Inhibition of dynamin-related protein 1-filamin interaction improves systemic glucose metabolism.

BACKGROUND AND PURPOSE: Maintaining mitochondrial quality is attracting attention as a new strategy to treat diabetes and diabetic complications. We previously reported that mitochondrial hyperfission by forming a protein complex between dynamin-related protein (Drp) 1 and filamin, mediates chronic heart failure and cilnidipine, initially developed as an L/N-type Ca2+ channel blocker, improves heart failure by inhibiting Drp1-filamin protein complex. We investigated whether cilnidipine improves hyperglycaemia of various diabetic mice models. EXPERIMENTAL APPROACH: Retrospective analysis focusing on haemoglobin A1c (HbA1c) was performed in hypertensive and hyperglycaemic patients taking cilnidipine and amlodipine. After developing diabetic mice by streptozotocin (STZ) treatment, an osmotic pump including drug was implanted intraperitoneally, followed by weekly measurements of blood glucose levels. Mitochondrial morphology was analysed by electron microscopy. A Ca2+ channel-insensitive cilnidipine derivative (1,4-dihydropyridine [DHP]) was synthesized and its pharmacological effect was evaluated using obese (ob/ob) mice fed with high-fat diet (HFD). KEY RESULTS: In patients, cilnidipine was superior to amlodipine in HbA1c lowering effect. Cilnidipine treatment improved systemic hyperglycaemia and mitochondrial morphological abnormalities in STZ-exposed mice, without lowering blood pressure. Cilnidipine failed to improve hyperglycaemia of ob/ob mice, with suppressing insulin secretion. 1,4-DHP improved hyperglycaemia and mitochondria abnormality in ob/ob mice fed HFD. 1,4-DHP and cilnidipine improved basal oxygen consumption rate of HepG2 cells cultured under 25 mM glucose. CONCLUSION AND IMPLICATIONS: Inhibition of Drp1-filamin protein complex formation becomes a new strategy for type 2 diabetes treatment.

Drp1 and neuroinflammation: Deciphering the interplay between mitochondrial dynamics imbalance and inflammation in neurodegenerative diseases.

Neuroinflammation and mitochondrial dysfunction are closely intertwined with the pathophysiology of neurological disorders. Recent studies have elucidated profound alterations in mitochondrial dynamics across a spectrum of neurological disorders. Dynamin-related protein 1 (DRP1) emerges as a pivotal regulator of mitochondrial fission, with its dysregulation disrupting mitochondrial homeostasis and fueling neuroinflammation, thereby exacerbating disease severity. In addition to its role in mitochondrial dynamics, DRP1 plays a crucial role in modulating inflammation-related pathways. This review synthesizes important functions of DRP1 in the central nervous system (CNS) and the impact of epigenetic modification on the progression of neurodegenerative diseases. The intricate interplay between neuroinflammation and DRP1 in microglia and astrocytes, central contributors to neuroinflammation, is expounded upon. Furthermore, the use of DRP1 inhibitors to influence the activation of microglia and astrocytes, as well as their involvement in processes such as mitophagy, mitochondrial oxidative stress, and calcium ion transport in CNS-mediated neuroinflammation, is scrutinized. The modulation of microglia to astrocyte crosstalk by DRP1 and its role in inflammatory neurodegeneration is also highlighted. Overall, targeting DRP1 presents a promising avenue for ameliorating neuroinflammation and enhancing the therapeutic management of neurological disorders.

ROS-mediated cytoplasmic localization of CARM1 induces mitochondrial fission through DRP1 methylation.

The dynamic regulation of mitochondria through fission and fusion is essential for maintaining cellular homeostasis. In this study, we discovered a role of coactivator-associated arginine methyltransferase 1 (CARM1) in mitochondrial dynamics. CARM1 methylates specific residues (R403 and R634) on dynamin-related protein 1 (DRP1). Methylated DRP1 interacts with mitochondrial fission factor (Mff) and forms self-assembly on the outer mitochondrial membrane, thereby triggering fission, reducing oxygen consumption, and increasing reactive oxygen species (ROS) production. This sets in motion a feedback loop that facilitates the translocation of CARM1 from the nucleus to the cytoplasm, enhancing DRP1 methylation and ROS production through mitochondrial fragmentation. Consequently, ROS reinforces the CARM1-DRP1-ROS axis, resulting in cellular senescence. Depletion of CARM1 or DRP1 impedes cellular senescence by reducing ROS accumulation. The uncovering of the above-described mechanism fills a missing piece in the vicious cycle of ROS-induced senescence and contributes to a better understanding of the aging process.

Novel insight into mitochondrial dynamin-related protein-1 as a new chemo-sensitizing target in resistant cancer cells.

Mitochondrial dynamics have pillar roles in several diseases including cancer. Cancer cell survival is monitored by mitochondria which impacts several cellular functions such as cell metabolism, calcium signaling, and ROS production. The equilibrium of death and survival rate of mitochondria is important for healthy cellular processes. Whereas inhibition of mitochondrial metabolism and dynamics can have crucial regulatory decisions between cell survival and death. The steady rate of physiological flux of both mitochondrial fission and fusion is strongly related to the preservation of cellular bioenergetics. Dysregulation of mitochondrial dynamics including fission and fusion is a critical machinery in cells accompanied by crosstalk in cancer progression and resistance. Many cancer cells express high levels of Drp-1 to induce cancer cell invasion, metastasis and chemoresistance including breast cancer, liver cancer, pancreatic cancer, and colon cancer. Targeting Drp-1 by inhibitors such as Midivi-1 helps to enhance the responsiveness of cancer cells towards chemotherapy. The review showed Drp-1 linked processes such as mitochondrial dynamics and relationship with cancer, invasion, and chemoresistance along with computational assessing of all publicly available Drp-1 inhibitors. Drp1-IN-1, Dynole 34-2, trimethyloctadecylammonium bromide, and Schaftoside showed potential inhibitory effects on Drp-1 as compared to standard Mdivi- 1. This emerging approach may have extensive strength in the context of cancer development and chemoresistance and further work is needed to aid in more effective cancer management.

ATAD3 is a limiting factor in mitochondrial biogenesis and adipogenesis of white adipocyte-like 3T3-L1 cells.

ATAD3 is a vital ATPase of the inner mitochondrial membrane of pluri-cellular eukaryotes, with largely unknown functions but early required for organism development as necessary for mitochondrial biogenesis. ATAD3 knock-down in C. elegans inhibits at first the development of adipocyte-like intestinal tissue so we used mouse adipocyte model 3T3-L1 cells to analyze ATAD3 functions during adipogenesis and lipogenesis in a mammalian model. ATAD3 function was studied by stable and transient modulation of ATAD3 expression in adipogenesis- induced 3T3-L1 cells using Knock-Down and overexpression strategies, exploring different steps of adipocyte differentiation and lipogenesis. We show that (i) an increase in ATAD3 is preceding differentiation-induced mitochondrial biogenesis; (ii) downregulation of ATAD3 inhibits adipogenesis, lipogenesis, and impedes overexpression of many mitochondrial proteins; (iii) ATAD3 re-expression rescues the phenotype of ATAD3 KD, and (iv) differentiation and lipogenesis are accelerated by ATAD3 overexpression, but inhibited by expression of a dominant-negative mutant. We further show that the ATAD3 KD phenotype is not due to altered insulin signal but involves a limitation of mitochondrial biogenesis linked to Drp1. These results demonstrate that ATAD3 is limiting for in vitro mitochondrial biogenesis and adipogenesis/lipogenesis and therefore that ATAD3 mutation/over- or under-expression could be involved in adipogenic and lipogenic pathologies.

Upregulation of EPSTI1/Drp1/AKT1 Signaling Pathways Using pDNA/Melittin Against Breast Cancer.

This study investigates the role of genes related to breast cancer in apoptosis control. A melittin nucleic acid sequence was synthesized and introduced into a pcDNA3.1(+) Mammalian Expression Plasmid. The cloning accuracy was assessed using PCR testing and enzyme digestion techniques. The vectors were transfected into cells using LipofectamineTM2000. The transfection efficacy of MCF-7 and 4T1 cells was evaluated using fluorescence and bright-field imaging. Pure melittin produced from bee venom had a notable hemolytic impact, with lower hemolytic activity levels than the positive control, Triton X-100. The growth rate of 4T1 and MCF-7 cancer cells was significantly inhibited. The apoptosis rates were 8.54%, 46.20%, and 78.82% for free pDNA, melittin, and pDNA-melittin, respectively. The C-pDNA/Melittin-treated group showed a statistically significant reduction in cancer factors compared to the control group. The treated tumors exhibited significant necrosis and late apoptosis, with a prevalence ranging from about 5% to 10% of the lesions. After exposure to pDNA-melittin, there was no significant increase in transcription levels of caspase-3, caspase-8, BCRA1, BAX, Drp1, AKT1, and EPSTI1 genes in the normal non-cancerous groups. The findings provide novel opportunities for the therapeutic targeting of malignancies via melittin and the stimulation of the EPSTI1/Drp1/AKT1 signaling cascades.

Downregulation of HHATL promotes cardiac hypertrophy via activation of SHH/DRP1.

HHATL, previously implicated in cardiac hypertrophy in the zebrafish model, has emerged as a prioritized HCM risk gene. We identified six rare mutations in HHATL, present in 6.94 % of nonsarcomeric HCM patients (5/72). Moreover, a decrease of HHATL in the heart tissue from HCM patients and cardiac hypertrophy mouse model using transverse aortic constriction was observed. Despite this, the precise pathogenic mechanisms underlying HHATL-associated cardiac hypertrophy remain elusive. In this study, we observed that HHATL downregulation in H9C2 cells resulted in elevated expression of hypertrophic markers and reactive oxygen species (ROS), culminating in cardiac hypertrophy and mitochondrial dysfunction. Notably, the bioactive form of SHH, SHHN, exhibited a significant increase, while the mitochondrial fission protein dynamin-like GTPase (DRP1) decreased upon HHATL depletion. Intervention with the SHH inhibitor RU-SKI 43 or DRP1 overexpression effectively prevented Hhatl-depletion-induced cardiac hypertrophy, mitigating disruptions in mitochondrial morphology and membrane potential through the SHH/DRP1 axis. In summary, our findings suggest that HHATL depletion activates SHH signaling, reducing DRP1 levels and thereby promoting the expression of hypertrophic markers, ROS generation, and mitochondrial dysfunction, ultimately leading to cardiac hypertrophy. This study provides additional compelling evidence supporting the association of HHATL with cardiac hypertrophy.

Serine 39 in the GTP-binding domain of Drp1 is involved in shaping mitochondrial morphology.

Continuous fusion and fission are critical for mitochondrial health. In this study, we further characterize the role played by dynamin-related protein 1 (Drp1) in mitochondrial fission. We show that a single amino acid change in Drp1 at position 39 from serine to alanine (S39A) within the GTP-binding (GTPase) domain results in a fused mitochondrial network in human SH-SY5Y neuroblastoma cells. Interestingly, the phosphorylation of Ser-616 and Ser-637 of Drp1 remains unaffected by the S39A mutation, and mitochondrial bioenergetic profile and cell viability in the S39A mutant were comparable to those observed in the control. This leads us to propose that the serine 39 residue of Drp1 plays a crucial role in mitochondrial distribution through its involvement in the GTPase activity. Furthermore, this amino acid mutation leads to structural anomalies in the mitochondrial network. Taken together, our results contribute to a better understanding of the function of the Drp1 protein.

Chondrocyte-targeted exosome-mediated delivery of Nrf2 alleviates cartilaginous endplate degeneration by modulating mitochondrial fission.

BACKGROUND: Cartilaginous endplate (CEP) degeneration, which is an important contributor to intervertebral disc degeneration (IVDD), is characterized by chondrocyte death. Accumulating evidence has revealed that dynamin-related protein 1 (Drp1)-mediated mitochondrial fission and dysfunction lead to apoptosis during CEP degeneration and IVDD. Exosomes are promising agents for the treatment of many diseases, including osteoporosis, osteosarcoma, osteoarthritis and IVDD. Despite their major success in drug delivery, the full potential of exosomes remains untapped. MATERIALS AND METHODS: In vitro and in vivo models of CEP degeneration were established by using lipopolysaccharide (LPS). We designed genetically engineered exosomes (CAP-Nrf2-Exos) expressing chondrocyte-affinity peptide (CAP) on the surface and carrying the antioxidant transcription factor nuclear factor E2-related factor 2 (Nrf2). The affinity between CAP-Nrf2-Exos and CEP was evaluated by in vitro internalization assays and in vivo imaging assays. qRT‒PCR, Western blotting and immunofluorescence assays were performed to examine the expression level of Nrf2 and the subcellular localization of Nrf2 and Drp1. Mitochondrial function was measured by the JC-1 probe and MitoSOX Red. Mitochondrial morphology was visualized by MitoTracker staining and transmission electron microscopy (TEM). After subendplate injection of the engineered exosomes, the degree of CEP degeneration and IVDD was validated radiologically and histologically. RESULTS: We found that the cargo delivery efficiency of exosomes after cargo packaging was increased by surface modification. CAP-Nrf2-Exos facilitated chondrocyte-targeted delivery of Nrf2 and activated the endogenous antioxidant defence system in CEP cells. The engineered exosomes inhibited Drp1 S616 phosphorylation and mitochondrial translocation, thereby preventing mitochondrial fragmentation and dysfunction. LPS-induced CEP cell apoptosis was alleviated by CAP-Nrf2-Exo treatment. In a rat model of CEP degeneration, the engineered exosomes successfully attenuated CEP degeneration and IVDD and exhibited better repair capacity than natural exosomes. CONCLUSION: Collectively, our findings showed that exosome-mediated chondrocyte-targeted delivery of Nrf2 was an effective strategy for treating CEP degeneration.

MAP4K4 exacerbates cardiac microvascular injury in diabetes by facilitating S-nitrosylation modification of Drp1.

Dynamin-related protein 1 (Drp1) is a crucial regulator of mitochondrial dynamics, the overactivation of which can lead to cardiovascular disease. Multiple distinct posttranscriptional modifications of Drp1 have been reported, among which S-nitrosylation was recently introduced. However, the detailed regulatory mechanism of S-nitrosylation of Drp1 (SNO-Drp1) in cardiac microvascular dysfunction in diabetes remains elusive. The present study revealed that mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) was consistently upregulated in diabetic cardiomyopathy (DCM) and promoted SNO-Drp1 in cardiac microvascular endothelial cells (CMECs), which in turn led to mitochondrial dysfunction and cardiac microvascular disorder. Further studies confirmed that MAP4K4 promoted SNO-Drp1 at human C644 (mouse C650) by inhibiting glutathione peroxidase 4 (GPX4) expression, through which MAP4K4 stimulated endothelial ferroptosis in diabetes. In contrast, inhibition of MAP4K4 via DMX-5804 significantly reduced endothelial ferroptosis, alleviated cardiac microvascular dysfunction and improved cardiac dysfunction in db/db mice by reducing SNO-Drp1. In parallel, the C650A mutation in mice abolished SNO-Drp1 and the role of Drp1 in promoting cardiac microvascular disorder and cardiac dysfunction. In conclusion, our findings demonstrate that MAP4K4 plays an important role in endothelial dysfunction in DCM and reveal that SNO-Drp1 and ferroptosis activation may act as downstream targets, representing potential therapeutic targets for DCM.

lncRNA Oip5-as1 inhibits excessive mitochondrial fission in myocardial ischemia/reperfusion injury by modulating DRP1 phosphorylation.

BACKGROUND: Aberrant mitochondrial fission, a critical pathological event underlying myocardial ischemia/reperfusion (MI/R) injury, has emerged as a potential therapeutic target. The long non-coding RNA (lncRNA) Oip5-as1 is increasingly recognized for its regulatory roles, particularly in MI/R injury. However, its precise mechanistic role in modulating mitochondrial dynamics remains elusive. This study aims to elucidate the mechanistic role of Oip5-as1 in regulating mitochondrial fission and evaluate its therapeutic potential against MI/R injury. METHODS: To simulate in vitro MI/R injury, HL-1 cardiomyocytes were subjected to hypoxia/reoxygenation (H/R). Lentiviral vectors were employed to achieve overexpression or knockdown of Oip5-as1 in HL-1 cells by expressing Oip5-as1 or shRNA targeting Oip5-as1, respectively. The impact of Oip5-as1 on mitochondrial dynamics in HL-1 cells was assessed using CCK-8 assay, flow cytometry, immunofluorescence staining, and biochemical assays. MI/R injury was induced in mice by ligating the left anterior descending coronary artery. Conditional knockout mice for Oip5-as1 were generated using the CRISPR/Cas9 genome editing technology, while overexpression of Oip5-as1 in mice was achieved via intramyocardial administration of AAV9 vectors. In mice, the role of Oip5-as1 was evaluated through echocardiographic assessment, histopathological staining, and transmission electron microscopy. Furthermore, Western blotting, RNA pull-down, RNA immunoprecipitation, and co-immunoprecipitation assays were conducted to investigate Oip5-as1's underlying mechanisms. RESULTS: The expression levels of Oip5-as1 are significantly decreased in MI/R-injured HL-1 cells and myocardium. In HL-1 cells undergoing H/R injury, overexpression of Oip5-as1 attenuated excessive mitochondrial fission, preserved mitochondrial functionality, and reduced cellular apoptosis, while knockdown of Oip5-as1 exhibited the opposite effects. Furthermore, in a mouse model of MI/R injury, overexpression of Oip5-as1 diminished mitochondrial fission, myocardial infarct size and improved cardiac function. However, knockout of Oip5-as1 exacerbated myocardial injury and cardiac dysfunction, which were significantly reversed by treatment with a mitochondrial division inhibitor-1 (Mdivi-1). Mechanistically, Oip5-as1 selectively interacts with AKAP1 and CaN proteins, inhibiting CaN activation and subsequent DRP1 dephosphorylation at Ser637, thereby constraining DRP1's translocation to the mitochondria and its involvement in mitochondrial fission. CONCLUSIONS: Our study underscores the pivotal role of Oip5-as1 in mitigating excessive mitochondrial fission during MI/R injury. The findings not only enhance our comprehension of the molecular mechanisms underlying MI/R injury but also identify Oip5-as1 as a potential therapeutic target for ameliorating MI/R injury.

Hypoxia-induced SENP3 promotes chemosensitivity and mitochondrial fission via deSUMOylation of Drp1.

OBJECTIVE: The study aimed to investigate the effect of the SUMOylation status of Drp1 on mitochondrial fission in CDDP-treated HNSCC cells cultured under hypoxic conditions. MATERIALS AND METHODS: The effect of hypoxia on the chemosensitivity of HNCC cells was evaluated by flow cytometry and CCK-8 assays. The biological function of SUMO-specific peptidase 3 (SENP3) was evaluated by loss-of-function assays both in vitro and in vivo. SENP3-regulated deSUMOylation of Drp1 were performed with co-IP assays. RESULTS: SENP3 expression correlated with chemosensitivity in clinical HNSCC samples subjected to hypoxic conditions. Hypoxia-induced ROS increased HIF-1α/SENP3 expression and mitochondrial fission in CDDP-treated HNSCC cells, and these effects were reversed by NAC treatment. SENP3 knockdown reversed hypoxia-induced mitochondrial fission and inhibited HNSCC cell apoptosis, which decreased CDDP sensitivity. Furthermore, hypoxia-induced SENP3 deconjugated SUMO2 from Drp1. CONCLUSION: Our findings revealed that hypoxia-induced SENP3 facilitates CDDP sensitivity and mitochondrial fission via deSUMOylation of Drp1.

Lactate facilitated mitochondrial fission-derived ROS to promote pulmonary fibrosis via ERK/DRP-1 signaling.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, fibrotic interstitial lung diseases, which mainly existed in middle-aged and elderly people. The accumulation of reactive oxygen species (ROS) is a common characteristic of IPF. Previous research also shown that lactate levels can be abnormally elevated in IPF patients. Emerging evidence suggested a relationship between lactate and ROS in IPF which needs further elucidation. In this article, we utilized a mouse model of BLM-induced pulmonary fibrosis to detect alterations in ROS levels and other indicators associated with fibrosis. Lactate could induce mitochondrial fragmentation by modulating expression and activity of DRP1 and ERK. Moreover, Increased ROS promoted P65 translocation into nucleus, leading to expression of lung fibrotic markers. Finally, Ulixertinib, Mdivi-1 and Mito-TEMPO, which were inhibitor activity of ERK, DRP1 and mtROS, respectively, could effectively prevented mitochondrial damage and production of ROS and eventually alleviate pulmonary fibrosis. Taken together, these findings suggested that lactate could promote lung fibrosis by increasing mitochondrial fission-derived ROS via ERK/DRP1 signaling, which may provide novel therapeutic solutions for IPF.

Cortistatin protects against septic cardiomyopathy by inhibiting cardiomyocyte pyroptosis through the SSTR2-AMPK-NLRP3 pathway.

BACKGROUND: Although the pathophysiological mechanism of septic cardiomyopathy has been continuously discovered, it is still a lack of effective treatment method. Cortistatin (CST), a neuroendocrine polypeptide of the somatostatin family, has emerged as a novel cardiovascular-protective peptide, but the specific mechanism has not been elucidated. PURPOSE: The aim of our study is to explore the role of CST in cardiomyocytes pyroptosis and myocardial injury in sepsis and whether CST inhibits cardiomyocytes pyroptosis through specific binding with somastatin receptor 2 (SSTR2) and activating AMPK/Drp1 signaling pathway. METHODS AND RESULTS: In this study, plasma CST levels were significantly high and were negatively correlated with N-terminal pro-B type natriuretic peptide (NT-proBNP), a biomarker for cardiac dysfunction, in patients with sepsis. Exogenous administration of CST significantly improved survival rate and cardiac function in mouse models of sepsis by inhibiting the activation of the NLRP3 inflammasome and pyroptosis of cardiomyocytes (decreased cleavage of caspase-1, IL-1ß and gasdermin D). Pharmacological inhibition and genetic ablation revealed that CST exerted anti-pyroptosis effects by specifically binding to somatostatin receptor subtype 2 (SSTR2), thus activating AMPK and inactivating Drp1 to inhibit mitochondrial fission in cardiomyocytes. CONCLUSIONS: This study is the first to report that CST attenuates septic cardiomyopathy by inhibiting cardiomyocyte pyroptosis through the SSTR2-AMPK-Drp1-NLRP3 pathway. Importantly, CST specifically binds to SSTR2, which promotes AMPK phosphorylation, inhibits Drp1-mediated mitochondrial fission, and reduces ROS levels, thereby inhibiting NLRP3 inflammasome activation-mediated pyroptosis and alleviating sepsis-induced myocardial injury.

Imbalanced mitochondrial dynamics contributes to the pathogenesis of X-linked adrenoleukodystrophy.

The peroxisomal disease adrenoleukodystrophy (X-ALD) is caused by loss of the transporter of very-long-chain fatty acids (VLCFAs), ABCD1. An excess of VLCFAs disrupts essential homeostatic functions crucial for axonal maintenance, including redox metabolism, glycolysis and mitochondrial respiration. As mitochondrial function and morphology are intertwined, we set out to investigate the role of mitochondrial dynamics in X-ALD models. Using quantitative 3D transmission electron microscopy, we revealed mitochondrial fragmentation in corticospinal axons in Abcd1- mice. In patient fibroblasts, an excess of VLCFAs triggers mitochondrial fragmentation through the redox-dependent phosphorylation of DRP1 (DRP1S616). The blockade of DRP1-driven fission by the peptide P110 effectively preserved mitochondrial morphology. Furthermore, mRNA inhibition of DRP1 not only prevented mitochondrial fragmentation but also protected axonal health in a Caenorhabditis elegans model of X-ALD, underscoring DRP1 as a potential therapeutic target. Elevated levels of circulating cell-free mtDNA in patients' CSF align this leukodystrophy with primary mitochondrial disorders. Our findings underscore the intricate interplay between peroxisomal dysfunction, mitochondrial dynamics and axonal integrity in X-ALD, shedding light on potential avenues for therapeutic intervention.

Nrf2 expression, mitochondrial fission, and neuronal apoptosis in the prefrontal cortex of methamphetamine abusers and rats.

Methamphetamine (MA), a representative amphetamine-type stimulant, is one of the most abused drugs worldwide. Studies have shown that MA-induced neurotoxicity is strongly associated with oxidative stress and apoptosis. While nuclear factor E2-related factor 2 (Nrf2), an antioxidant transcription factor, is known to exert neuroprotective effects, its role in MA-induced dopaminergic neuronal apoptosis remains incompletely understood. In the present study, we explored the effects of MA on the expression levels of Nrf2, dynamin-related protein 1 (Drp1), mitofusin 1 (Mfn1), cytochrome c oxidase (Cyt-c), and cysteine aspartate-specific protease 3 (Caspase 3), as well as the correlations between Nrf2 and mitochondrial dynamics and apoptosis. Brain tissue from MA abusers was collected during autopsy procedures. An MA-dependent rat model was also established by intraperitoneal administration of MA (10 mg/kg daily) for 28 consecutive days, followed by conditioned place preference (CPP) testing. Based on immunohistochemical staining and western blot analysis, the protein expression levels of Nrf2 and Mfn1 showed a decreasing trend, while levels of Drp1, Cyt-c, and Caspase 3 showed an increasing trend in the cerebral prefrontal cortex of both MA abusers and MA-dependent rats. Notably, the expression of Nrf2 was positively associated with the expression of Mfn1, but negatively associated with the expression levels of Drp1, Cyt-c, and Caspase 3. These findings suggest that oxidative stress and mitochondrial fission contribute to neuronal apoptosis, with Nrf2 potentially playing a critical role in MA-induced neurotoxicity.

The Effect of Cold-Water Swimming on Energy Metabolism, Dynamics, and Mitochondrial Biogenesis in the Muscles of Aging Rats.

Our study aimed to explore the potential positive effects of cold water exercise on mitochondrial biogenesis and muscle energy metabolism in aging rats. The study involved 32 male and 32 female rats aged 15 months, randomly assigned to control sedentary animals, animals training in cold water at 5 ± 2 °C, or animals training in water at thermal comfort temperature (36 ± 2 °C). The rats underwent swimming training for nine weeks, gradually increasing the duration of the sessions from 2 min to 4 min per day, five days a week. The results demonstrated that swimming in thermally comfortable water improved the energy metabolism of aging rat muscles (increased metabolic rates expressed as increased ATP, ADP concentration, TAN (total adenine nucleotide) and AEC (adenylate energy charge value)) and increased mRNA and protein expression of fusion regulatory proteins. Similarly, cold-water swimming improved muscle energy metabolism in aging rats, as shown by an increase in muscle energy metabolites and enhanced mitochondrial biogenesis and dynamics. It can be concluded that the additive effect of daily activity in cold water influenced both an increase in the rate of energy metabolism in the muscles of the studied animals and an intensification of mitochondrial biogenesis and dynamics (related to fusion and fragmentation processes). Daily activity in warm water also resulted in an increase in the rate of energy metabolism in muscles, but at the same time did not cause significant changes in mitochondrial dynamics.

Mitochondrial dysfunction induced by HIF-1α under hypoxia contributes to the development of gastric mucosal lesions.

INTRODUCTION: Hypoxia is an important characteristic of gastric mucosal diseases, and hypoxia-inducible factor-1α (HIF-1α) contributes to microenvironment disturbance and metabolic spectrum abnormalities. However, the underlying mechanism of HIF-1α and its association with mitochondrial dysfunction in gastric mucosal lesions under hypoxia have not been fully clarified. OBJECTIVES: To evaluate the effects of hypoxia-induced HIF-1α on the development of gastric mucosal lesions. METHODS: Portal hypertensive gastropathy (PHG) and gastric cancer (GC) were selected as representative diseases of benign and malignant gastric lesions, respectively. Gastric tissues from patients diagnosed with the above diseases were collected. Portal hypertension (PHT)-induced mouse models in METTL3 mutant or NLRP3-deficient littermates were established, and nude mouse gastric graft tumour models with relevant inhibitors were generated. The mechanisms underlying hypoxic condition, mitochondrial dysfunction and metabolic alterations in gastric mucosal lesions were further analysed. RESULTS: HIF-1α, which can mediate mitochondrial dysfunction via upregulation of METTL3/IGF2BP3-dependent dynamin-related protein 1 (Drp1) N6-methyladenosine modification to increase mitochondrial reactive oxygen species (mtROS) production, was elevated under hypoxic conditions in human and mouse portal hypertensive gastric mucosa and GC tissues. While blocking HIF-1α with PX-478, inhibiting Drp1-dependent mitochondrial fission via mitochondrial division inhibitor 1 (Mdivi-1) treatment or METTL3 mutation alleviated this process. Furthermore, HIF-1α influenced energy metabolism by enhancing glycolysis via lactate dehydrogenase A. In addition, HIF-1α-induced Drp1-dependent mitochondrial fission also enhanced glycolysis. Drp1-dependent mitochondrial fission and enhanced glycolysis were associated with alterations in antioxidant enzyme activity and dysfunction of the mitochondrial electron transport chain, resulting in massive mtROS production, which was needed for activation of NLRP3 inflammasome to aggravate the development of the PHG and GC. CONCLUSIONS: Under hypoxic conditions, HIF-1α enhances mitochondrial dysfunction via Drp1-dependent mitochondrial fission and influences the metabolic profile by altering glycolysis to increase mtROS production, which can trigger NLRP3 inflammasome activation and mucosal microenvironment alterations to contribute to the development of benign and malignant gastric mucosal lesions.

The CaMK Family Differentially Promotes Necroptosis and Mouse Cardiac Graft Injury and Rejection.

Organ transplantation is associated with various forms of programmed cell death which can accelerate transplant injury and rejection. Targeting cell death in donor organs may represent a novel strategy for preventing allograft injury. We have previously demonstrated that necroptosis plays a key role in promoting transplant injury. Recently, we have found that mitochondria function is linked to necroptosis. However, it remains unknown how necroptosis signaling pathways regulate mitochondrial function during necroptosis. In this study, we investigated the receptor-interacting protein kinase 3 (RIPK3) mediated mitochondrial dysfunction and necroptosis. We demonstrate that the calmodulin-dependent protein kinase (CaMK) family members CaMK1, 2, and 4 form a complex with RIPK3 in mouse cardiac endothelial cells, to promote trans-phosphorylation during necroptosis. CaMK1 and 4 directly activated the dynamin-related protein-1 (Drp1), while CaMK2 indirectly activated Drp1 via the phosphoglycerate mutase 5 (PGAM5). The inhibition of CaMKs restored mitochondrial function and effectively prevented endothelial cell death. CaMKs inhibition inhibited activation of CaMKs and Drp1, and cell death and heart tissue injury (n = 6/group, p < 0.01) in a murine model of cardiac transplantation. Importantly, the inhibition of CaMKs greatly prolonged heart graft survival (n = 8/group, p < 0.01). In conclusion, CaMK family members orchestrate cell death in two different pathways and may be potential therapeutic targets in preventing cell death and transplant injury.

Inhibition of Drp1-mediated mitochondrial fission improves contrast-induced acute kidney injury by targeting the mROS-TXNIP-NLRP3 inflammasome axis.

Acute kidney injury (AKI) is a critical complication known for their extremely high mortality rate and lack of effective clinical therapy. Disorders in mitochondrial dynamics possess a pivotal role in the occurrence and progression of contrast-induced nephropathy (CIN) by activating NLRP3 inflammasome. The activation of dynamin-related protein-1 (Drp1) can trigger mitochondrial dynamic disorders by regulating excessive mitochondrial fission. However, the precise role of Drp1 during CIN has not been clarified. In vivo experiments revealed that inhibiting Drp1 through Mdivi-1 (one selective inhibitor of Drp1) can significantly decrease the expression of p-Drp1 (Ser616), mitochondrial p-Drp1 (Ser616), mitochondrial Bax, mitochondrial reactive oxygen species (mROS), NLRP3, caspase-1, ASC, TNF-α, IL-1ß, interleukin (IL)-18, IL-6, creatinine (Cr), malondialdehyde (MDA), blood urea nitrogen (BUN), and KIM-1. Moreover, Mdivi-1 reduced kidney pathological injury and downregulated the interaction between NLRP3 and thioredoxin-interacting protein (TXNIP), which was accompanied by decreased interactions between TRX and TXNIP. This resulted in increasing superoxide dismutase (SOD) and CAT activity, TRX expression, up-regulating mitochondrial membrane potential, and augmenting ATP contents and p-Drp1 (Ser616) levels in the cytoplasm. However, it did not bring impact on the expression of p-Drp1 (Ser637) and TXNIP. Activating Drp-1though Acetaldehyde abrogated the effects of Mdivi-1. In addition, the results of in vitro studies employing siRNA-Drp1 and plasmid-Drp1 intervention in HK-2 cells treated with iohexol were consistent with the in vivo experiments. Our findings revealed inhibiting Drp1 phosphorylation at Ser616 could ameliorate iohexol -induced acute kidney injury though alleviating the activation of the TXNIP-NLRP3 inflammasome pathway.