Alzheimer's disease (AD) is characterized by amyloid-β (Aβ) accumulation, impaired proteostatic clearance, and oxidative damage, all of which contribute to neuronal dysfunction and disease progression. Fucoxanthin (FX), a marine-derived carotenoid abundant in brown algae, has shown antioxidant and neuroprotective potential. However, its role in autophagy-lysosome dysfunction and ferroptosis-associated oxidative injury under amyloidogenic conditions remains unclear. In this study, the effects of FX were investigated in APP Swedish mutant-expressing Neuro2a (SweAPP N2a) cells treated with 0.1-5 μM FX and in 5XFAD transgenic mice orally administered FX at 200 mg kg-1. FX treatment increased LC3-II expression and reduced p62 accumulation in SweAPP N2a cells, indicating enhanced autophagic degradation. FX also increased the expression of the lysosomal markers LAMP1 and cathepsin D (CTSD), suggesting enhanced lysosome-associated degradative capacity. These responses were accompanied by AMPK activation and suppression of mTOR signaling, together with increased autophagic flux as confirmed by bafilomycin A1-based analysis. Moreover, FX significantly reduced intracellular ROS levels and lipid peroxidation marker 4-hydroxynonenal (4-HNE), while modulating ferroptosis-associated proteins, including GPX4 and FTH1. Consistent with the cellular findings, FX administration in 5XFAD mice modulated autophagy-lysosome-related and ferroptosis-associated proteins in the brain and significantly reduced ThS-positive amyloid plaque burden. Collectively, these findings demonstrate that FX enhances autophagy-lysosome-associated proteostatic regulation through AMPK/mTOR signaling and attenuates ferroptosis-linked oxidative injury under amyloidogenic conditions. These results provide mechanistic evidence supporting the role of FX as a marine-derived bioactive compound for modulating AD-related pathological processes.

Thapsigargin-induced autophagic flux impairment and inflammation are potentiated by CLN3 deficiency and alleviated by 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) in human ARPE-19 cells.

Fluoride exposure impairs lysosomal biogenesis and autophagic flux in myocardial injury: Involvement of the FNIP1/mTOR/TFEB pathway.

Acute pancreatitis is caused by a premature activation of digestive proteases. One hypothesis is based on the proteolytic activation of the serine protease trypsinogen by the lysosomal enzyme cathepsin B (CTSB) after co-localization in the same subcellular compartment. The ER-cargo receptor protein CLN8 (ceroid lipofuscinosis, neuronal) mediates cathepsin transport from the endoplasmic reticulum (ER), the site of enzyme synthesis, to the trans-Golgi system, from which they are distributed to their final destinations. The aim of this study is to investigate the role of CLN8 in acute pancreatitis and intracellular cathepsin trafficking by using isolated pancreatic acinar cells, a CLN8-deficient (Cln8mnd/MsrJ) mouse model, and 266–6 mouse pancreatic acinar tumor cells in which the Cln8 gene was inactivated by CRISPR/Cas9. Loss of CLN8 mitigated the early phase of acute pancreatitis but did not prevent it completely. We still observed CTSB expression in the endo-lysosomal and secretory compartment albeit enzyme activation was decreased. At later disease stages pancreatic injury increased along with an upregulation of ER-phagy shown by an overexpression of LC3B and the ER-phagy receptor FAM134B as well as autophagolysosome formation and increased ER stress. In summary, our data show that acute pancreatitis still occurs despite disruption of the EGRESS (ER-to-Golgi relaying of enzymes of the lysosomal system) complex implicating alternative intracellular enzyme delivery routes. They also illustrate that ER-stress and ER-phagy aggravate severity at later course of pancreatitis.Supplementary InformationThe online version contains supplementary material available at 10.1186/s43556-026-00479-4.

CLN3 Batten disease, also known as Juvenile Neuronal Ceroid Lipofuscinosis, is a childhood-onset neurodegenerative disorder caused by mutations in the CLN3 gene, frequently accompanied by emotional and behavioral symptoms. Although these symptoms and a reduced quality of life have been previously reported in individuals with CLN3 disease and their parents, no published comparisons with siblings exist. Furthermore, age and sex differences require further study. In a nationally representative sample (n = 20, age 7-29 years), the present study applied the ASEBA Behavior Checklists, the Inventory of Life Quality to examine the prevalence of emotional and behavioral problems and quality of life in affected individuals and sibling controls. Additionally, we assessed parental quality of life using the Satisfaction With Life Scale. We found that individuals with CLN3 disease scored in the borderline clinical range for the ASEBA total problem scores (mean 63.1, SD 11.6), whereas siblings fell within the normal variation range (mean 46.4, SD 11.4). A severely reduced quality of life was reported for both affected individuals and their parents, while siblings scored within the normal range. No significant sex differences were found. Despite facing significant challenges, families also reported strengths in their offspring: grit, determination and a sense of humor in affected individuals, and empathy and dutifulness in sibling controls. This study underscores the significant emotional and behavioral challenges faced by individuals with CLN3 Batten disease, which affect the quality of life for them and their parents. These findings emphasize the need for targeted interventions for both the patients and their families.

Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by dopaminergic neuron loss and α-synuclein (α-syn) aggregation, often linked to lysosomal dysfunction. Cathepsin D (CTSD), a lysosomal hydrolase essential for α-syn clearance, becomes functionally impaired when its maturation is disrupted, exacerbating proteostatic stress. This study investigated whether ginsenoside Rg1(Rg1) restores CTSD maturation and lysosomal function to mitigate PD pathology. MPTP-induced zebrafish and mouse PD models, as well as MPP+-treated SH-SY5Y cells, animals and cells were treated with Rg1 at different concentrations. Motor behavior, dopaminergic neuron survival, α-syn clearance, CTSD maturation, lysosomal activity, endoplasmic reticulum (ER) stress, oxidative stress, autophagic flux, and apoptosis were systematically evaluated. Rg1 improved locomotor performance and preserved dopaminergic neurons, promoted α-syn clearance, and enhanced CTSD maturation in lysosomes. These effects coincided with reduced ER and oxidative stress, normalized autophagic flux, and decreased apoptosis. Rg1 functions as a natural lysosomal enhancer, restoring lysosome-ER homeostasis and counteracting multiple pathogenic pathways in PD. The findings reveal a CTSD-dependent regulatory axis in α-syn homeostasis and highlight Rg1 as a promising multi-target therapeutic candidate for PD.

Mechanism of liquiritin from Codonopsis pilosula-Glycyrrhiza uralensis extract in alleviating piglet weaning stress via binding to alpha-1-microglobulin/bikunin precursor by machine learning and molecular dynamics simulation.

This study aimed to investigate the effect of downhill running on mitophagic flux and autophagosome-lysosome fusion in rat soleus muscle. Sprague-Dawley rats were trained on a treadmill at a speed of 16 m·min -1 and a decline of -16° for 90 min, and the soleus muscle was sampled at 0 h, 12 h, 24 h, 48 h, and 72 h after exercise. Mitochondrial ultrastructural changes were observed by using a transmission electron microscope. Protein levels of Cathepsin D, Vacuolar H + -ATPase (V-ATPase), mitochondrial respiratory complex Ⅰ (NDUFB8), complex Ⅲ (UQCRC2), and microtubule-associated protein 1 light chain 3 (LC3) were determined by Western blot. Mitochondria co-localizations with LC3 and lysosomal-associated membrane protein 2 (LAMP2), syntaxin 17 (STX17) co-localizations with LC3, LAMP2, SNAP29, and vesicle-associated membrane protein 8 (VAMP8), as well as the SNAP29 co-localizations with VAMP8, were measured by immunofluorescence. To assess mitophagic flux in vivo , colchicine or saline was injected intraperitoneally 3 d before exercise, and the protein expression of mitochondrial LC3-Ⅱ was detected by Western blot. After downhill running, mitochondrial structure appeared to be abnormal and contained autophagosomes and autophagolysosomes. The expression levels of Cathepsin D, Vacuolar H + -ATPase, and mitochondrial LC3, as well as the co-localizations of STX17 with LC3 and SNAP29 were significantly higher, whereas the expression levels of mitochondrial NDUFB8, UQCRC2, and LAMP2, along with the co-localizations of STX17 with LAMP2 and VAMP8 were significantly lower than those in the control group. Specifically, mitochondrial LC3-Ⅱ flux was significantly lower following downhill running. A bout of downhill running may block mitophagic flux by impairing mitophagosome-lysosome fusion, which is accompanied by increased recruitment of STX17 and SNAP29 to autophagosome and reduced co-localizations of STX17 and SNAP29 with lysosomal VAMP8.

TFEB has a protective effect in cisplatin induced AKI through regulating exosome-MVBs pathway.

Lysosomal storage, mitochondrial pathology, and autophagy in knockout of tripeptidyl peptidase 1 in human neuroblastoma cells in vitro.

Per- and polyfluoroalkyl substances (PFAS) are persistent pollutants linked to breast cancer (BC), but their role in perineural invasion (PNI) of triple-negative breast cancer (TNBC) is unclear. Cathepsin D (CTSD), a lysosomal protease, is hypothesized to mediate PFAS-induced PNI, though systematic evidence is lacking. We integrated multi-omics data from TCGA-BRCA, METABRIC, and single-cell RNA-seq datasets. Analyses included differential gene expression, Mendelian randomization, consensus clustering, and machine learning for prognostic modeling. Single-cell analyses were performed using Seurat, Monocle2, and CellChat. GraphBan screened natural CTSD-binding compounds, with binding affinity evaluated by molecular docking and dynamics simulations. Experimental validation included immunohistochemistry, immunofluorescence, Transwell, and Western blot assays. We identified 5 PFAS-associated PNI-related genes (PPGs), with CTSD central to TNBC PNI. PPG-based molecular subtyping revealed a high-risk subgroup exhibiting enhanced epithelial-mesenchymal transition (EMT) activity, proliferation capacity, and significantly poorer overall survival. The PPG-based prognostic model effectively stratified patient outcomes and immunotherapy response. Mendelian randomization confirmed a causal link between genetically predicted CTSD levels and BC risk. Single-cell analysis showed CTSD specifically enriched in myeloid cells; CTSD⁺ myeloid cells displayed immunosuppressive signatures and therapy resistance. CTSD⁺ epithelial cells interacted with cancer-associated fibroblasts via FGF signaling and showed altered metabolism. GraphBan predicted and experiments confirmed Aurantio-obtusin as a high-affinity CTSD inhibitor. Molecular simulations demonstrated stable binding of both PFAS and Aurantio-obtusin to CTSD. Histologically, elevated CTSD expression co-localized with CD68⁺ macrophages in PNI-positive TNBC tissues, while Aurantio-obtusin suppressed CTSD expression and inhibited TNBC cell proliferation and migration. This study suggests that PFAS exposure is associated with PNI and malignant progression in TNBC, potentially involving dysregulation of CTSD. The robust PPG-based prognostic signature and the natural inhibitor Aurantio-obtusin offer novel biomarkers and a potential therapeutic strategy for mitigating PFAS-related cancer risks.

BackgroundJuvenile neuronal ceroid lipofuscinosis (JNCL, Batten Disease) is a childhood-onset, neurodegenerative, lysosomal storage disorder caused by mutations in the lysosomal gene CLN3. Progressive cognitive decline is characteristic clinical feature, and no definitive treatment is currently available. The neuronal function of CLN3 is unknown, and the pathomechanisms leading to cognitive impairment are poorly understood hindering the development of targeted therapies.MethodsWhole-cell patch clamp and high-density microelectrode array recordings were performed in acute brain slices from Cln3Δex7/8 mice to assess synaptic properties, intrinsic excitability, and network activity. High-resolution confocal imaging was used to quantify dendritic spine density. To explore pre- and postsynaptic roles of CLN3, adeno-associated viral (AAV) re-expression of CLN3 was combined with optogenetics, allowing assessment of CLN3 function in each compartment selectively.ResultsLoss of CLN3 caused defective synaptic vesicle release and reduced synaptic strength, reflecting impairments in both pre- and postsynaptic function in Cln3Δex7/8 mice. We also observed reduced network bursting and deficits in intrinsic neuronal excitability, indicating early functional disturbances independent of storage burden and neuronal loss. Further, we report non-redundant requirements for CLN3 at both pre- and postsynaptic sites to sustain function. Importantly, AAV9-mediated gene rescue at early disease stages corrected preexisting synaptic defects and restored function.ConclusionsOur findings demonstrate a critical requirement for CLN3 in maintaining synaptic function and show that targeted gene therapy can restore established functional deficits in Cln3-deficient mice. This suggests that the therapeutic window may extend to stages already characterized by functional impairments, raising hope that targeted interventions could not only slow disease progress but to also potentially restore neuronal function and thereby improve clinical outcome. Moreover, these early synaptic deficits provide sensitive and robust functional readouts that can support preclinical research.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12967-026-08304-w.

BackgroundCLN3 Batten disease is a severe pediatric neurodegenerative disorder caused by mutations in the CLN3 gene, most commonly a 1 kb deletion encompassing exons 7 and 8. CLN3 deficiency is associated with lysosomal dysfunction, impaired cellular clearance and disrupted metabolism. While neurons are particularly vulnerable in CLN3 Batten disease and have been the primary focus of research, glial cells are increasingly recognized as active contributors to disease pathology. Among them, astrocytes—the most abundant glial cell type in the brain—play critical roles in maintaining neuronal health and homeostasis. However, astrocytes remain understudied in CLN3 patient-derived models.MethodsWe present the first iPSC-derived astrocyte model from a skin biopsy of a CLN3 patient carrying the common 1 kb deletion. Cellular and molecular features of iPSC and astrocytes derived from both healthy controls and the CLN3 patient were characterized via qPCR, immunocytochemistry and targeted mass spectrometry. In addition, comprehensive omics-based profiling, through transcriptomic and label-free quantitative proteomics, was performed to uncover novel molecular mechanisms and generate hypotheses that can guide future mechanistic and functional studies.ResultsTranscriptomic and proteomic analyses during astrocyte differentiation revealed an upregulation of mitochondrial respiratory chain complexes I and IV—contrasting with the downregulation typically observed in CLN3-deficient neurons. We also identified a metabolic shift favoring the elongation of very-long-chain saturated fatty acids, accompanied by reduced lipid synthesis and enhanced fatty acid oxidation. These metabolic alterations were paralleled by an upregulation of proteins involved in oxidative stress responses, likely reflecting a compensatory adaptation to mitochondrial and lipid metabolic dysregulation. Furthermore, we observed significant changes in chromatin organization during astrocyte differentiation in CLN3 cells, suggesting epigenetic remodeling as a contributing factor to disease pathology.ConclusionOur findings prompt the hypothesis that mitochondrial dysfunction may precede lysosomal defects in CLN3-deficient astrocytes. Restoring mitochondrial health could improve brain metabolism, inflammation control, neurotransmitter regulation, and neuronal survival, highlighting mitochondria as promising therapeutic targets in CLN3 Batten disease.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12929-026-01253-y.

PSA testing is widely used for the early detection of prostate cancer (PCa), but its low specificity leads to overdiagnosis and unnecessary interventions. Proclarix, a novel blood test combining serum levels of prostate specific antigen (PSA), percentage of free PSA (%fPSA), Cathepsin D (CTSD) and Thrombospondin 1 (THBS1) with age into a risk score, aims to improve risk stratification by predicting clinically significant PCa (csPCa). This study evaluated its diagnostic performance in a Danish population using retrospective serum samples collected consecutively from patients with suspected PCa. Proclarix' ability to reduce biopsies and detection of clinically insignificant PCa (ciPCa, defined as Grade Group < 2) was assessed in men with a PSA 2-10 ng/ml and a prostate volume of ≥ 35ml (targeted population) compared with the percentage of free PSA (%fPSA) and the European Randomized Study of Screening for Prostate Cancer Risk Calculator (ERSPC-RC). The secondary analysis included the performance of Proclarix' and Proclarix density compared with the %fPSA and PSA density (PSA-D) in a broader population with a PSA 2-20 ng/ml regardless of both prostate volume and DRE (extended population). Proclarix score is considered negative when it's below the cutoff 10%. In the targeted population (n = 373), a negative Proclarix test significantly reduced the probability of csPCa from 27% (pretest) to 5% (posttest, 95%CI: 0-10%), (p < 0.028) outperforming %fPSA (posttest 14%, 95%CI: 4-24%) and ERSPC-RC (posttest 20%, 95%CI: 4-36%). For the diagnosis of csPCa, Proclarix had a significantly (p < 0.01) greater specificity of 22% (95%CI: 17-27%) at 97% sensitivity (95%CI: 94-100%) and 95% NPV (95%CI: 90-100%) than did %fPSA and the ERSPC-RC, with 14% (95%CI: 10-18%) and 7% (95%CI: 4-11%) specificity, respectively. In the extended population (n = 656), Proclarix density had significantly (p < 0.01) greater specificity (39%, 95%CI: 35-44%) than did PSA-D (32%, 95%CI: 27-36%) at an equal sensitivity of 90%. Proclarix reduces prostate biopsies and ciPCa detection while maintaining a low risk of missing csPCa.

Melatonin is a promising drug for improving bone mass in postmenopausal women. This study investigated the mechanism behind the regulation of melatonin on osteoblast differentiation. Mouse embryonic osteoblast precursor MC3T3-E1 cells were treated with melatonin or transfected with cathepsin D (CTSD) vectors. A cellular model was established by H2O2 treatment. Alkaline phosphatase (ALP) staining and Alizarin Red S staining were performed. An osteoporosis mouse model was established by ovariectomy (OVX) and treated with melatonin or transfected with a CTSD knockdown vector. Bone biomechanical testing was performed to assess bone strength. Histological bone damage was assessed, osteoclast differentiation was visualized using tartrate-resistant acid phosphatase staining, osteocalcin (OCN) and type I collagen α1 chain (COL1A1) expression was visualized using immunohistochemistry, and serum bone turnover markers procollagen type I N-propeptide (PINP) and C-terminal telopeptide of type I collagen (CTX-1) levels were measured using enzyme-linked immunosorbent assay. Receptor activator of NF-κB ligand (RANKL), osteoprotegerin (OPG), Wnt3a, and β-catenin protein levels were determined using western blotting. Melatonin treatment or CTSD overexpression promoted ALP activity and mineralization in MC3T3-E1 cells. Melatonin upregulated CTSD expression. Melatonin treatment enhanced bone strength, inhibited osteoclast differentiation, increased OCN and COL1A1 expression, elevated PINP levels, and reduced CTX-1 in OVX mice. Moreover, melatonin suppressed RANKL expression and promoted OPG, Wnt3a, and β-catenin expression. CTSD knockdown abolished the regulatory effects of melatonin on MC3T3-E1 cells and OVX mice. In conclusion, melatonin increases CTSD expression to promote osteoblast differentiation and regulate the RANKL/OPG/Wnt signaling pathway, thereby slowing down the progression of osteoporosis.

Lysosomes are crucial for the function of fetal vacuolated enterocytes in neonatal piglets, yet how they are regulated by miRNAs remains poorly defined. Therefore, this study aimed to elucidate how miRNAs govern lysosomal homeostasis in the developing intestine. Using a neonatal piglet model of lysosomal dysfunction induced by imipramine (IMI), we identified ssc-miR-214-3p as a key down-regulated miRNA implicated in lysosomal pathways. In IPEC-J2 enterocytes, the miR-214-3p mimic ameliorated IMI cytotoxicity by restoring cell viability and migration while suppressing apoptosis. Further analysis revealed that miR-214-3p directly reversed the lysosomal defects triggered by IMI treatment. Specifically, it alleviated lysosomal alkalinization and markedly restored acid phosphatase (ACP) activity, indicating a recovery of the acidic hydrolytic environment. This restoration was also accompanied by the preservation of lysosomal membrane integrity and a consequent reduction in the nuclear translocation of transcription factor EB (TFEB). Furthermore, cathepsin D (CTSD) was validated as a direct target of miR-214-3p by luciferase assay, and its overexpression reversed the protective effects of the mimic on lysosomal acidification and lysosome-associated membrane protein 1 (LAMP1) levels. Collectively, our findings reveal a novel miR-214-3p/CTSD axis that regulates lysosomal homeostasis during neonatal intestinal maturation, providing a potential therapeutic target for porcine intestinal disorders.

Avian pathogenic Escherichia coli virulence protein Hcp2a induces incomplete autophagy in chicken HD11 cells

Neuronal ceroid lipofuscinosis or Batten disease comprises a category of autosomal recessive neurodegenerative disorders that primarily affect children. Mutations in different genes lead to different forms of neuronal ceroid lipofuscinoses (CLN1–14). At present, there is no established therapy to cure most of the neuronal ceroid lipofuscinoses and the treatments are symptomatic. Enzyme replacement therapy, gene therapy, stem cell transplantation, and pharmacological chaperone therapy are being tested in different animal models and human patients. Peroxisome proliferator-activated receptor alpha (PPARα) is a member of the nuclear hormone receptor superfamily, which along with its transcription partner retinoid X receptor alpha (RXRα) regulates the expression of their target genes. This review highlights the potential role of PPARα and RXRα in the regulation of CLN genes. Here, using the MatInspector program of the Genomatix software, we performed promoter analyses of all CLN genes and observed that most of the CLN genes harbor one or more potential binding sites for PPAR and RXR in their promoter region. We further grouped them according to a binding prediction of the transcription factors to indicate high affinity binding of PPAR to CLN2, CLN3, CLN4, CLN5, CLN7, CLN10, CLN11, CLN12, and CLN14. On the other hand, we observed high affinity binding of RXR to CLN1, CLN3, CLN6, CLN7, CLN8, CLN10, and CLN13. Since PPARα and RXRα have been demonstrated to control the transcription of CLN2 gene, our current promoter analysis findings highlight a possible treatment strategy for neuronal ceroid lipofuscinoses using agonists of PPARα and RXRα.

CLN1 disease, caused by mutations in the PPT1 gene, is a fatal neurodegenerative lysosomal storage disorder. While central nervous system (CNS) pathology is well documented, the impact on peripheral tissues remains unclear. Having previously described severe spinal cord pathology, we investigated whether PPT1 deficiency also impacts the neuromuscular junction (NMJ) and skeletal muscle, and whether early systemic gene therapy can prevent these disease manifestations. NMJ morphology, terminal Schwann cell (tSC) coverage, and skeletal muscle structure were examined in symptomatic and end-stage Ppt1-/- mice. Neonatal mice received systemic AAV9-hCLN1 gene therapy via intravenous injection. Untreated Ppt1-/- mice exhibited pronounced NMJ pathology, including progressive tSC loss, apparently reduced innervation, and increased abnormal acetylcholine receptor clustering. In parallel, we observed skeletal muscle atrophy, with decreased myofiber diameter and reduced myonuclear content, despite preserved sciatic nerve morphology. Systemic AAV9-hCLN1 therapy partially prevented or ameliorated these phenotypes, preserving NMJ innervation and muscle fiber structure. These findings identify peripheral NMJ and muscle abnormalities as previously unrecognized features of CLN1 disease and provide proof-of-concept that early systemic gene therapy can mitigate these effects. Our results highlight the systemic nature of CLN1 pathology and support the need for treatments that address both CNS and peripheral targets for comprehensive disease modification.

The Juvenile form of Batten disease is a neurodegenerative disease with symptoms starting in the first decade and ending in death in the third decade of life.The gene defective in this form of Batten disease, CLN3, is conserved in eukaryotes, suggesting that the gene product serves a basic function in the cell, though the function is unknown. We have investigated the expression and regulation of the yeast homolog BTN1.Reanalysis of publicly available gene expression data suggests that transcription of BTN1 increases in response to oxidative stress, treatment with rapamycin or arsenate, amino acid starvation, and sporulation conditions. Similar to GCN4, there are upstream open reading frames (uORF) in front of BTN1, suggesting translational regulation. We developed reporter strains in which the HIS3 open reading frame replaced that of the BTN1 gene, with and without the uORFs.These reporters show that one or more of the uORFs decrease the expression of the HIS3 reporter.When expressed in the reporter strain using a high copy vector, GCN3, tRNA , and tRNA , increase expression, suggesting the involvement of the TORC1 pathway.BIT61 abuts BTN1 but is encoded on the opposite strand; 3' RACE analysis indicates that the mRNA of BIT61 overlaps with that of BTN1.BIT61 is involved in the TORC2 pathway, which interacts with the TORC1 pathway, suggesting a possible cis-acting mechanism of co-regulation.Lastly, we demonstrate that a yeast strain with a null mutation in BTN1 is sensitive to selective amino acid starvation, further supporting the association of BTN1 with TORC1.