured in db/db mice and streptozotocin (STZ) + high-fat diet (HFD)-induced diabetic mice. Irg1 knockout mice were generated. db/db mice were treated with 4-octyl itaconate (4-OI, 50 mg/kg), a derivative of itaconate, for 4 weeks. Renal function and morphological changes were investigated. Ultrastructural alterations were determined by transmission electron microscopy. Results: Renal IRG1 levels were reduced in two diabetic models. STZ+HFD-treated Irg1 knockout mice exhibited aggravated renal tubular injury and worsened renal function. Treatment with 4-OI lowered urinary albumin-to-creatinine ratio and blood urea nitrogen levels, and restored renal histological changes in db/db mice. It improved mitochondrial damage, increased expressions of peroxisome-proliferator-activated receptor γ coactivator-1α (PGC-1α) and mitochondrial transcription factor A (TFAM) in the renal cortex of db/db mice. These were confirmed in vitro; 4-OI improved high glucose-induced abnormal mitochondrial morphology and TFAM expression in HK-2 cells, effects that were inhibited by PGC-1α silencing. Moreover, 4-OI reduced the number of apoptotic cells in the renal cortex of db/db mice. Further study showed that 4-OI increased renal Nrf2 expression and decreased oxidative stress levels in db/db mice. In HK-2 cells, 4-OI decreased high glucose-induced mitochondrial ROS production, which was reversed by Nrf2 silencing. Nrf2 depletion also inhibited 4-OI-mediated regulation of PGC-1α, TFAM, and mitochondrial apoptotic protein expressions. Conclusions: 4-OI attenuates renal tubular injury in db/db mice by activating Nrf2 and promoting PGC-1α-mediated mitochondrial biogenesis.

Alternative splicing of Mff regulates AMPK-mediated phosphorylation, mitochondrial fission and antiviral response

Mitochondrial morphology and function change dynamically in response to intracellular signaling and the surrounding environment. The mitochondrial fission factor Mff, which localizes to the outer mitochondrial membrane, mediates not only mitochondrial fission by recruiting the dynamin-related GTPase Drp1 to mitochondrial fission sites but also the double-stranded RNA-induced antiviral response on mitochondria through mitochondrial antiviral signaling (MAVS). Mff is reported to be regulated by AMP-activated protein kinase (AMPK)-mediated protein phosphorylation and alternative pre-mRNA splicing; however, the relationships among RNA splicing, phosphorylation, and multiple functions of Mff have not been fully understood. Here, we showed that mouse Mff has a tissue-specific splicing pattern, and at least eight Mff splice isoforms were expressed in mouse embryonic fibroblasts (MEFs). We introduced single Mff isoforms into Mff knockout MEFs and found that insertion of exon 6 just after the phosphorylation site, by the alternative splicing, reduced its phosphorylation by AMPK and its functions in mitochondrial fission and the antiviral response. In addition, the underlying mechanism repressing these functions was independent of phosphorylation. These results indicate that multiple functions of Mff on mitochondria are regulated by AMPK-mediated phosphorylation and alternative splicing, under the control of energy metabolism and cellular differentiation.

Adipose stem cell exosomes promote mitochondrial autophagy through the PI3K/AKT/mTOR pathway to alleviate keloids

BackgroundFibrosis with unrelieved chronic inflammation is an important pathological change in keloids. Mitochondrial autophagy plays a crucial role in reducing inflammation and inhibiting fibrosis. Adipose stem cell-derived exosomes, a product of adipose stem cell paracrine secretion, have pharmacological effects, such as anti-inflammatory and antiapoptotic effects, and mediate autophagy. Therefore, this study aims to investigate the function and mechanism of adipose stem cell exosomes in the treatment of keloids.MethodWe isolated adipose stem cell exosomes under normoxic and hypoxic condition to detect their effects on keloid fibroblast proliferation, migration, and collagen synthesis. Meanwhile, 740YPDGFR (PI3K/AKT activator) was applied to detect the changes in autophagic flow levels and mitochondrial morphology and function in keloid fibroblasts. We constructed a human keloid mouse model by transplanting human keloid tissues into six-week-old (20–22 g; female) BALB/c nude mice, meanwhile, we applied adipose stem cell exosomes to treat the mouse model and observed the retention and effect of ADSC exosomes in vivo.ResultsADSC exosomes can inhibit the PI3K/AKT/mTOR signaling pathway. The exosomes of ADSCs decreased the inflammatory level of KFs, enhanced the interaction between P62 and LC3, and restored the mitochondrial membrane potential. In the human keloid mouse model, ADSC exosomes can exist stably, promote mitochondrial autophagy in keloid tissue, improve mitochondrial morphology, reduce inflammatory reaction and fibrosis. Meanwhile, At the same time, the exosomes derived from hypoxic adipose stem cells have played a more effective role in both in vitro and in vivo experiments.ConclusionsAdipose stem cell exosomes inhibited the PI3K/AKT/mTOR pathway, activated mitochondrial autophagy, and alleviated keloid scars.Graphical

Characterization of fission and fusion mitochondrial dynamics in HD fibroblasts according to patient's severity status

Huntington's Disease (HD) is an inheritable neurodegenerative condition caused by an expanded CAG trinucleotide repeat in the HTT gene with a direct correlation between CAG repeats expansion and disease severity with earlier onset-of- disease. Previously we have shown that primary skin fibroblasts from HD patients exhibit unique phenotype disease features, including distinct nuclear morphology and perturbed actin cap linked with cell motility, that are correlated with the HD patient disease severity. Here we provide further evidence that mitochondrial fission-fusion morphology balance dynamics, classified using a custom image-based high-content analysis (HCA) machine learning tool, that improved correlation with HD severity status. This mitochondrial phenotype is supported by appropriate changes in fission-fusion biomarkers (Drp1, MFN1, MFN2, VAT1) levels in the HD patients' fibroblasts. These findings collectively point towards a dysregulation in mitochondrial dynamics, where both fission and fusion processes may be disrupted in HD cells compared to healthy controls. This study shows for the first time a methodology that enables identification of HD phenotype before patient's disease onset (Premanifest). Therefore, we believe that this tool holds a potential for improving precision in HD patient's diagnostics bearing the potential to evaluate alterations in mitochondrial dynamics throughout the progression of HD, offering valuable insights into the molecular mechanisms and drug therapy evaluation underlying biological differences in any disease stage.

Ferrostatin-1 ameliorates Cis-dichlorodiammineplatinum(II)-induced ovarian toxicity by inhibiting ferroptosis

Cis-dichlorodiammineplatinum(II) (CDDP), while widely utilized in tumor therapy, results in toxic side effects that patients find intolerable. The specific mechanism by which CDDP inflicts ovarian damage remains unclear. This study aimed to explore the involvement of ferrostatin-1 (FER-1) and ferroptosis in CDDP-induced ovarian toxicity. This study established models of CDDP-induced injury in granulosa cells (GCs) and rat model of premature ovarian failure (POF). CCK-8 assessed the effects of CDDP and FER-1 on GC viability. FerroOrange and Mito-FerroGreen, DCFH-DA and MitoSox-Red, Rhodamine 123 and Transmission electron microscopy (TEM) measured Fe2+, reactive oxygen species (ROS), mitochondrial membrane potential and the mitochondrial morphology in GC cells, respectively. Serum hormone levels; organ indices; malondialdehyde, superoxide dismutase, and glutathione analyses; and western blotting were performed to examine ferroptosis's role in vitro. Molecular docking simulation was evaluated the interaction between FER-1 and GPX4 or FER-1 and NRF2. Molecular docking simulations were conducted to evaluate the interactions between FER-1 and GPX4, as well as FER-1 and NRF2. The findings revealed that CDDP-induced ovarian toxicity involved iron accumulation, increased ROS accumulation, and mitochondrial dysfunction, leading to endocrine disruption and tissue damage in rats. These changes correlated with NRF2, HO-1, and GPX4 levels. However, FER-1 decreased the extent of ferroptosis. Thus, ferroptosis appears to be a crucial mechanism of CDDP-induced ovarian injury, with GPX4 as potential protective targets.

Subcellular localization and function analysis of PINK1 mitron in PD progression: Mitron modulates mitochondrial morphology to regulate neuronal death

Parkinson’s disease (PD) is a multi-factorial neurodegenerative disorder. Loss or degeneration of the dopaminergic neurons in the substantia nigra and development of Lewy Bodies in dopaminergic neurons were the defining pathologic changes. MiRNAs fine-tune the protein levels by post-transcriptional gene regulation. MiR-7019-3p is encoded within the 5th intron of PD associated protein PINK1. In present study, we firstly demonstrated miR-7019-3p expression is significantly up regulated in PD mice model and neuron cell models, miR-7019-3p mainly existed in mitochondria, miR-7019-3p could regulate the structure and function of mitochondria in neuronal cells. We predicted and verified that mitochondria associated protein OPA1 and 12s rRNA, 16s rRNA and polycistronic RNA are target genes of miR-7019-3p. Finally, we proved that SP1 protein could independently regulate the expression of miR-7019-3p at the upstream. The evidences in the study suggest the role miR-7019-3p in the regulation of mitochondrial structure and function, and this kind of regulation could be implemented or promoted through the pathway of SP1-miR-7019-3p-OPA1/12s rRNA, 16s rRNA and polycistronic RNA.Our results have suggested a promising and potential therapeutic target for reversing mitochondria dysregulation in neuronal cells during Parkinson's disease process.

Dysregulated Ca2+ signaling, fluid secretion, and mitochondrial function in a mouse model of early Sjögren's disease.

The molecular mechanisms leading to saliva secretion are largely established, but factors that underlie secretory hypofunction, specifically related to the autoimmune disease Sjögren's syndrome (SS) are not fully understood. A major conundrum is the lack of association between the severity of salivary gland immune cell infiltration and glandular hypofunction. SS-like disease was induced by treatment with DMXAA, a small molecule agonist of murine STING. We have previously shown that the extent of salivary secretion is correlated with the magnitude of intracellular Ca2+ signals (Takano et al., 2021). Contrary to our expectations, despite a significant reduction in fluid secretion, neural stimulation resulted in enhanced Ca2+ signals with altered spatiotemporal characteristics in vivo. Muscarinic stimulation resulted in reduced activation of the Ca2+-activated Cl- channel, TMEM16a, although there were no changes in channel abundance or absolute sensitivity to Ca2+. Super-resolution microscopy revealed a disruption in the colocalization of Inositol 1,4,5-trisphosphate receptor Ca2+ release channels with TMEM16a, and channel activation was reduced when intracellular Ca2+ buffering was increased. These data indicate altered local peripheral coupling between the channels. Appropriate Ca2+ signaling is also pivotal for mitochondrial morphology and bioenergetics. Disrupted mitochondrial morphology and reduced oxygen consumption rate were observed in DMXAA-treated animals. In summary, early in SS disease, dysregulated Ca2+ signals lead to decreased fluid secretion and disrupted mitochondrial function contributing to salivary gland hypofunction.

Endoplasmic reticulum-mitochondrial encounter structure regulates the mitochondrial morphology, DON biosynthesis and toxisome formation in Fusarium graminearum

The endoplasmic reticulum-mitochondrial encounter structure (ERMES) complex is known to play crucial roles in various cellular processes. However, its functional significance in filamentous fungi, particularly its impact on deoxynivalenol (DON) biosynthesis in Fusarium graminearum, remains inadequately understood. In this study, we aimed to investigate the regulatory function of the ERMES complex in F. graminearum. Our findings indicate significant changes in mitochondrial morphology of ERMES mutants, accompanied by decreased ATP content and ergosterol production. Notably, the toxisome formation in the ERMES mutant ΔFgMDM10 was defective, resulting in a substantial reduction in DON biosynthesis. This suggests a pivotal role of ERMES in toxisome formation, as evidenced by the pronounced inhibition of toxisome formation when ERMES was disrupted by boscalid. Furthermore, ERMES deficiencies were shown to diminish the virulence of F. graminearum towards host plants significantly. In conclusion, our results suggest ERMES is an important regulator of mitochondrial morphology, DON biosynthesis, and toxisome formation in F. graminearum.

Sex-dependent regulation of retinal pigment epithelium and retinal function by Pgc-1α.

Age-related macular degeneration (AMD) is a major cause of blindness that affects people over 60. While aging is the prominent factor in AMD, studies have reported a higher prevalence of AMD in women compared to age-matched men. Higher levels of the innate immune response's effector proteins complement factor B and factor I were also found in females compared to males in intermediate AMD. However, the mechanisms underlying these differences remain elusive. Peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) is a key regulator of mitochondrial biogenesis and metabolic pathways. Previously, we showed that Pgc-1α repression and high-fat diet induce drastic AMD-like phenotypes in mice. Our recent data revealed that Pgc-1α repression alone can also induce retinal pigment epithelium (RPE) and retinal dysfunction in mice, and its inhibition in vitro results in lipid droplet accumulation in human RPE. Whether sex is a contributing factor in these phenotypes remains to be elucidated. Using electroretinography, we demonstrate that sex could influence RPE function during aging independent of Pgc-1α in wild-type (WT) mice. We further show that Pgc-1α repression exacerbates RPE and retinal dysfunction in females compared to aged-match male mice. Gene expression analyses revealed that Pgc-1α differentially regulates genes related to antioxidant enzymes and mitochondrial dynamics in males and females. RPE flat mounts immunolabeled with TOMM20 and DRP1 indicated a sex-dependent role for Pgc-1α in regulating mitochondrial fission. Analyses of mitochondrial network morphology suggested sex-dependent effects of Pgc-1α repression on mitochondrial dynamics. Together, our study demonstrates that inhibition of Pgc-1α induces a sex-dependent decline in RPE and retinal function in mice. These observations on the sex-dependent regulation of RPE and retinal function could offer novel insights into targeted therapeutic approaches for age-related RPE and retinal degeneration.