Nerve regeneration following traumatic peripheral nerve injuries and neuropathies is a complex process modulated by diverse factors and intricate molecular mechanisms. Past studies have focused on factors that stimulate axonal outgrowth and myelin regeneration. However, recent studies have highlighted the pivotal role of autophagy in peripheral nerve regeneration, particularly in the context of traumatic injuries. Consequently, autophagy-targeting modulation has emerged as a promising therapeutic approach to enhancing peripheral nerve regeneration. Our current understanding suggests that activating autophagy facilitates the rapid clearance of damaged axons and myelin sheaths, thereby enhancing neuronal survival and mitigating injury-induced oxidative stress and inflammation. These actions collectively contribute to creating a favorable microenvironment for structural and functional nerve regeneration. A range of autophagy-inducing drugs and interventions have demonstrated beneficial effects in alleviating peripheral neuropathy and promoting nerve regeneration in preclinical models of traumatic peripheral nerve injuries. This review delves into the regulation of autophagy in cell types involved in peripheral nerve regeneration, summarizing the potential drugs and interventions that can be harnessed to promote this process. We hope that our review will offer novel insights and perspectives on the exploitation of autophagy pathways in the treatment of peripheral nerve injuries and neuropathies.

Autophagy is a "self-eating" biological process that degrades cytoplasmic contents to ensure cellular homeostasis. Its response to stimuli occurs in two stages: Within a few to several hours of exposure to a stress condition, autophagic flow rapidly increases, which is mediated by post-translational modification (PTM). Subsequently, the transcriptional program is activated and mediates the persistent autophagic response. O-linked β-N-acetylglucosamine (O-GlcNAc) modification is an inducible and dynamically cycling PTM; mounting evidence suggests that O-GlcNAc modification participates in the total autophagic process, including autophagy initiation, autophagosome formation, autophagosome-lysosome fusion, and transcriptional process. In this review, we summarize the current knowledge on the emerging role of O-GlcNAc modification in regulating autophagy-associated proteins and explain the different regulatory effects on autophagy exerted by O-GlcNAc modification.

The pathogenesis of depression is associated with synaptic impairment and dysfunction in autophagy processes. Mendelian randomization (MR) analysis revealed that six GWAS IDs revealed a significant association between Beclin-1 levels and depression risk. Besides, all SNPs had a positive effect on depression risk. Analyzing neurons from depressed individuals using single-cell RNA sequencing (scRNA-seq) uncovered decreased expression of AKT, mTOR, and genes linked to synaptic plasticity. The activation of the PI3K/AKT/mTOR signaling has been demonstrated to control autophagy and have a protective effect on the nervous system. Hydrogen sulfide (H2S) is an endogenous gasotransmitter that can potentially treat various neurological disorders by improving neuronal synaptic plasticity. However, whether H2S regulates autophagy through PI3K/AKT/mTOR signaling, improves neuronal synaptic plasticity damage, and plays an antidepressant role is unclear. Our current research revealed that the reduction in the expression of p-PI3K, p-AKT, and p-mTOR proteins increase in neuronal autophagy activity and decline synaptic plasticity in mice with depression induced by chronic unpredictable mild stress (CUMS). Treatment with the exogenous hydrogen sulfide donor NaHS for one day and continuous treatment for one week improved the depression-like behaviors in the mice. Compared with those after one day of NaHS treatment, the above protein expression levels were restored and maintained, and the antidepressant effect was more significant after one week of continuous treatment with NaHS. Moreover, the PI3K inhibitor LY294002 was used to demonstrate that NaHS suppresses autophagy through activating the PI3K/AKT/mTOR signaling and ameliorates synaptic plasticity impairments. This study provides novel insights into the antidepressant mechanisms of H2S, highlighting its antidepressant therapeutic potential.

Enhancer of zeste homolog 2 (EZH2), an epigenetic regulator that primarily inhibits downstream gene expression by tri-methylating histone H3, which is usually overexpressed in tumors and participates in many processes such as tumor occurrence and development, invasion, migration, drug resistance, and anti-tumor immunity as an oncogene, making it an important biomarker in cancer therapy. Collectively, several transcription factors and RNAs cooperate to facilitate the elevated expression of EZH2 in cancer. Although the significance of blocking EZH2 in cancer for inhibiting cancer progression is widely recognized, the clinical application of EZH2 inhibitors continues to encounter numerous challenges. In this review, drawing upon our comprehensive understanding of the factual underpinnings of EZH2's role in cancer, we aim to clarify the crucial importance of targeting EZH2 in cancer treatment. Furthermore, we summarize the current research landscape surrounding targeted EZH2 inhibitors and offer insights into potential future applications of these inhibitors.

The interaction between vitamin D and the immune system is perhaps the most well recognised extraskeletal facet of vitamin D, encompassing early studies of therapy for TB and leprosy through to more recent links with autoimmune disease. However, the spotlight on vitamin D and immune function has been particularly intense in the last five years following the COVID-19 pandemic. This was due, in part, to the many association studies of vitamin D status and COVID-19 infection and disease prognosis, as well as the smaller number of clinical trials of vitamin D supplementation. However, a potential role for vitamin D in COVID-19 also stemmed from the basic biology of vitamin D that provides a plausible mechanistic rationale for beneficial effects of vitamin D for improved immune health in the setting of respiratory infection. The aim of this review is to summarise the different strands of mechanistic evidence supporting a beneficial effect of vitamin D in COVID-19, how this was modified during the pandemic itself, and the potential new aspects of vitamin D and immune function that are likely to arise in the near future. Key topics that feature in this review are: antibacterial versus antiviral innate immune responses to 1,25-dihydroxyvitamin D (1,25(OH)2D); the function of immune 1α-hydroxylase (CYP27B1) activity and metabolism of 25-hydroxyvitamin D (25(OH)D) beyond antigen-presenting cells; advances in immune cell target gene responses to 1,25-dihydroxyvitamin D (notably changes in metabolic profile). Whilst much of the interest during the COVID-19 era has focused on vitamin D and public health, the continued evolution of our understanding of how vitamin D interacts with different components of the immune system continues to support a beneficial role for vitamin D in immune health.

Acute kidney injury (AKI) represents a rapid decline in kidney function, often associated with significant morbidity and mortality. Inefficient management of acute-phase inflammation and inadequate repair of established damage exacerbate AKI and facilitate its progression to chronic kidney disease (CKD). Platelet membrane (PM) has emerged as a promising targeting ligand in various studies. PM proteins can also facilitate the recruitment and differentiation of CD34+ cells (hematopoietic stem cells and endothelial progenitor cells) through both direct and indirect mechanisms, including enhancing adhesion of CD34+ cells to damaged tissues and elevating stromal cell-derived factor-1 (SDF-1) levels in ischemia-reperfusion injury (IRI) kidneys. In parallel, extensive research has demonstrated that rapamycin shows high potential as an anti-inflammatory therapy for AKI. Herein, we design a PM hybrid rapamycin liposome (Rapa@PM-Lipo), which not only improves the delivery efficiency of rapamycin, but also leverages the potential of PM to achieve long-lasting repair. Rapa@PM-Lipo significantly reduced Acute renal Tubular Necrosis (ATN) score in IRI kidneys following intravenous administration, both as a single and multiple doses. This study exploits the therapeutic potential of PM and explores its novel applications for facilitating tissue repair, presenting a promising strategy for the treatment of AKI and mitigating its progression to CKD.

Hepatic very low-density lipoprotein (VLDL) is essential for maintaining lipid metabolism in the liver. Sphingosine kinases (SphKs) are essential rate-limiting enzymes that catalyze sphingosine phosphorylation to Sphingosine-1-phosphate (S1P). SphKs exist as two isoforms, SphK1 and SphK2, both highly expressed in the liver. SphK1 plays a critical role in regulating hepatic inflammation and drug metabolism. This study aimed to determine whether SphK2 regulates hepatic lipid metabolism, particularly VLDL secretion. Immunohistochemical staining revealed decreased SphK2 protein levels within regions proximal to hepatic lipid accumulation in individuals diagnosed with metabolic dysfunction-associated steatotic liver disease (MASLD). Sphk2-/- mice exhibited spontaneous hepatocyte lipid accumulation and reduced VLDL secretion. Proteomic analysis revealed that SphK2 deficiency impaired soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNARE) complex interactions involved in vesicular transport and organelle membrane fusion. Furthermore, SphK2 deficiency results in accelerated degradation of the SEC22B, STX5A, and GS28 proteins via chaperone-mediated autophagy (CMA), impeding VLDL transport to the Golgi apparatus. MYH1485, a specific activator of mTOR, induces mTORC2 phosphorylation, thereby inhibiting the degradation of SNARE complexes by CMA and counteracting the lipid accumulation induced by SphK2 deficiency. Exogenous S1P supplementation markedly reversed the reduction in mTORC2 phosphorylation and suppressed CMA, thereby improving VLDL secretion. Our study elucidates an inventive regulatory mechanism by which SphK2 modulates CMA by activating mTORC2 phosphorylation, promoting VLDL secretion, and balancing lipid metabolism in the liver. These findings provide insights into SphK2 function and the underlying mechanisms involved in the regulation of VLDL secretion, which may facilitate MASLD treatment. Proposed model for the role of SphK2 in hepatic VLDL secretion. In hepatocytes, the inhibition of SphK2 activity decreased S1P production, which subsequently downregulates the mTORC2 pathway. This process accelerates the degradation of the SNARE complex components STX5A, GS28, and SEC22B via CMA, which regulates the mutual recognition between VTVs and the Golgi apparatus, ultimately reducing VLDL secretion in hepatocytes.

Biomolecular condensates are cellular compartments without enveloping membranes, enabling them to dynamically adjust their composition in response to environmental changes through post-translational modifications. Recent work has revealed that interferon-induced ADP-ribosylation (ADPr), which can be reversed by a SARS-CoV-2-encoded hydrolase, is enriched within a condensate. However, the identity of the condensate and the responsible host ADP-ribosyltransferase remain elusive. Here, we demonstrate that interferon induces ADPr through transcriptional activation of PARP14, requiring both the physical presence and catalytic activity of PARP14 for condensate formation. Interferon-induced ADPr colocalizes with PARP14 and its associated E3 ligase, DTX3L. These PARP14/ADPr condensates contain key components of p62 bodies-including the selective autophagy receptor p62, its binding partner NBR1 and the associated protein TAX1BP1, along with K48-linked and K63-linked polyubiquitin chains-but lack the autophagosome marker LC3B. Knockdown of p62 disrupts the formation of these ADPr condensates. Importantly, these structures are unaffected by autophagy inhibition, but depend on ubiquitination and proteasome activity. Taken together, these findings demonstrate that interferon triggers PARP14-mediated ADP-ribosylation in p62 bodies, which requires an active ubiquitin-proteasome system.

Stress memory is an adaptive mechanism that enables organisms to develop resilience in response to environmental changes. Among them, somatic stress memory is an important means for organisms to cope with contemporary repeated stress, and is accompanied by transcription memory. Sulfide is a common environmental pollutant; however, some organisms have adapted to survive in sulfur-rich environments. Urechis unicinctus is a sulfur-tolerant organism that enhances sulfide stress tolerance by establishing a somatic sulfide stress memory mechanism. However, the molecular mechanisms that regulate sulfide stress memory remain unclear. To explore whether epigenetics, which plays a role in the response of organisms to environmental stress, is involved in regulating somatic sulfide stress memory, we performed a combined analysis of DNA methylation and transcriptome data. We found that elevated levels of DNA methylation under repetitive sulfide stress regulated gene expression and resulted in enhanced sulfide stress tolerance in U. unicinctus, a phenomenon verified using DNA methylase inhibitors. Transcriptional memory can be induced in genes related to oxidative stress, regulation of autophagy, and maintenance of protein homeostasis by altering the level of DNA methylation to facilitate sulfide stress acclimation. Our results provide new insights into adaptive mechanisms to cope with environmental fluctuations.

Bangle, a perennial herb belonging to the ginger family with antiinflammatory properties, has been under-researched in ulcerative colitis. This study aimed to investigate the effects of Bangle extract (BaE) on inflammation and autophagy in the colons of mice with dextran sulfate sodium (DSS)-induced colitis. Male C57BL/6J mice were assigned to four groups: control, DSS + 0% BaE, DSS + 1% BaE, and DSS + 3% BaE. The BaE groups were fed BaE diets for 3 weeks, followed by an additional week of BaE diets and 3% DSS in the water. The control group received a standard chow diet and water for 4 weeks. Plasma leucine-rich α2-glycoprotein (LRG) levels, macrophage count, and the levels of nuclear factor kappa B (NFκB) p65, tumor necrosis factor-α (TNF-α), adenosine monophosphate-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), mechanistic target of rapamycin (mTOR), and autophagy markers were analyzed. In the DSS + 0% BaE group, LRG levels, macrophage count, NFκB p65 protein, and TNF-α mRNA levels were significantly higher compared to the control group. However, in the DSS + 3% BaE group, these levels were significantly reduced. Additionally, PGC-1α and phosphorylated AMPK levels were increased, while phosphorylated mTOR levels decreased, and autophagy marker microtubule-associated protein 1 light chain 3B (LC3B)-II levels were increased in the DSS + 3% BaE group. BaE may ameliorate colonic inflammation and upregulate autophagy via the modulation of the AMPK/mTOR/NFκB pathway in DSS-induced colitis.

Bovine viral diarrhoea (BVD), a condition triggered by bovine viral diarrhoea virus (BVDV), is recognized globally as a prevalent pathogen among ruminants and markedly affects the economics of animal husbandry. MicroRNAs, a class of small noncoding RNAs, play pivotal roles in regulating a myriad of biological processes.The ATG7-LC3 pathway, a canonical autophagy mechanism, is integral in defending against pathogenic invasion and maintaining cellular homeostasis.

In this study, we observed significant downregulation of bta-miR-221 in cells infected with BVDV. We further established that overexpression of bta-miR-221 markedly attenuated BVDV replication in Madin‒Darby bovine kidney (MDBK) cells. Through bioinformatics prediction analysis, we identified ATG7, an autophagy-related gene, as a direct downstream target of bta-miR-221. However, the intricate relationships among bta-miR-221, the ATG7-LC3 pathway, and BVDV infection remained unclear. Our study revealed that ATG7 expression was significantly elevated in BVDV-infected cells, whereas bta-miR-221 mimics repressed both endogenous and exogenous ATG7 expression. Following BVDV infection, we noted a decrease in LC3I expression, its conversion to LC3II, a significant increase in ATG7 expression, and a notable decrease in SQSTM1/p62 expression. By employing laser confocal microscopy and immunoprecipitation assays, we elucidated the regulation of the ATG7-LC3 pathway by bta-miR-221 in MDBK cells. Our findings recealed that BVDV infection enhanced the ATG7-LC3 interaction, inducing autophagy through the suppression of bta-miR-221 in MDBK cells. Consequently, bta-miR-221 emerged as a potent inhibitor of BVDV, impacting its proliferation and replication within the host.

This research sheds light on novel aspects of virus-host interactions and lays a foundation for the development of antiviral therapeutics.

The prevalence and spread of canine influenza virus (CIV) pose a threat to the health of dogs and humans. Some studies have shown that autophagy is closely related to virus replication, but the exact relationship between CIV replication and autophagy is still unclear. Therefore, this study investigated the effects of autophagy on CIV replication in vitro and in vivo. The data showed that CIV infection significantly caused respiratory tract damage in mice, upregulated the mRNA/protein levels of CIV replication-related genes and autophagy-related genes. In addition, the activation of autophagy by rapamycin (Rapa) significantly intensified the CIV replication and the respiratory tract damage of mice, while the inhibition of autophagy by 3-Methyladenine (3-MA) significantly alleviated these effects. Data of MDCK cells also demonstrated that CIV promoted self-replication through activating autophagy, and the upregulation of AKT/mTOR by insulin significantly inhibited the CIV replication. In summary, this study showed that CIV could promote self-replication by activating AKT/mTOR mediated autophagy, which provides new ideas for the prevention and treatment of canine influenza.

Traumatic brain injury (TBI) precipitates a neuroinflammatory cascade, with the NLRP3 inflammasome emerging as a critical mediator. This review scrutinizes the complex activation pathways of the NLRP3 inflammasome by underscoring the intricate interplay between calcium signaling, mitochondrial disturbances, redox imbalances, lysosomal integrity, and autophagy. It is hypothesized that a combination therapy approach-integrating NF-κB pathway inhibitors with NLRP3 inflammasome antagonists-holds the potential to synergistically dampen the inflammatory storm associated with TBI.

A comprehensive analysis of literature detailing NLRP3 inflammasome activation pathways and therapeutic interventions was conducted. Empirical evidence supporting the concurrent administration of MCC950 and Rapamycin was reviewed to assess the efficacy of dual-action strategies compared to single-agent treatments.

Findings highlight potassium efflux and calcium signaling as novel targets for intervention, with cathepsin B inhibitors showing promise in mitigating neuroinflammation. Dual therapies, particularly MCC950 and Rapamycin, demonstrate enhanced efficacy in reducing neuroinflammation. Autophagy promotion, alongside NLRP3 inhibition, emerges as a complementary therapeutic avenue to reverse neuroinflammatory damage.

Combination therapies targeting the NLRP3 inflammasome and related pathways offer significant potential to enhance recovery in TBI patients. This review presents compelling evidence for the development of such strategies, marking a new frontier in neuroinflammatory research and therapeutic innovation.

Renal fibrosis is a common pathological process of progressive chronic kidney disease (CKD). However, effective therapy is constrained currently. Autophagy is an important mechanism in kidney injury and repairment but its exact role in renal fibrosis was discrepant according to previous studies. Sulforaphane (SFN), a natural plant compound, has been explored as a promising nutritional therapy for a variety of diseases. But the salutary effect and underlying mechanism of SFN on CKD have not been fully elucidated. In this study, we investigated the effect of SFN on renal fibrosis in unilateral ureteral obstruction (UUO) mice. Then we examined the regulatory effect of SFN on autophagy-related proteins in renal fibroblasts and renal tubular epithelial cells. Our results showed that sulforaphane could significantly alleviate renal fibrosis in UUO mice. In vitro, the expression levels of autophagy-related protein showed that SFN could upregulate the autophagy activity of renal interstitial fibroblasts and downregulate the autophagy activity of renal tubular epithelial cells. Furthermore, we found that phosphorylated mTOR protein levels was reduced in renal fibroblasts and increased in renal tubular epithelial cells after SFN treatment. Our results strongly suggested that SFN could alleviate renal fibrosis through dual regulation of mTOR-mediated autophagy pathway. This finding may provide a new perspective on the renal salutary effect of SFN and provide a preclinical rationale for exploring the therapeutic potential of SFN to slow down renal fibrosis.

Various forms of tissue damage can lead to fibrosis, an abnormal reparative reaction. In the industrialized countries, 45% of deaths are attributable to fibrotic disorders. Autophagy is a highly preserved process. Lysosomes break down organelles and cytoplasmic components during autophagy. The cytoplasm is cleared of pathogens and dysfunctional organelles, and its constituent components are recycled. With the growing body of research on autophagy, it is becoming clear that autophagy and its associated mechanisms may have a role in the development of numerous fibrotic disorders. However, a comprehensive understanding of autophagy in fibrosis is still lacking and the progression of fibrotic disease has not yet been thoroughly investigated in relation to autophagy‑associated processes. The present review focused on the latest findings and most comprehensive understanding of macrophage autophagy, endoplasmic reticulum stress‑mediated autophagy and autophagy‑mediated endothelial‑to‑mesenchymal transition in the initiation, progression and treatment of fibrosis. The article also discusses treatment strategies for fibrotic diseases and highlights recent developments in autophagy‑targeted therapies.

The 5‑year survival rate of patients with esophageal squamous cell carcinoma (ESCC) is <20%, highlighting the need for the development of novel therapeutic targets. Neuregulin‑1 (NRG1), a transmembrane protein involved in cell proliferation and survival signaling, has unclear biological functions and clinical value in ESCC. The present study investigated the association between NRG1 expression and ESCC by analyzing data from both patients with ESCC and The Cancer Genome Atlas database. Reverse transcription‑quantitative PCR and immunohistochemistry staining were used to determine the levels of gene and protein in the tissue. The findings revealed that NRG1 gene and protein levels were significantly higher in tumor tissues compared with the normal tissues. Elevated expression of NRG1 was associated with poor outcomes, particularly in patients with advanced ESCC. Silencing NRG1 decreased both its mRNA and protein levels, disrupting key signaling pathways, such as phosphorylated (p‑)AKT and cellular rapidly accelerated fibrosarcoma (p‑cRAF), which led to decreased cancer cell proliferation, migration and tumor sphere formation, along with increased cell death. High expression levels of NRG1 and cRAF were significantly associated with poor prognosis. Additionally, silencing NRG1 promoted autophagosome and autolysosome formation, decreasing LC3B levels. The use of the autophagy inhibitor chloroquine significantly enhanced cell death induced by NRG1 silencing, suggesting that autophagy functions as a survival mechanism in ESCC cells in which NRG1 is silenced. Furthermore, high co‑expression of NRG1 and LC3B was associated with a worse prognosis. On the whole, the present study demonstrated that targeting NRG1 with autophagy inhibitors may serve as a potential therapeutic strategy for ESCC.

Chronic Obstructive Pulmonary Disease (COPD) is a poorly reversible respiratory disorder distinguished by dyspnea, cough, expectoration and exacerbations due to abnormality of airways or emphysema. In this review, we consider the therapeutic potential of targeting Mammalian target of Rapamycin (mTOR) for treating COPD. The mTOR is a highly conserved serine-threonine protein kinase that integrates signals from growth factors and nutrients to control protein synthesis, lipid biogenesis and metabolism. Dysregulated mTOR pathway signaling due to genetic factors or cigarette smoking impairs autophagy, driving the buildup of abnormal cells and damaged proteins, resulting in inflammation and oxidative stress. Persistent mTOR activation also contributes to pulmonary vascular cell proliferation, facilitating the development of pulmonary resistance in COPD. Rapamycin, an inhibitor of mTOR, prevents the buildup of senescent cells in the lungs of COPD patients and inhibits the release of lung tissue-damaging proteases. mTOR also impacts the corticosteroid sensitivity in COPD patients by regulating the levels of histone deacetylases. The emerging role of gut-lung axis dysbiosis in the progression of COPD and its influence on mTOR further highlights the relevance of the mTOR pathway in COPD pathophysiology.

To investigate the effects of Huayu Qutan recipe (, HYQT) on the atherosclerosis (AS) model of ApoE-/- mice with a high-fat diet and to illustrate the underlying mechanisms from modern patho-physiological conceptualizations.

High performance liquid chromatography of quadrupole time of flight-tandem mass spectrometry (HPLC-Q-TOF-MS/MS) analysis was used to identify the active compounds in the recipe. The mice were randomly allocated into 7 groups: control (CTRL) group, normal diet (ND) group, high-fat diet (HFD) group, HYQT groups (low dose, medium dose, and high dose), and simvastatin (SIM) group. Deferent doses of HYQT were gavaged twice a day, and then the protective effect of HYQT on plaque formation in ApoE-/- mice with a high-fat diet was verified viahematoxylin-eosin (HE) staining and oil red o (ORO) staining. We observed the co-localization in aortic macrophages and lipid droplets (LDs) by CD68 and the Bodipy fluorescence probe. Light chain 3 phosphoprotein class Ⅱ/light chain 3 phosphoprotein class Ⅰ (LC3Ⅱ/LC3Ⅰ) was examined by western blotting, and sequestosome 1 (SQSTM1/p62), Beclin1, Lamp1, mammalian target of rapamycin (mTOR), phosphorylated mammalian target of rapamycin (p-mTOR), and ATP-binding cassette transporter A1 (ABCA1) were examined by real-time polymerase chain reaction (RT-PCR) and Western blotting. Transcription factor EB (TFEB) nuclear translocation was determined by immunofluorescence analysis.

Five active compounds were identified using HPLC-Q-TOF-MS/MS analysis: ferulic acid, chlorogenic acid, calycosin, formononetin, and 8,2'-dihydroxy-7,4'-dimethoxy-isoflavane. The effect of HYQT on atherosclerotic plaque formation in ApoE-/- mice was investigated. These findings showed that HYQT decreased the co-localization of CD68 and Bodipy and increased the co-localization of CD68 and LC3B. Medium and high doses of HYQT increased autophagosome formation and promoted the maturation of LC3Ⅱ/LC3Ⅰ. Additionally, HYQT decreased the expression of SQSTM1/p62. Medium and high doses of HYQT also increased the expression of Beclin1 and Lamp1. RT-PCR and Western blot results suggested that HYQT enhanced the expression of ABCA1 mRNA and protein and regulated the mTORC1/TFEB signaling pathway.

The results indicate that HYQT is an effective traditional Chinese herbal remedy for the treatment of AS. HYQT mitigates macrophage-derived foam cell formation by activating autophagy in atherosclerosis. The mTOR/TFEB signaling pathway and ABCA1 are therapeutic targets of HYQT for the treatment of AS.

The abnormal effector function of CD4+ T cells plays a key role in the pathogenesis of Sjogren's syndrome (SS) and its associated systematic autoimmune response. Cellular metabolism, including glucose metabolism, lipid metabolism and amino acid metabolism, supports proliferation, migration, survival and differentiation into distinct CD4+ T-cell subsets. Different subtypes of T cells have significantly different demands for related metabolic processes, which enables us to finely regulate CD4+ T cells through different metabolic processes in autoimmune diseases such as SS. In this review, we summarize the effects of disturbances in distinct metabolic processes, such as glycolysis, fatty acid metabolism, glutamine decomposition, mitochondrial dynamics, and ferroptosis, on how to support the effector functions of CD4+ T cells in the SS. We also discuss potential drugs with high value in the treatment of SS through metabolic normalization in CD4+ T cells. Finally, we propose possible directions for future targeted therapy for immunometabolism in SS.

Metabolic dysfunction-associated steatohepatitis (MASH), a severe form of metabolic dysfunction-associated steatotic liver disease (MASLD), involves hepatic lipid accumulation, inflammation, and fibrosis. It can progress to cirrhosis or hepatocellular carcinoma without timely treatment. Current treatment options for MASH are limited. This study explores the therapeutic effects of fluorofenidone (AKF-PD), a novel small-molecule compound with antifibrotic and anti-inflammatory properties, on MASH in mouse model. Mice fed a choline-deficient, l-amino acid-defined, high-fat diet (CDAHFD) were treated with AKF-PD, resulting in reduced serum ALT, AST, hepatic lipid accumulation, liver inflammation, and fibrosis. Network pharmacology and RNA-sequencing analyses suggested that AKF-PD influenced multiple metabolic, inflammatory, and fibrosis-related pathways. Further experiments verified that AKF-PD activated hepatic AMPK signaling, leading to the inhibition of the downstream SREBF1/SCD1 pathway and the activation of autophagy. Additionally, AKF-PD suppressed the expression of various inflammatory factors, reduced macrophage infiltration, and inhibited NLRP3 inflammasome activation. Moreover, AKF-PD attenuated liver fibrosis by inhibiting TGFβ1/SMAD signaling. In conclusion, this study reveals that AKF-PD effectively decreases hepatic lipid accumulation, liver inflammation and fibrosis in a CDAHFD-induced MASH model, positioning AKF-PD as a promising candidate for the treatment of MASH.

Activating PIK3CA mutations, present in up to 40% of hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (Her2-) breast cancer (BC) patients, can be effectively targeted with the alpha isoform-specific PI3K inhibitor Alpelisib. This treatment significantly improves outcomes for HR+, Her2-, and PIK3CA-mutated metastatic BC patients. However, acquired resistance, often due to aberrant activation of the mTOR complex 1 (mTORC1) pathway, remains a significant clinical challenge. Our study, using in vitro and orthotopic xenograft mouse models, demonstrates that constitutively active mTORC1 signaling renders PI3K inhibitor-resistant BC exquisitely sensitive to various drugs targeting cancer metabolism. Mechanistically, mTORC1 suppresses the induction of autophagy during metabolic perturbation, leading to energy stress, a critical depletion of aspartate, and ultimately cell death. Supporting this mechanism, BC cells with CRISPR/Cas9-engineered knockouts of canonical autophagy genes showed similar vulnerability to metabolically active drugs. In BC patients, high mTORC1 activity, indicated by 4E-BP1T37/46 phosphorylation, correlated with p62 accumulation, a sign of impaired autophagy. Together, these markers predicted poor overall survival in multiple BC subgroups. Our findings reveal that aberrant mTORC1 signaling, a common cause of PI3K inhibitor resistance in BC, creates a druggable metabolic vulnerability by suppressing autophagy. Additionally, the combination of 4E-BP1T37/46 phosphorylation and p62 accumulation serves as a biomarker for poor overall survival, suggesting their potential utility in identifying BC patients who may benefit from metabolic therapies.

To investigate the effect of Didang Decoction-medicated serum on autophagy in high glucose (HG)-induced rat glomerular endothelial cells (RGECs) and explore the pathway that mediates its effect.

Primary RGECs were isolated and cultured using sequential sieving combined with collagenase digestion, followed by identification using immunofluorescence assay for factor VIII. High glucose medium was used to induce RGECs to simulate a diabetic environment, and the effects of Didang Decoction-medicated serum and 3-MA (an autophagy inhibitor), either alone or in combination, on autophagy of HG-exposed cells were evaluated by observing autophagic vacuoles using monodansylcadaverine (MDC) staining. RT-qPCR and Western blotting were employed to measure mRNA and protein expression levels of Beclin-1, p62, LC3B, p-PI3K, p-Akt, and p-mTOR.

Compared with the control cells, the HG-exposed RGECs showed significantly reduced autophagic fluorescence intensity, decreased Beclin-1 mRNA expression, increased p62 mRNA expression, downregulated Beclin-1 protein and LC3-II/I ratio, and upregulated p62, p-PI3K, p-Akt, and p-mTOR protein levels. Didang Decoction-medicated serum significantly enhanced autophagic fluorescence intensity in HG-exposed cells, increased Beclin-1 mRNA expression, decreased p62 mRNA expression, upregulated Beclin-1 protein, and downregulated p62, p-PI3K, p-Akt, and p-mTOR protein levels.

Didang Decoction-medicated serum enhances autophagy in HG-exposed RGECs by regulating the PI3K/Akt/mTOR signaling pathway, which sheds light on a new therapeutic strategy for diabetic nephropathy.

Rescue therapies of relapsed/refractory (r/r) Hodgkin's lymphoma (HL) in the third to sixth-line provide major, yet unresolved problems. The MEPED regimen includes nuclear receptor agonists such as pioglitazone and dexamethasone, which counterbalance HL homeostasis, HL stress response inhibitors, everolimus and COX-2 inhibitor, and a stress response inducer, low-dose metronomic treosulfan. CR (six of seven patients) and long-term cCR in patients receiving no consolidating allogeneic stem cell transplantation highlight MEPED as a potent salvage therapy in advanced refractory HL. MEPED edits everolimus activities in such a way that mTORC1 becomes a non-oncogene addiction bottleneck, hence determining long-term therapy outcome. The implications of the therapeutic paradigm shift toward editing of HL tissue, and particularly mTOR addiction, could prove to be profound for clinical practice, both in terms of outcome and treatment tolerability. The long-term results of MEPED treatment indicate the urgent evaluation of the schedule in a multicenter trial for r/r HL.

Cerebral ischemia is a prevalent cerebrovascular disorder, with the restoration of blocked blood vessels serving as the current standard clinical treatment. However, reperfusion can exacerbate neuronal damage and neurological dysfunction, resulting in cerebral ischemia-reperfusion (I/R) injury. Presently, clinical treatment strategies for cerebral I/R injury are limited, creating an urgent need to identify new effective therapeutic targets. The PI3K/AKT signaling pathway, a pro-survival pathway associated with cerebral I/R injury, has garnered significant attention. We conducted a comprehensive review of the literature on the PI3K/AKT pathway in the context of cerebral I/R. Our findings indicate that activation of the PI3K/AKT signaling pathway following cerebral I/R can alleviate oxidative stress, reduce endoplasmic reticulum stress (ERS), inhibit inflammatory responses, decrease neuronal apoptosis, autophagy, and pyroptosis, mitigate blood-brain barrier (BBB) damage, and promote neurological function recovery. Consequently, this pathway ultimately reduces neuronal death, alleviates brain tissue damage, decreases the volume of cerebral infarction, and improves behavioral impairments. These results suggest that the PI3K/AKT signaling pathway is a promising therapeutic target for further research and drug development, holding significant potential for the treatment of cerebral I/R injury.

Huntington's disease (HD) is caused by expansion of the polyglutamine stretch in huntingtin protein (HTT) resulting in hallmark aggresomes/inclusion bodies (IBs) composed of mutant huntingtin protein (mHTT) and its fragments. Stimulating autophagy to enhance mHTT clearance is considered a potential therapeutic strategy for HD. Our recent evaluation of the autophagic-lysosomal pathway (ALP) in human HD brain reveals upregulated lysosomal biogenesis and relatively normal autophagy flux in early Vonsattel grade brains, but impaired autolysosome clearance in late grade brains, suggesting that autophagy stimulation could have therapeutic benefits as an earlier clinical intervention. Here, we tested this hypothesis by crossing the Q175 HD knock-in model with our autophagy reporter mouse TRGL ( T hy-1- R FP- G FP- L C3) to investigate in vivo neuronal ALP dynamics. In the Q175 and/or TRGL/Q175 mice, mHTT was detected in autophagic vacuoles and also exhibited a high level of colocalization with autophagy receptors p62/SQSTM1 and ubiquitin in the IBs. Compared to the robust lysosomal pathology in late-stage human HD striatum, ALP alterations in Q175 models are also late-onset but milder that included a lowered phospho-p70S6K level, lysosome depletion and autolysosome elevation including more poorly acidified autolysosomes and larger-sized lipofuscin granules, reflecting impaired autophagic flux. Administration of a mTOR inhibitor to 6-mo-old TRGL/Q175 normalized lysosome number, ameliorated aggresome pathology while reducing mHTT-, p62- and ubiquitin-immunoreactivities, suggesting the beneficial potential of autophagy modulation at early stages of disease progression.

Emerging evidences suggest that autophagy, a key cellular process responsible for degrading and recycling damaged organelles and proteins, plays a crucial role in maintaining neuronal health. Dysfunctional autophagy has been linked to the pathogenesis of Alzheimer's disease (AD), contributing to the accumulation of misfolded proteins and cellular debris. Molecular mechanisms underlying autophagy dysfunction in AD involve amyloid-beta (Aβ) and tau accumulation, neuroinflammation, mitochondrial dysfunction, oxidative stress and endoplasmic reticulum stress. Disrupted signaling pathways such as TRIB3, Nmnat and BAG3 that regulate key processes like autophagosome initiation, lysosome function, and protein homeostasis also play a crucial role in the pathogenesis. Restoration of autophagy by modulating these molecular and signaling pathways may be an effective therapeutic strategy for AD. Studies have found few drugs targeting autophagy dysregulation in AD. These drugs include metformin that has been found to modulate the expression of TRIB3 for autophagy regulation. Another drug, resveratrol has been reported to augment the activity of Nmnat thus, increases autophagy flux. BACE1 and mTOR inhibitors like arctigenin, nilvadipine and dapagliflozin were also found to restore autophagy. This study elaborates recent advances in signaling and molecular pathways and discusses current and emerging therapeutic interventions targeting autophagy dysfunction in AD.

Among all malignancies, colorectal cancer ranks third in incidence rate and second in mortality rate. Human beta-defensin-1 (hBD-1) has broad-spectrum antimicrobial properties, and it plays an important role in the tumor microenvironment. Circular ribonucleic acids (circRNAs) regulate the proliferation and progression of colorectal cancer cells via cancer-related signaling pathways.

Cell proliferation was assessed using the Cell Counting Kit-8 assay to determine the optimal hBD-1 concentration. Intracellular autophagic vesicles were visualized via monodansylcadaverine staining. In addition, the levels of AKT and mammalian target of rapamycin (mTOR)-associated signaling proteins were analyzed via Western blot analysis. CircRNA microarrays and quantitative real-time polymerase chain reaction were used to identify differentially expressed circRNAs in colon cancer cell lines. The functional role of HAS-CIRCpedia-5280 in vitro was demonstrated by overexpressing HAS-CIRCpedia-5280 and inhibiting miR-4712-5p. HAS-CIRCpedia-5280 could be a sponge of miR-4712-5p, mimicking the effect induced by HAS-CIRCpedia-5280 overexpression in colon cancer cells.

hBD-1 inhibited the proliferation of colon cancer cells and increased the number of intracellular autophagic vesicles. In addition, hBD-1 inhibited the AKT/mTOR signaling pathway, thereby enhancing cellular autophagy. Further, the interaction of HAS-CIRCpedia-5280 and miR-4712-5p was investigated. hBD-1 upregulated the expression level of HAS-CIRCpedia-5280 and downregulated the expression level of miR-4712-5p in colon cancer cells. Subsequently, the overexpression of HAS-CIRCpedia-5280 or the inhibition of miR-4712-5p activated the AKT/mTOR signaling pathway, leading to cellular autophagy inhibition. Conversely, the mimicry of miR-4712-p counteracted the effect of HAS-CIRCpedia 5280 overexpression in colon cancer cells by inhibiting the activation of the AKT/mTOR signaling pathway and, thereby, enhancing cellular autophagy.

hBD-1 can have an inhibitory effect against cell proliferation in colon cancer SW-620/HCT-116 cells via the HAS-CIRCpedia-5280/miR-4712-5p-mediated activation of autophagy.

Diabetic kidney disease (DKD) is one of the complications of diabetes mellitus, which triggers kidney fibrosis and eventually develops into end-stage renal disease. Nuciferine (NF) is one of the most important functional components in lotus leaves (LL), but its role and mechanism for the treatment of DKD are unclear. A high-fat-diet (HFD)-induced DKD model in KK-AY mice was established in this study. NF treatment significantly improved blood glucose and blood biochemical indices in DKD mice. Furthermore, NF reduced the levels of mALB, UCRE, Scr, and BUN in mice urine. Further, the extent of renal lesions in the mice in this study was at stage IV according to the Mogensen staging method. NF treatment was effective in ameliorating renal injury during this period. Concurrently, the protein levels of FN, N-cadherin, TGFβ, p-Smad3, p-PI3K, p-AKT, p-mTOR, and p62 were decreased. In contrast, the level of expression of Beclin-1 was increased. In the high glucose-exposed HK-2 cell model, the expression of p-PI3K, p-AKT, and p-mTOR was all downregulated, and autophagy proteins were increased after NF intervention. In addition, HK-2 cells were treated with high glucose in combination with Wortmannin and 3-MA, respectively. The results demonstrated that NF inhibited the expression of TGFβ and p-Smad3 by regulating autophagy through the PI3K-AKT-mTOR pathway, thereby ameliorating renal fibrosis at stage IV in mice. Therefore, LL can be used as a dietary component for the prevention of renal fibrosis in DKD patients.

Adenomyosis (AM) is a common and challenging disease in gynecological clinics, which adversely affects women's physical and mental health. Despite the growing number of studies, the mechanisms associated with the growth of the lesion are poorly understood. Studies show that abnormal proliferation, apoptosis, and migration in ectopic endometrial stromal cells (EESc) of AM may contribute to the development and progression of AM. Understanding the underlying molecular mechanisms can significantly contribute to diagnosing and treating AM. In the present study, EESc was isolated and cultured from the ectopic endometrium of patients with AM. These cells were treated with a PI3K/AKT activator (740 Y-P) and an inhibitor (LY294002), while the expression of microRNA-21 (miR-21) was interfered with. The effects of miR-21 on the apoptosis and autophagy of EESc, as well as the associated mechanisms, were investigated from multiple perspectives. Here, we found that 740 Y-P could significantly promote proliferation, inhibit apoptosis of EESc, and increase the expression of mTOR and p-mTOR proteins in EESc. Moreover, activating miR-21 enhanced the pro-migration effect of 740 Y-P and reversed the pro-apoptotic effect of LY294002, reducing the apoptosis rate and increasing the migration ability of EESc. Our investigation revealed that miR-21 can inhibit apoptosis and autophagy and promote migration of EESc. This effect is likely mediated via the PI3K/AKT/mTOR pathway.

The adverse effects of elevated temperatures on human health are becoming progressively severe. This research established a mouse model of cognitive dysfunction induced by heat stress to examine the impact of omega-3 PUFAs on the cognitive capabilities of heat-stressed mice. The study also aimed to elucidate the role and potential mechanisms of autophagy regulation in cognitive enhancement through omega-3 PUFAs interventions. Administration of omega-3 PUFAs ameliorated cognitive deficits in heat-stressed mice and increased brain concentrations of these fatty acids. Notably, omega-3 PUFAs significantly protected hippocampal neurons' morphology, quantity, and synaptic architecture in heat-stressed mice. Additionally, omega-3 PUFAs intake reduced the prevalence of damaged mitochondria in the hippocampus and mitigated oxidative harm. Further investigation revealed that heat stress induces autophagy. However, the autophagic process becomes dysfunctional, leading to impaired autophagic activity. Omega-3 PUFAs supplementation markedly augmented hippocampal autophagy in the heat-stressed mice. Moreover, heat stress upregulated the phosphorylation of the PI3K-Akt-mTOR pathway in both the mouse hippocampus and HT22 cells. In contrast, omega-3 PUFAs intake significantly diminished the phosphorylation levels within this pathway, alleviating the autophagic fusion barrier imposed by heat stress and promoting autophagic flux. The findings suggest that omega-3 PUFAs supplementation during heat stress may bolster autophagic function by inhibiting the phosphorylation of the PI3K-Akt-mTOR pathway. This modulation reduces structural and oxidative stress damage, ultimately enhancing cognitive function in mice subjected to heat stress.

This study aims to explore the therapeutic efficacy of evodiamine (EVO) in the treatment of dry eye disease (DED).

Mouse models of DED was developed using benzalkonium chloride eye drops and subcutaneous atropine injections. Corneal epithelial defects were assessed by fluorescein sodium staining, and tear secretion was measured with the phenol red thread test. For the in vitro model, human corneal epithelial cells were cultured in a sodium chloride-enriched medium. Phenotypic and mechanistic analyses were conducted using real-time quantitative PCR, Western blotting, flow cytometry, and immunofluorescence staining.

The administration of EVO eye drops significantly enhanced tear secretion in mice, ameliorated ocular surface damage, decreased the expression of corneal inflammatory factors, and increased the density of conjunctival goblet cells. Furthermore, EVO reduced oxidative stress by promoting autophagy. Mechanistically, EVO-induced autophagy was mediated via the p53/mammalian target of rapamycin pathway.

These findings suggest that EVO is a potential therapeutic agent for the treatment of DED, with its beneficial effects attributed to the activation of autophagy through the p53/mammalian target of rapamycin pathway.

The activation of STING1 can lead to the production and secretion of cytokines, initiating antitumor immunity. Here, we screened an ion channel ligand library and identified tetrandrine, a bis-benzylisoquinoline alkaloid, as an immunological adjuvant that enhances antitumor immunity by preventing the autophagic degradation of the STING1 protein. This tetrandrine effect is independent of its known function as a calcium or potassium channel blocker. Instead, tetrandrine inhibits lysosomal function, impairing cathepsin maturation, and autophagic degradation. Proteomic analysis of lysosomes identified TAX1BP1 as a novel autophagic receptor for the proteolysis of STING1. TAX1BP1 recognizes STING1 through the physical interaction of its coiled-coil domain with the cyclic dinucleotide binding domain of STING1. Systematic mutation of lysine (K) residues revealed that K63-ubiquitination of STING1 at the K224 site ignites TAX1BP1-dependent STING1 degradation. Combined treatment with tetrandrine and STING1 agonists promotes antitumor immunity by converting "cold" pancreatic cancers into "hot" tumors. This process is associated with enhanced cytokine release and increased infiltration of cytotoxic T-cells into the tumor microenvironment. The antitumor immunity mediated by tetrandrine and STING1 agonists is limited by neutralizing antibodies to the type I interferon receptor or CD8+ T cells. Thus, these findings establish a potential immunotherapeutic strategy against pancreatic cancer by preventing the autophagic degradation of STING1.

Spermine (SPM) and spermidine (SPD) are polyamines found in all organisms, and their concentrations can be regulated by ingestion. We demonstrated that yeast-fermented garlic (YF) extract significantly increased autophag flux in OUMS-36T-1 and HeLa cells expressing the fluorescent probe (GFP-LC3-RFP-LC3ΔG). YF-induced increase of autophagy occurred independently of mTORC1 signaling, and RNA-sequencing analysis revealed that EGR1 was the most significantly altered gene in YF-treated OUMS-36T-1 cells. YF-treated EGR1-deficient HAP1 cells displayed reduced autophagic flux (p < 0.05). YF-induced increasing of autophagic flux occurred via a specific SPM/SPD ratio. HAP1 cells treated with equivalent amounts of SPD or SPM as that found in YF did not increase autophagic flux (p > 0.05); however, treatment with SPD and SPM in the same ratio as that found in YF increased autophagic flux (p < 0.05). This specific SPM/SPD ratio reduced MG132-induced proteostress via EGR1-dependent pathways (p < 0.05). Thus, the SPM/SPD balance may regulate autophagy via EGR1-dependent pathways, and controlling this balance may provide a strategy to maintain cellular homeostasis.

Chronic atrophic gastritis (CAG) is a critical initial step in gastric cancer tumorigenesis accompanied by high malignancy. Erianin has been proposed as a promising agent in treating precancerous lesions of gastric cancer. Considering that little work has been implemented concerning the specific role and possible regulatory mechanism of Erianin in CAG, the goal of the study is to disclose the effects and mechanism of erianin on the malignant transformation in the process of CAG. CAG cell model was generated in human gastric epithelium GES-1 cells induced by Nmethyl-N'-nitro-N-nitrosoguanidine (MNNG). CCK-8 method determined cell viability. ELISA and corresponding assay kits severally appraised the contents of inflammatory cytokines and oxidative stress markers. Cellular reactive oxygen species (ROS) formation was measured by flow cytometry analysis using DCFH-DA probe. GFP-LC3 immunofluorescence staining and Western blotting evaluated autophagy. Also, Western blotting analyzed the expression of components in mitogen activated protein kinase (MAPK)/mechanistic target of rapamycin (mTOR) signaling. The results manifested that MNNG treatment diminished the viability and autophagy whereas intensified the inflammation and oxidative stress in GES-1 cells, which were all reversed by Erianin. Besides, Erianin blocked mTOR/MAPK signaling in MNNG-exposed GES-1 cells. Autophagy inhibitor 3-methyladenine (3-MA) or p38 MAPK agonist asiatic acid partially counteracted the protection elicited by Erianin against viability loss, inflammatory reaction as well as oxidative stress in MNNG-induced GES-1 cells. Combined with the findings, Erianin might mediate autophagy to improve MNNG-elicited CAG via MAPK/mTOR signaling.

In most types of erectile dysfunction, particularly in advanced stages, typical pathological features observed are reduced parenchymal cells coupled with increased tissue fibrosis. However, the current treatment methods have shown limited success in reversing these pathologic changes. Recent research has revealed that changes in autophagy levels, along with alterations in apoptosis and fibrosis-related proteins, are linked to the progression of erectile dysfunction, suggesting a significant association. Autophagy, known to significantly affect cell fate and tissue fibrosis, is currently being explored as a potential treatment modality for erectile dysfunction. However, these present studies are still in their nascent stage, and there are limited experimental data available. This review analyzes erectile dysfunction from a pathological perspective. It provides an in-depth overview of how autophagy is involved in the apoptotic processes of smooth muscle and endothelial cells and its role in the fibrotic processes occurring in the cavernosum. This study aimed to develop a theoretical framework for the potential effectiveness of autophagy in preventing and treating erectile dysfunction, thus encouraging further investigation among researchers in this area.

Autophagy shapes CD8 T cell fate; yet the timing, triggers and targets of this process are poorly defined. Herein, we show that naive CD8 T cells have high autophagic flux, and we identify an autophagy checkpoint whereby antigen receptor engagement and inflammatory cytokines acutely repress autophagy by regulating amino acid transporter expression and intracellular amino acid delivery. Activated T cells with high levels of amino acid transporters have low autophagic flux in amino-acid-replete conditions but rapidly reinduce autophagy when amino acids are restricted. A census of proteins degraded and fueled by autophagy shows how autophagy shapes CD8 T cell proteomes. In cytotoxic T cells, dominant autophagy substrates include cytolytic effector molecules, and amino acid and glucose transporters. In naive T cells, mitophagy dominates and selective mitochondrial pruning supports the expression of molecules that coordinate T cell migration and survival. Autophagy thus differentially prunes naive and effector T cell proteomes and is dynamically repressed by antigen receptors and inflammatory cytokines to shape T cell differentiation.

Metabolic reprogramming is pivotal in cancer stem cell (CSC) self-renewal. However, the intricate regulatory mechanisms governing the crosstalk between metabolic reprogramming and liver CSCs remain elusive. Here, using a metabolic CRISPR-Cas9 knockout screen, we identify ATP6V1D, a subunit of the vacuolar-type H+-translocating ATPase (V-ATPase), as a key metabolic regulator of hepatocellular carcinoma (HCC) stemness. Elevated ATP6V1D expression correlates with poor clinical outcomes in HCC patients. ATP6V1D knockdown inhibits HCC stemness and malignant progression both in vitro and in vivo. Mechanistically, ATP6V1D enhances HCC stemness and progression by maintaining macroautophagic/autophagic flux. Specifically, ATP6V1D not only promotes lysosomal acidification, but also enhances the interaction between CHMP4B and IST1 to foster ESCRT-III complex assembly, thereby facilitating autophagosome-lysosome fusion to maintain autophagic flux. Moreover, silencing CHMP4B or IST1 attenuates HCC stemness and progression. Notably, low-dose bafilomycin A1 targeting the V-ATPase complex shows promise as a potential therapeutic strategy for HCC. In conclusion, our study highlights the critical role of ATP6V1D in driving HCC stemness and progression via the autophagy-lysosomal pathway, providing novel therapeutic targets and approaches for HCC treatment.Abbreviations: 3-MA: 3-methyladenine; ANT: adjacent normal liver tissues; ATP6V1D: ATPase H+ transporting V1 subunit D; BafA1: bafilomycin A1; CHMP: charged multivesicular body protein; co-IP: co-immunoprecipitation; CSC: cancer stem cell; ESCRT: endosomal sorting complex required for transport; HCC: hepatocellular carcinoma; IF: immunofluorescence; IHC: immunohistochemical; LCSCs: liver cancer stem cells; qRT-PCR: quantitative real time PCR; V-ATPase: vacuolar-type H+- translocating ATPase; WB: western blot.