Intermittent theta burst stimulation (iTBS), a time-saving and cost-effective repetitive transcranial magnetic stimulation regime, has been shown to improve cognition in patients with Alzheimer's disease (AD). However, the specific mechanism underlying iTBS-induced cognitive enhancement remains unknown. Previous studies suggested that mitochondrial functions are modulated by magnetic stimulation. Here, we showed that iTBS upregulates the expression of iron-sulfur cluster assembly 1 (ISCA1, an essential regulatory factor for mitochondrial respiration) in the brain of APP/PS1 mice. In vivo and in vitro studies revealed that iTBS modulates mitochondrial iron-sulfur cluster assembly to facilitate mitochondrial respiration and function, which is required for ISCA1. Moreover, iTBS rescues cognitive decline and attenuates AD-type pathologies in APP/PS1 mice. The present study uncovers a novel mechanism by which iTBS modulates mitochondrial respiration and function via ISCA1-mediated iron-sulfur cluster assembly to alleviate cognitive impairments and pathologies in AD. We provide the mechanistic target of iTBS that warrants its therapeutic potential for AD patients.
Humans ; Mice ; Animals ; Transcranial Magnetic Stimulation ; Alzheimer Disease/therapy* ; Cognitive Dysfunction/therapy* ; Cognition ; Sulfur ; Iron ; Iron-Sulfur Proteins ; Mitochondrial Proteins

Apoptosis Regulatory Proteins ; Apoptosis ; Antiviral Agents/pharmacology* ; Mitochondrial Proteins ; Cell Line, Tumor ; Antineoplastic Agents/pharmacology*

This study aimed to investigate the effect of treadmill exercise on neuropathic pain and to determine whether mitophagy of the anterior cingulate cortex (ACC) contributes to exercise-mediated amelioration of neuropathic pain. Chronic constriction injury of the sciatic nerve (CCI) was used to establish a neuropathic pain model in Sprague-Dawley (SD) rats. Von-Frey filaments were used to assess the mechanical paw withdrawal threshold (PWT), and a thermal radiation meter was used to assess the thermal paw withdrawal latency (PWL) in rats. qPCR was used to evaluate the mRNA levels of Pink1, Parkin, Fundc1, and Bnip3. Western blot was used to evaluate the protein levels of PINK1 and PARKIN. To determine the impact of the mitophagy inducer carbonyl cyanide m-chlorophenylhydrazone (CCCP) on pain behaviors in CCI rats, 24 SD rats were randomly divided into CCI drug control group (CCI+Veh group), CCI+CCCP low-dose group (CCI+CCCP0.25), CCI+CCCP medium-dose group (CCI+CCCP2.5), and CCI+CCCP high-dose group (CCI+CCCP5). Pain behaviors were assessed on 0, 1, 3, 5, and 7 days after modeling. To explore whether exercise regulates pain through mitophagy, 24 SD rats were divided into sham, CCI, and CCI+Exercise (CCI+Exe) groups. The rats in the CCI+Exe group underwent 4-week low-moderate treadmill training one week after modeling. The mechanical pain and thermal pain behaviors of the rats in each group were assessed on 0, 7, 14, 21, and 35 days after modeling. Western blot was used to detect the levels of the mitophagy-related proteins PINK1, PARKIN, LC3 II/LC3 I, and P62 in ACC tissues. Transmission electron microscopy was used to observe the ultrastructure of mitochondrial morphology in the ACC. The results showed that: (1) Compared with the sham group, the pain thresholds of the ipsilateral side of the CCI group decreased significantly (P < 0.001). Meanwhile, the mRNA and protein levels of Pink1 were significantly higher, and those of Parkin were lower in the CCI group (P < 0.05). (2) Compared with the CCI+Veh group, each CCCP-dose group showed higher mechanical and thermal pain thresholds, and the levels of PINK1 and LC3 II/LC3 I were elevated significantly (P < 0.05, P < 0.01). (3) The pain thresholds of the CCI+Exe group increased significantly compared with those of the CCI group after treadmill intervention (P < 0.001, P < 0.01). Compared with the CCI group, the protein levels of PINK1 and P62 were decreased (P < 0.001, P < 0.01), and the protein levels of PARKIN and LC3 II/LC3 I were increased in the CCI+Exe group (P < 0.01, P < 0.05). Rod-shaped mitochondria were observed in the ACC of CCI+Exe group, and there were little mitochondrial fragmentation, swelling, or vacuoles. The results suggest that the mitochondrial PINK1/PARKIN autophagy pathway is blocked in the ACC of neuropathic pain model rats. Treadmill exercise could restore mitochondrial homeostasis and relieve neuropathic pain via the PINK1/PARKIN pathway.
Rats ; Animals ; Mitophagy/physiology* ; Rats, Sprague-Dawley ; Carbonyl Cyanide m-Chlorophenyl Hydrazone/pharmacology* ; Gyrus Cinguli ; Neuralgia ; Ubiquitin-Protein Ligases/metabolism* ; Protein Kinases ; Membrane Proteins/metabolism* ; Mitochondrial Proteins/metabolism*

Objective To study the effect and mechanism of blueberry on regulating the mitochondrial inner membrane protein mitofilin/Mic60 in an in vitro model of metabolic dysfunction-associated liver disease (MAFLD). Methods L02 human hepatocytes were induced by free fatty acids (FFA) to establish MAFLD cell model. A normal group, a model group, an 80 μg/mL blueberry treatment group, a Mic60 short hairpin RNA (Mic60 shRNA) transfection group, and Mic60 knockdown combined with an 80 μg/mL blueberry treatment group were established. The intracellular lipid deposition was observed by oil red O staining, and the effect of different concentrations of blueberry pulp on the survival rate of L02 cells treated with FFA was measured by MTT assay. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), superoxide dismutase (SOD) activity, glutathione (GSH) and malondialdehyde (MDA) contents were measured by visible spectrophotometry. The expression of reactive oxygen species (ROS) in hepatocytes was observed by fluorescence microscopy, and the mRNA and protein expression of Mic60 were detected by real-time quantitative PCR and Western blot analysis, respectively. Results After 24 hours of FFA stimulation, a large number of red lipid droplets in the cytoplasm of L02 cells was observed, and the survival rate of L02 cells treated with 80 μg/mL blueberry was higher. The results of ALT, AST, TG, TC, MDA and the fluorescence intensity of ROS in blueberry treated group were lower than those in model group, while the levels of SOD, GSH, Mic60 mRNA and protein in blueberry treated group were higher than those in model group. Conclusion Blueberry promotes the expression of Mic60, increases the levels of SOD and GSH in hepatocytes, and reduces the production of ROS, thus alleviating the injury of MAFLD hepatocytes and regulating the disorder of lipid metabolism.
Humans ; Blueberry Plants/chemistry* ; Hepatocytes/metabolism* ; Liver/metabolism* ; Liver Diseases/metabolism* ; Reactive Oxygen Species/metabolism* ; Superoxide Dismutase/metabolism* ; Superoxides/metabolism* ; Mitochondrial Membranes/metabolism* ; Mitochondrial Proteins/metabolism* ; Plant Extracts/pharmacology*

OBJECTIVES: To study the genetic characteristics, clinical characteristics, and prognosis of children with primary dilated cardiomyopathy (DCM). METHODS: A retrospective analysis was performed on the medical data of 44 children who were diagnosed with DCM in Hebei Children's Hospital from July 2018 to February 2023. According to the genetic testing results, they were divided into two groups: gene mutation-positive group (n=17) and gene mutation-negative group (n=27). The two groups were compared in terms of clinical data at initial diagnosis and follow-up data. RESULTS: Among the 44 children with DCM, there were 21 boys (48%) and 23 girls (52%). Respiratory symptoms including cough and shortness of breath were the most common symptom at initial diagnosis (34%, 15/44). The detection rate of gene mutations was 39% (17/44). There were no significant differences between the two groups in clinical characteristics, proportion of children with cardiac function grade Ⅲ or Ⅳ, brain natriuretic peptide levels, left ventricular ejection fraction, and left ventricular fractional shortening at initial diagnosis (P>0.05). The median follow-up time was 23 months, and 9 children (20%) died, including 8 children from the gene mutation-positive group, among whom 3 had TTN gene mutation, 2 had LMNA gene mutation, 2 had TAZ gene mutation, and 1 had ATAD3A gene mutation. The gene mutation-positive group had a significantly higher mortality rate than the gene mutation-negative group (P<0.05). CONCLUSIONS There is no correlation between the severity of DCM at initial diagnosis and gene mutations in children. However, children with gene mutations may have a poorer prognosis.
Male ; Female ; Humans ; Child ; Stroke Volume ; Retrospective Studies ; Ventricular Function, Left ; Phenotype ; Cardiomyopathy, Dilated/diagnosis* ; Mutation ; ATPases Associated with Diverse Cellular Activities/genetics* ; Membrane Proteins/genetics* ; Mitochondrial Proteins/genetics*

BACKGROUND: Epidermal growth factor receptor (EGFR) gene mutations are the most common driver mutations in non-small cell lung cancer (NSCLC). To prolong the survival of the patients, EGFR tyrosine kinase inhibitors (TKIs) resistance in NSCLC is a major challenge that needs to be addressed urgently, and this study focuses on investigating the mechanism of cigarette smoke (CS) induced Gefitinib resistance in NSCLC. METHODS: PC-9 and A549 cells were cultured in vitro and treated with 1 µmol/L Gefitinib for 4 h and 10% cigarette smoke extract (CSE) for 48 h. Western blot was used to detect Sirtuin 3 (Sirt3) and superoxide dismutase 2 (SOD2) protein expressions; DCFH-DA probe was used to detect intracellular reactive oxygen species (ROS); CCK-8 kit was used to detect cell activity, and EdU was used to detect cell proliferation ability. Sirt3 overexpression plasmid (OV-Sirt3) was transfected in PC-9 and A549 cells and treated with 1 µmol/L Gefitinib for 4 h and 10% CSE for 48 h after N-acetylcysteine (NAC) action. The expressions of Sirt3 and SOD2 were detected by Western blot; the ROS level in the cells was detected by DCFH-DA probe, and the cell activity was detected by CCK-8. RESULTS: CSE induced an increase in the 50% inhibitory concentration (IC50) of both PC-9 and A549 cells to Gefitinib (P<0.01) and enhanced the proliferation of PC-9 and A549 cells, suggesting that CS induced Gefitinib resistance in NSCLC. ROS was involved in CSE-induced Gefitinib resistance (P<0.05). CSE induced low expressions of Sirt3 and SOD2 (P<0.01), and Sirt3/SOD2 was associated with poor prognosis in lung cancer patients (P<0.05). OV-Sirt3 in PC-9 and A549 cells reversed CSE-induced Gefitinib resistance (P<0.05) and significantly reduced ROS production. NAC reversed CSE-induced Gefitinib resistance in PC-9 and A549 cells (P<0.05). CONCLUSIONS The ROS/Sirt3/SOD2 pathway is involved in CS-induced Gefitinib resistance in NSCLC.
Humans ; Gefitinib/therapeutic use* ; Carcinoma, Non-Small-Cell Lung/metabolism* ; Sirtuin 3/therapeutic use* ; Lung Neoplasms/metabolism* ; Reactive Oxygen Species/therapeutic use* ; Antineoplastic Agents/therapeutic use* ; Cigarette Smoking ; Sincalide/therapeutic use* ; ErbB Receptors/metabolism* ; Drug Resistance, Neoplasm/genetics* ; Cell Line, Tumor

OBJECTIVE: To investigate the growth-inhibitory and pro-apoptotic effects of piroctone olamine (PO) on glioma cells and explore the underlying mechanism. METHODS: Human glioma cell lines U251 and U373 were treated with PO and the changes in cell proliferation were detected using CCK-8 assay and EdU assay. Clone formation assay and flow cytometry were used to examine the changes in clone formation ability and apoptosis of the treated cells. Mitochondrial membrane potential of the cells and morphological changes of the mitochondria were detected using JC-1 staining and a fluorescence probe, respectively. The expressions of mitochondrial fission protein DRP1 and the fusion protein OPA1 were determined with Western blotting. Transcriptome sequencing and differential gene enrichment analysis was performed, and the expression levels of PI3K, AKT and p-AKT in the treated cells were verified using Western blotting. RESULTS: CCK-8 assay showed that PO significantly inhibited the proliferation of U251 and U373 cells in a time- and dose-dependent manner (P < 0.001). EdU test showed that the proliferative activity of PO-treated cells was significantly decreased, and the number of cell colonies also decreased significantly (P < 0.01). PO treatment significantly increased apoptotic rates (P < 0.01), decreased mitochondrial membrane potential and caused obvious changes in mitochondrial morphology of the cells. Pathway enrichment analysis showed that the down-regulated genes were significantly enriched in the PI3K/AKT pathway, which was verified by Western blotting showing significantly down-regulated expression levels of PI3K, AKT and p-AKT in PO-treated cells (P < 0.05). CONCLUSION PO interferes with mitochondrial fusion and fission function through the PI3K/AKT pathway, thereby inhibiting the proliferation and increasing apoptosis of glioma cells.
Humans ; Glioma ; Mitochondrial Dynamics ; Phosphatidylinositol 3-Kinases ; Proto-Oncogene Proteins c-akt

We identified distinct senescence-related molecular subtypes and critical genes among prostate cancer (PCa) patients undergoing radical prostatectomy (RP) or radical radiotherapy (RT). We conducted all analyses using R software and its suitable packages. Twelve genes, namely, secreted frizzled-related protein 4 (SFRP4), DNA topoisomerase II alpha (TOP2A), pleiotrophin (PTN), family with sequence similarity 107 member A (FAM107A), C-X-C motif chemokine ligand 14 (CXCL14), prostate androgen-regulated mucin-like protein 1 (PARM1), leucine zipper protein 2 (LUZP2), cluster of differentiation 38 (CD38), cartilage oligomeric matrix protein (COMP), vestigial-like family member 3 (VGLL3), apolipoprotein E (APOE), and aldehyde dehydrogenase 2 family member (ALDH2), were eventually used to subtype PCa patients from The Cancer Genome Atlas (TCGA) database and GSE116918, and the molecular subtypes showed good correlations with clinical features. In terms of the tumor immune environment (TME) analysis, compared with cluster 1, cancer-associated fibroblasts (CAFs) scored significantly higher, while endothelial cells scored lower in cluster 2 in TCGA database. There was a statistically significant correlation between both CAFs and endothelial cells with biochemical recurrence (BCR)-free survival for PCa patients undergoing RP. For the GSE116918 database, cluster 2 had significantly lower levels of CAFs and tumor purity and higher levels of stromal, immune, and Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data (ESTIMATE) scores than cluster 1; in addition, patients with high levels of CAFs, stromal scores, immune scores, and ESTIMATE scores and low levels of tumor purity tended to suffer from BCR. Based on the median of differentially expressed checkpoints, high expression of CD96, hepatitis A virus cellular receptor 2 (HAVCR2), and neuropilin 1 (NRP1) in GSE116918 and high expression of CD160 and tumor necrosis factor (ligand) superfamily member 18 (TNFSF18) in TCGA database were associated with a significantly higher risk of BCR than their counterparts. In conclusion, we first constructed distinct molecular subtypes and critical genes for PCa patients undergoing RP or RT from the fresh perspective of senescence.
Male ; Humans ; Endothelial Cells ; Ligands ; Prostatic Neoplasms/pathology* ; Prostate/pathology* ; Prostatectomy ; Aldehyde Dehydrogenase, Mitochondrial ; DNA-Binding Proteins ; Transcription Factors

OBJECTIVE: To explore the protective effect and possible mechanisms of bloodletting acupuncture at Jing-well points (BAJP) pre-treatment on acute hypobaric hypoxia (AHH)-induced myocardium injury rat. METHODS: Seventy-five rats were randomly divided into 5 groups by a random number table: a control group (n=15), a model group (n=15), a BAJP group (n=15), a BAJP+3-methyladenine (3-MA) group (n=15), and a BANA (bloodletting at nonacupoint; tail bleeding, n=15) group. Except for the control group, the AHH rat model was established in the other groups, and the corresponding treatment methods were adopted. Enzyme-linked immunosorbent assay (ELISA) was used to detect creatine kinase isoenzyme MB (CK-MB) and cardiac troponins I (CTnI) levels in serum and superoxide dismutase (SOD) and malondialdehyde (MDA) levels in myocardial tissue. Hematoxylin-eosin (HE) staining was used to observe myocardial injury, and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) staining was used to observe cell apoptosis. Transmission electron microscopy detection was used to observe mitochondrial damage and autophagosomes in the myocardium. The mitochondrial membrane potential of the myocardium was analyzed with the fluorescent dye JC-1. Mitochondrial respiratory chain complex (complex I, III, and IV) activities and ATPase in the myocardium were detected by mitochondrial respiratory chain complex assay kits. Western blot analysis was used to detect the autophagy index and hypoxia inducible factor-1α (HIF-1α)/Bcl-2 and adenovirus E1B 19k Da-interacting protein 3 (BNIP3) signaling. RESULTS: BAJP reduced myocardial injury and inhibited myocardial cell apoptosis in AHH rats. BAJP pretreatment decreased MDA levels and increased SOD levels in AHH rats (all P<0.01). Moreover, BAJP pretreatment increased the mitochondrial membrane potential (P<0.01), mitochondrial respiratory chain complex (complexes I, III, and IV) activities (P<0.01), and mitochondrial ATPase activity in AHH rats (P<0.05). The results from electron microscopy demonstrated that BAJP pretreatment improved mitochondrial swelling and increased the autophagosome number in the myocardium of AHH rats. In addition, BAJP pretreatment activated the HIF-1α/BNIP3 pathway and autophagy. Finally, the results of using 3-MA to inhibit autophagy in BAJP-treated AHH rats showed that suppression of autophagy attenuated the treatment effects of BAJP in AHH rats, further proving that autophagy constitutes a potential target for BAJP treatment of AHH. CONCLUSION BAJP is an effective treatment for AHH-induced myocardial injury, and the mechanism might involve increasing HIF-1α/BNIP3 signaling-mediated autophagy and decreasing oxidative stress.
Animals ; Rats ; Acupuncture Therapy ; Altitude ; Apoptosis ; Autophagy ; Bloodletting ; Hypoxia/metabolism* ; Membrane Proteins/pharmacology* ; Mitochondrial Proteins/pharmacology* ; Oxidative Stress ; Rats, Sprague-Dawley

Humans ; Brain Neoplasms/metabolism* ; Cell Line, Tumor ; Cell Proliferation ; Gene Expression Regulation, Neoplastic ; Glioma/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Signal Transduction ; ATPases Associated with Diverse Cellular Activities/metabolism* ; Mitochondrial Proteins/metabolism* ; GTP-Binding Protein alpha Subunits/metabolism*