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Although immunogenic cell death (ICD) induced by lysosomal membrane permeabilization (LMP) evidently enhance the effectiveness of antitumor immunity for triple-negative breast cancer (TNBC) with poor immunogenicity, their potential is increasingly restricted by the development of other death pathways and the repair of lysosomes by endoplasmic reticulum (ER) during LMP induction. Herein, a polydopamine nanocomposite with i-motif DNA modified and BNN6 loaded is prepared toward boosting LMP and immunotherapy of TNBC by synergy of spatially confined photoacoustic (PA) effects and nitric oxide. Combining the high-frequency pulsed laser (4000 kHz) with the intra-lysosomal assembly of nanocomposites produced spatially confined and significantly boosted PA effects (4.8-fold higher than the individually dispersed particles extracellular), suppressing damage to other cellular components and selectively reducing lysosomal integrity to 19.2 %. Simultaneously, the releasing of nitric oxide inhibited the repair of lysosomes by ER stress, causing exacerbated LMP. Consequently, efficient immune activation was achieved, including the abundant releasing of CRT/HMGB1 (5.93–6.8-fold), the increasing maturation of dendritic cells (3.41-fold), and the fostered recruitment of CD4+/CD8+ T cells (3.99–3.78-fold) in vivo. The study paves a new avenue for the rational design and synergy of confined energy conversion and responsive nanostructures to achieve the treatment of low immunogenicity tumors. A strategy of boosting lysosomal membrane permeabilization (LMP) and concomitantly preventing the repair was developed to address the immunotherapy challenge of triple-negative breast cancer. Spatially confined and significantly enhanced photoacoustic (PA) effects were achieved through DNA-guided pH-responsive assembly of polydopamine nanocomposites in lysosomes and application of a high-frequency pulsed laser. Efficient immunogenic cell death was guaranteed by selective and powerful damage of lysosomal membranes through the significant contrast of PA intensities for dispersed/assembled particles and nitric oxide release induced endoplasmic reticulum stress. The study paves a new avenue for the rational design and synergy of confined energy conversion and responsive nanostructures to achieve the treatment of low immunogenicity tumors. [Display omitted] [ABSTRACT FROM AUTHOR]

Fluoride is known to induce nephrotoxicity; however, the underlying mechanisms remain incompletely understood. Therefore, this study aims to explore the roles and mechanisms of lysosomal membrane permeabilization (LMP) and the GSDME/HMGB1 axis in fluoride-induced nephrotoxicity and the protective effects of rutin. Rutin, a naturally occurring flavonoid compound known for its antioxidative and anti-inflammatory properties, is primarily mediated by inhibiting oxidative stress and reducing proinflammatory markers. To that end, we established in vivo and in vitro models. In the in vivo study, rats were exposed to sodium fluoride (NaF) throughout pregnancy and up until 2 months after birth. In parallel, we employed in vitro models using HK-2 cells treated with NaF, n-acetyl-L-cysteine (NAC), or rutin. We assessed lysosomal permeability through immunofluorescence and analyzed relevant protein expression via western blotting. Our findings showed that NaF exposure increased ROS levels, resulting in enhanced LMP and increased cathepsin B (CTSB) and D (CTSD) expression. Furthermore, the exposure to NaF resulted in the upregulation of cleaved PARP1, cleaved caspase-3, GSDME-N, and HMGB1 expressions, indicating cell death and inflammation-induced renal damage. Rutin mitigates fluoride-induced nephrotoxicity by suppressing ROS-mediated LMP and the GSDME/HMGB1 axis, ultimately preventing fluoride-induced renal toxicity occurrence and development. In conclusion, our findings suggest that NaF induces renal damage through ROS-mediated activation of LMP and the GSDME/HMGB1 axis, leading to pyroptosis and inflammation. Rutin, a natural antioxidative and anti-inflammatory dietary supplement, offers a novel approach to prevent and treat fluoride-induced nephrotoxicity.

Abstract RNA interference (RNAi) is a gene-silencing mechanism triggered by the cytosolic entry of double-stranded RNAs (dsRNAs). Many animal cells internalize extracellular dsRNAs via endocytosis for RNAi induction. However, it is not clear how the endocytosed dsRNAs are translocated into the cytosol across the endo/lysosomal membrane. Herein, we show that in Drosophila S2 cells, endocytosed dsRNAs induce lysosomal membrane permeabilization (LMP) that allows cytosolic dsRNA translocation. LMP mediated by dsRNAs requires the lysosomal Cl−/H+ antiporter ClC-b/DmOstm1. In clc-b or dmostm1 knockout S2 cells, extracellular dsRNAs are endocytosed and reach the lysosomes normally but fail to enter the cytosol. Pharmacological induction of LMP restores extracellular dsRNA-directed RNAi in clc-b or dmostm1-knockout cells. Furthermore, clc-b or dmostm1 mutant flies are defective in extracellular dsRNA-directed RNAi and its associated antiviral immunity. Therefore, endocytosed dsRNAs have an intrinsic ability to induce ClC-b/DmOstm1-dependent LMP that allows cytosolic dsRNA translocation for RNAi responses in Drosophila cells.

Abstract Purpose Radiotherapy-activated NBTXR3 (NBTXR3 + RT) has demonstrated superior efficacy in cancer cell destruction and tumor growth control, compared to radiotherapy (RT), in preclinical and clinical settings. Previous studies highlighted the immunomodulatory properties of NBTXR3 + RT, such as modification of tumor cell immunogenicity/adjuvanticity, producing an effective local tumor control and abscopal effect, related to an enhanced antitumor immune response. Furthermore, NBTXR3 + RT has shown potential in restoring anti-PD1 efficacy in a refractory tumor model. However, the early events leading to these results, such as NBTXR3 endocytosis, intracellular trafficking and primary biological responses induced by NBTXR3 + RT remain poorly understood. Methods We analyzed by transmission electron microscopy endocytosis and intracellular localization of NBTXR3 nanoparticles after endocytosis in various cell lines, in vitro and in vivo. A kinetic of NBTXR3 endocytosis and its impact on lysosomes was conducted using LysoTracker staining, and a RNAseq analysis was performed. We investigated the ability of NBTXR3 + RT to induce lysosomal membrane permeabilization (LMP) and ferroptosis by analyzing lipid peroxidation. Additionally, we evaluated the recapture by cancer cells of NBTXR3 released from dead cells. Results NBTXR3 nanoparticles were rapidly internalized by cells mainly through macropinocytosis and in a less extend by clathrin-dependent endocytosis. NBTXR3-containing endosomes were then fused with lysosomes. The day following NBTXR3 addition, we measured a significant increase in LysoTracker lysosome labeling intensity, in vitro as in vivo. Following RT, a significant lysosomal membrane permeabilization (LMP) was measured exclusively in cells treated with NBTXR3 + RT, while RT had no effect. The day post-irradiation, a significant increase in lipid peroxidation, a biomarker of ferroptosis, was measured with NBTXR3 + RT compared to RT. Moreover, we demonstrated that NBTXR3 nanoparticles released from dead cells can be recaptured by cancer cells. Conclusions Our findings provide novel insights into the early and specific biological effects induced by NBTXR3 + RT, especially LMP, not induced by RT in our models. The subsequent significant increase in lipid peroxidation partially explains the enhanced cancer cell killing capacity of NBTXR3 + RT compared to RT, potentially by promoting ferroptosis. This study improves our understanding of the cellular mechanisms underlying NBTXR3 + RT and highlights its potential as an agnostic therapeutic strategy for solid cancers treatment. Graphical Abstract

Bleomycin exhibits effective chemotherapeutic activity against multiple types of tumors, and also induces various side effects, such as pulmonary fibrosis and neuronal defects, which limit the clinical application of this drug. Macroautophagy/autophagy has been recently reported to be involved in the functions of bleomycin, and yet the mechanisms of their crosstalk remain insufficiently understood. Here, we demonstrated that reactive oxygen species (ROS) produced during bleomycin activation hampered autophagy flux by inducing lysosomal membrane permeabilization (LMP) and obstructing lysosomal degradation. Exhaustion of ROS with N-acetylcysteine relieved LMP and autophagy defects. Notably, we observed that LMP and autophagy blockage preceded the emergence of cellular senescence during bleomycin treatment. In addition, promoting or inhibiting autophagy-lysosome degradation alleviated or exacerbated the phenotypes of senescence, respectively. This suggests the alternation of autophagy activity is more a regulatory mechanism than a consequence of bleomycin-induced cellular senescence. Taken together, we reveal a specific role of bleomycin-induced ROS in mediating defects of autophagic degradation and further regulating cellular senescence in vitro and in vivo . Our findings, conversely, indicate the autophagy-lysosome degradation pathway as a target for modulating the functions of bleomycin. These provide a new perspective for optimizing bleomycin as a clinically applicable chemotherapeutics devoid of severe side-effects. Abbreviations : AT2 cells: type II alveolar epithelial cells; ATG7: autophagy related 7; bEnd.3: mouse brain microvascular endothelial cells; BNIP3L: BCL2/adenovirus E1B interacting protein 3-like; CCL2: C-C motif chemokine ligand 2; CDKN1A: cyclin dependent kinase inhibitor 1A; CDKN2A: cyclin dependent kinase inhibitor 2A; FTH1: ferritin heavy polypeptide 1; γ-H2AX: phosphorylated H2A.X variant histone; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HUVEC: human umbilical vein endothelial cells; HT22: hippocampal neuronal cell lines; Il: interleukin; LAMP: lysosomal-associated membrane protein; LMP: lysosome membrane permeabilization; MTORC1: mechanistic target of rapamycin kinase complex 1; NAC: N-acetylcysteine; NCOA4: nuclear receptor coactivator 4; PI3K: phosphoinositide 3-kinase; ROS: reactive oxygen species; RPS6KB/S6K: ribosomal protein S6 kinase; SA-GLB1/β-gal: senescence-associated galactosidase, beta 1; SAHF: senescence-associated heterochromatic foci; SASP: senescence-associated secretory phenotype; SEC62: SEC62 homolog, preprotein translocation; SEP: superecliptic pHluorin; SQSTM1/p62: sequestosome 1; TFEB: transcription factor EB.

Background and Purpose: Autophagy is a protective factor for controlling neuronal damage, while necroptosis promotes neuroinflammation after spinal cord injury (SCI). DADLE (D-Ala 2 , D-Leu 5 ]-enkephalin) is a selective agonist for delta (δ) opioid receptor and has been identified as a promising drug for neuroprotection. The aim of this study was to investigate the mechanism/s by which DADLE causes locomotor recovery following SCI., Experimental Approach: Spinal cord contusion model was used and DADLE was given by i.p. (16 mg·kg -1 ) in mice for following experiments. Motor function was assessed by footprint and Basso mouse scale (BMS) score analysis. Western blotting used to evaluate related protein expression. Immunofluorescence showed the protein expression in each cell and its distribution. Network pharmacology analysis was used to find the related signalling pathways., Key Results: DADLE promoted functional recovery after SCI. In SCI model of mice, DADLE significantly increased autophagic flux and inhibited necroptosis. Concurrently, DADLE restored autophagic flux by decreasing lysosomal membrane permeabilization (LMP). Additionally, chloroquine administration reversed the protective effect of DADLE to inhibit necroptosis. Further analysis showed that DADLE decreased phosphorylated cPLA 2 , overexpression of cPLA 2 partially reversed DADLE inhibitory effect on LMP and necroptosis, as well as the promotion autophagy. Finally, AMPK/SIRT1/p38 pathway regulating cPLA 2 is involved in the action DADLE on SCI and naltrindole inhibited DADLE action on δ receptor and on AMPK signalling pathway., Conclusion and Implication: DADLE causes its neuroprotective effects on SCI by promoting autophagic flux and inhibiting necroptosis by decreasing LMP via activating δ receptor/AMPK/SIRT1/p38/cPLA 2 pathway., (© 2023 British Pharmacological Society.)

The synthesis of carborane-1,8-naphthalimide conjugates and evaluation of their DNA-binding ability and anticancer activity were performed. A series of 4-carboranyl-3-nitro-1,8-naphthalimide derivatives, mitonafide and pinafide analogs, were synthesised via amidation and reductive amination reactions, and their calf thymus DNA (ct-DNA)-binding properties were investigated using circular dichroism, UV–vis spectroscopy, and thermal denaturation. Results showed that conjugates 34–37 interacted very strongly with ct-DNA (ΔTm = 10.00–13.00 °C), indicating their ability to intercalate with DNA, but did not inhibit the activity of topoisomerase II. The conjugates inhibited the cell growth of the HepG2 cancer cell line in vitro. The same compounds caused the G2M phase arrest. Cell lines treated with these conjugates showed an increase in reactive oxygen species, glutathione, and Fe2+ levels, lipid peroxidation, and mitochondrial membrane potential relative to controls, indicating the involvement of ferroptosis. Furthermore, these conjugates caused lysosomal membrane permeabilization in HepG2 cells but not in MRC-5 cells.

PM2.5 exposure can be associated with the onset of neurodegenerative diseases, with oxidative stress-induced cellular homeostasis disruption and cell death as one of the main mechanisms. However, the exact cellular and molecular processes are still rarely investigated. Autophagy and KEAP1-NRF2 (Kelch-like ECH-Associating protein 1-nuclear factor erythroid 2 related factor 2) signaling pathway are two main cellular defense systems for maintaining cellular homeostasis and resisting oxidative stress. In this study, we primarily investigated the role of autophagy and KEAP1-NRF2 in regulating cell death resulting from PM2.5 exposure in mouse neuroblastoma N2a cells. Our results showed that PM2.5 exposure disrupted autophagic flux by impairing lysosomal function, including lysosomal alkalinization, increased lysosome membrane permeabilization (LMP), and Cathepsin B release. Furthermore, dysregulated autophagy enhances NRF2 activity in a p62-dependent manner, which then initiates the expression of a series of antioxidant genes and increases cellular insensitivity to ferroptosis. Meanwhile, autophagy dysfunction impairs the intracellular degradation of ferroptosis related proteins such as GPX4 and ferritin. As these proteins accumulate, cells also become less sensitive to ferroptosis. LMP-associated cell death may be the main mechanism of PM2.5-induced N2a cytotoxicity. Our results may provide insights into the mechanisms of PM2.5-induced neurotoxicity and predict effective prevention and treatment strategies.

Loss of lysosomal membrane integrity results in leakage of lysosomal hydrolases to the cytosol which might harm cell function and induce cell death. Destabilization of lysosomes often precede apoptotic or necrotic cell death and occur during both physiological and pathological conditions. The weak base acridine orange readily enters cells and accumulates in the acidic environment of lysosomes. Vital staining with acridine orange is a well-proven technique to observe lysosomal destabilization using fluorescence microscopy and flow cytometry. These analyses are, however, time consuming and only adapted for discrete time points, which make them unsuitable for large-scale approaches. Therefore, we have developed a time-saving, high-throughput microplate reader-based method to follow destabilization of the lysosomal membrane in real-time using acridine orange. This protocol can easily be adopted for patient samples since the number of cells per sample is low and the time for analysis is short.

Abstract Serine hydroxymethyltransferase 2 (SHMT2) plays an important role in converting serine to glycine and supplying carbon to one-carbon metabolism to sustain cancer cell proliferation. However, the expression, function, and underlying mechanisms of SHMT2 in clear cell renal cell carcinoma (ccRCC) remain largely unknown. In this study, we demonstrated that SHMT2 was upregulated in ccRCC tissues compared with controls and associated with patient survival. SHMT2 knockdown inhibited proliferation, migration, and invasion in ccRCC cells. Overexpression of SHMT2 promoted tumor progression. Mechanistically, SHMT2 depletion disrupted one-carbon metabolism, increased reactive oxygen species (ROS) levels, and decreased ATP levels via metabolic reprogramming, which destroyed cell homeostasis. The SHMT2 knockdown-induced stress activated autophagy. A mass of autophagosomes fused with lysosomes, resulting in lysosomal membrane permeabilization (LMP) and leakage of lysosomal contents into the cytoplasm, which eventually led to apoptosis. Our work reveals that SHMT2 functions as an oncogenic gene to promote ccRCC progression. SHMT2 depletion induces apoptosis by causing LMP through excessive activation of the autophagy-lysosome pathway via metabolic reprogramming.

Abstract Spinal cord injury (SCI) is a devastating injury that may result in permanent motor impairment. The active ingredients of medications are unable to reach the affected area due to the blood‒brain barrier. Elamipretide (SS-31) is a new and innovative aromatic cationic peptide. Because of its alternating aromatic and cationic groups, it freely crosses the blood‒brain barrier. It is also believed to decrease inflammation and protect against a variety of neurological illnesses. This study explored the therapeutic value of SS-31 in functional recovery after SCI and its possible underlying mechanism. A spinal cord contusion injury model as well as the Basso Mouse Scale, footprint assessment, and inclined plane test were employed to assess how well individuals could function following SCI. The area of glial scarring, the number of dendrites, and the number of synapses after SCI were confirmed by HE, Masson, MAP2, and Syn staining. Western blotting, immunofluorescence, and enzyme-linked immunosorbent assays were employed to examine the expression levels of pyroptosis-, autophagy-, lysosomal membrane permeabilization (LMP)- and MAPK signalling-related proteins. The outcomes showed that SS-31 inhibited pyroptosis, enhanced autophagy and attenuated LMP in SCI. Mechanistically, we applied AAV vectors to upregulate Pla2g4A in vivo and found that SS-31 enhanced autophagy and attenuated pyroptosis and LMP by inhibiting phosphorylation of cPLA2. Ultimately, we applied asiatic acid (a p38-MAPK agonist) to test whether SS-31 regulated cPLA2 partially through the MAPK-P38 signalling pathway. Our group is the first to suggest that SS-31 promotes functional recovery partially by inhibiting cPLA2-mediated autophagy impairment and preventing LMP and pyroptosis after SCI, which may have potential clinical application value.

Abstract Lysosomes are single-membraned organelles that mediate the intracellular degradation of macromolecules. Various stress can induce lysosomal membrane permeabilization (LMP), translocating intralysosomal components, such as cathepsins, to the cytoplasm, which induces lysosomal-dependent cell death (LDCD). This study reports that p53 regulates LMP in response to DNA-damaging drugs. Treating wild-type TP53 A549 cells with DNA-damaging drugs (namely, doxorubicin, carboplatin, and etoposide) induced LMP and accelerated cell death more rapidly than treating TP53-knockout (KO) A549 cells. This suggested p53-dependent LMP and LDCD induction in response to DNA damage. LMP was induced by p53-dependent BID upregulation and activation, followed by translocation of truncated BID to lysosomes. Simultaneously, autophagy for damaged lysosome elimination (lysophagy) was activated via the p53–mTOR–TEFB/TFE3 pathways in response to DNA damage. These data suggested the dichotomous nature of p53 for LMP regulation; LMP induction and repression via the p53–BID axis and p53–mTOR–TFEB/TFE3 pathway, respectively. Blocking autophagy with hydroxychloroquine or azithromycin as well as ATG5 KO enhanced LMP and LDCD induction after exposure to DNA-damaging drugs. Furthermore, lysosomal membrane stabilization using U18666A, a cholesterol transporter Niemann-Pick disease C1 (NPC1) inhibitor, suppressed LMP as well as LDCD in wild-type TP53, but not in TP53-KO, A549 cells. Thus, LMP is finely regulated by TP53 after exposure to DNA-damaging drugs.

Background and Purpose: Autophagy is a protective factor for controlling neuronal damage, while necroptosis promotes neuroinflammation after spinal cord injury (SCI). DADLE (D‐Ala2, D‐Leu5]‐enkephalin) is a selective agonist for delta (δ) opioid receptor and has been identified as a promising drug for neuroprotection. The aim of this study was to investigate the mechanism/s by which DADLE causes locomotor recovery following SCI. Experimental approach: Spinal cord contusion model was used and DADLE was given by i.p. (16 mg·kg−1) in mice for following experiments. Motor function was assessed by footprint and Basso mouse scale (BMS) score analysis. Western blotting used to evaluate related protein expression. Immunofluorescence showed the protein expression in each cell and its distribution. Network pharmacology analysis was used to find the related signalling pathways. Key Results: DADLE promoted functional recovery after SCI. In SCI model of mice, DADLE significantly increased autophagic flux and inhibited necroptosis. Concurrently, DADLE restored autophagic flux by decreasing lysosomal membrane permeabilization (LMP). Additionally, chloroquine administration reversed the protective effect of DADLE to inhibit necroptosis. Further analysis showed that DADLE decreased phosphorylated cPLA2, overexpression of cPLA2 partially reversed DADLE inhibitory effect on LMP and necroptosis, as well as the promotion autophagy. Finally, AMPK/SIRT1/p38 pathway regulating cPLA2 is involved in the action DADLE on SCI and naltrindole inhibited DADLE action on δ receptor and on AMPK signalling pathway. Conclusion and Implication: DADLE causes its neuroprotective effects on SCI by promoting autophagic flux and inhibiting necroptosis by decreasing LMP via activating δ receptor/AMPK/SIRT1/p38/cPLA2 pathway. [ABSTRACT FROM AUTHOR]

Lysosomal membrane permeabilization (LMP) has emerged as a significant component of cellular signaling pathway by which autophagy or cell death is regulated under many pathological situations including alcohol-associated liver disease (ALD). However, the mechanisms involved in the regulation of LMP in ALD remain obscure. Recently, we demonstrated that lipotoxicity serves as a causal factor to trigger LMP in hepatocytes. We identified that the apoptotic protein BAX (BCL2 associated X, apoptosis regulator) could recruit MLKL (mixed lineage kinase domain-like pseudokinase), a necroptotic executive protein, to lysosomes and induce LMP in various ALD models. Importantly, the pharmacological or genetic inhibition of BAX or MLKL protects hepatocytes from lipotoxicity-induced LMP. Thus, our study reveals a novel molecular mechanism that activation of BAX/MLKL signaling contributes to the pathogenesis of ALD through mediating lipotoxicity-induced LMP. Abbreviations: ALD: alcohol-associated liver disease; BAX: BCL2 associated X; LAMP2: lysosomal associated membrane protein 2; LMP: lysosomal membrane permeabilization; MLKL: mixed lineage kinase domain-like pseudokinase; PA: palmitic acid.

Lysosomes are acidic organelles that mediate the degradation and recycling of cellular waste materials. Damage to lysosomes can cause lysosomal membrane permeabilization (LMP) and trigger different types of cell death, including apoptosis. Newcastle disease virus (NDV) can naturally infect most birds. Additionally, it serves as a promising oncolytic virus known for its effective infection of tumor cells and induction of intensive apoptotic responses. However, the involvement of lysosomes in NDV-induced apoptosis remains poorly understood. Here, we demonstrate that NDV infection profoundly triggers LMP, leading to the translocation of cathepsin B and D and subsequent mitochondria-dependent apoptosis in various tumor and avian cells. Notably, the released cathepsin B and D exacerbate NDV-induced LMP by inducing the generation of reactive oxygen species. Additionally, we uncover that the viral Hemagglutinin neuraminidase (HN) protein induces the deglycosylation and degradation of lysosome-associated membrane protein 1 (LAMP1) and LAMP2 dependent on its sialidase activity, which finally contributes to NDV-induced LMP and cellular apoptosis. Overall, our findings elucidate the role of LMP in NDV-induced cell apoptosis and provide novel insights into the function of HN during NDV-induced LMP, which provide innovative approaches for the development of NDV-based oncolytic agents. Author summary: Our study investigates the role of lysosomes in Newcastle disease virus (NDV)-induced cell apoptosis. We found that NDV infection leads to lysosomal membrane permeabilization (LMP) and the release of cathepsin B and D enzymes. This triggers mitochondria-dependent apoptosis in various tumor and avian cells. The released cathepsin B and D exacerbate LMP by generating reactive oxygen species. Additionally, the viral Hemagglutinin neuraminidase protein degrades lysosome-associated membrane proteins (LAMP1 and LAMP2), contributing to LMP and apoptosis. These findings provide insights into NDV-induced cell apoptosis and highlight the potential of targeting lysosomes for the development of NDV-based oncolytic therapies. [ABSTRACT FROM AUTHOR]

For a long time, lysosomes were purely seen as organelles in charge of garbage disposal within the cell. They destroy any cargo delivered into their lumen with a plethora of highly potent hydrolytic enzymes, including various proteases. In case of damage to their limiting membranes, the lysosomes release their soluble content with detrimental outcomes for the cell. In recent years, however, this view of the lysosome changed towards acknowledging it as a platform for integration of manifold intracellular and extracellular signals. Even impaired lysosomal membrane integrity is no longer considered to be a one‐way street to cell death. Increasing evidence suggests that lysosomal enzymes, mainly cathepsin proteases, can be released in a spatially and temporarily restricted manner that is compatible with cellular survival. This way, cathepsins can act in the cytosol and the nucleus, where they affect important cellular processes such as cell division. Here, we review this evidence and discuss the routes and molecular mechanisms by which the cathepsins may reach their unusual destination.

The synthesis of carborane-1,8-naphthalimide conjugates and evaluation of their DNA-binding ability and anticancer activity were performed. A series of 4-carboranyl-3-nitro-1,8-naphthalimide derivatives, mitonafide and pinafide analogs, were synthesised via amidation and reductive amination reactions, and their calf thymus DNA (ct-DNA)-binding properties were investigated using circular dichroism, UV-vis spectroscopy, and thermal denaturation. Results showed that conjugates 34 - 37 interacted very strongly with ct-DNA (Δ T m = 10.00-13.00 °C), indicating their ability to intercalate with DNA, but did not inhibit the activity of topoisomerase II. The conjugates inhibited the cell growth of the HepG2 cancer cell line in vitro levels, lipid peroxidation, and mitochondrial membrane potential relative to controls, indicating the involvement of ferroptosis. Furthermore, these conjugates caused lysosomal membrane permeabilization in HepG2 cells but not in MRC-5 cells.2+ levels, lipid peroxidation, and mitochondrial membrane potential relative to controls, indicating the involvement of ferroptosis. Furthermore, these conjugates caused lysosomal membrane permeabilization in HepG2 cells but not in MRC-5 cells.

Sodium sulfite is a widely used preservative in the food industry. Ferroptosis has been a newly discovered form of iron-dependent oxidative cell death in recent years. However, the potential connection between sodium sulfite and ferroptosis has not been explored. In our study, we observed the abnormal expression of ferroptosis marker protein in vivo , suggesting that sodium sulfite caused ferroptosis in vivo . Next, our study revealed that sodium sulfite caused the overproduction of mitochondrial reactive oxygen species (mtROS) in the AML-12 cells. It is well established that reactive oxygen species (ROS) can induce lysosomal membrane permeabilization. After lysosomal membrane permeabilization occurs, the outflow of Fe 2+ in lysosomes triggers the Fenton reaction and subsequently results in the increase of intracellular ROS level, which is closely related to ferroptosis. As speculated, acridine orange (AO) staining and LysoTracker red staining showed that sodium sulfite-induced lysosomal membrane permeabilization could be alleviated by mtROS scavenger TEMPO. In addition, TEMPO, lysosomal stabilizer mannose, and lysosomal iron chelator deferoxamine (DFO) inhibited sodium sulfite-induced ferroptosis. Overall, the results showed that sodium sulfite induced lysosomal iron efflux through the mtROS-lysosomal membrane permeabilization pathway and eventually led to ferroptosis. Our study might provide a new mechanism for the hepatotoxicity of sodium sulfite and a theoretical basis for the risk assessment of sodium sulfite as a food additive.

Cancer is a global health and economic issue. The majority of anticancer therapies become ineffective due to frequent genomic turnover and chemoresistance. Furthermore, chemotherapy and radiation are non-specific, killing all rapidly dividing cells including healthy cells. In this review, we examine the ability of some natural products to induce lysosomal-mediated cell death in neoplastic cells as a way to kill them more specifically than conventional therapies. This list is by no means exhaustive. We postulate mechanisms to explain lysosomal membrane permeabilization and its role in triggering cell death in cancer cells.

Titanium dioxide nanoparticles (TiO2 NPs) have wide commercial applications, owing to their small size; however, the biosafety of TiO2 NPs should be evaluated further. In this study, we aimed to investigate the cytotoxicity of TiO2 NPs in the presence and absence of ultraviolet A (UVA) irradiation in human keratinocyte HaCaT cells. TiO2 NPs did not significantly affect cell viability in the absence of UVA irradiation. Nonetheless, UVA-irradiated TiO2 NPs induced caspase-dependent apoptosis of HaCaT cells. Exposure of HaCaT cells to TiO2 NPs and UVA resulted in reactive oxygen species (ROS) generation and lysosomal membrane permeabilization (LMP); both effects were not observed in the absence of UVA irradiation. An analysis of the relationship between LMP and ROS, using CA-074 as a cathepsin inhibitor or NAC as an antioxidant, showed that LMP stimulates ROS generation under these conditions. These results imply that LMP-dependent oxidative stress plays a critical role in the UVA phototoxicity of TiO2 NPs in HaCaT cells.

Purpose: Emerging evidence suggests that 27-Hydroxycholesterol (27-OHC) causes neurodegenerative diseases through the induction of cytotoxicity and cholesterol metabolism disorder. The objective of this study is to determine the impacts of 27-OHC on lysosomal membrane permeabilization (LMP) and pyroptosis in neurons in the development of neural degenerative diseases.Methods: In this study, SH-SY5Y cells and C6 cells were co-cultured in vitro to investigate the influence of 27-OHC on the function of lysosome, LMP and pyroptosis related factors in neuron. Lyso Tracker Red (LTR) was used to detect the changes of lysosome pH, volume and number. Acridine orange (AO) staining was also used to detect the LMP in neurons. Then the morphological changes of cells were observed by a scanning electron microscope (SEM). The content of lysosome function associated proteins [including Cathepsin B (CTSB), Cathepsin D (CTSD), lysosomal-associated membraneprotein-1 (LAMP-1), LAMP-2] and the pyroptosis associated proteins [including nod-like recepto P3 (NLRP3), gasdermin D (GSDMD), caspase-1 and interleukin (IL)-1β] were detected through Western blot.Results: Results showed higher levels of lysosome function associated proteins, such as CTSB (p < 0.05), CTSD (p < 0.05), LAMP-1 (p < 0.01), LAMP-2; p < 0.01) in 27-OHC treated group than that in the control group. AO staining and LTR staining showed that 27-OHC induced lysosome dysfunction with LMP. Content of pyroptosis related factor proteins, such as GSDMD (p < 0.01), NLRP3 (p < 0.001), caspase-1 (p < 0.01) and IL-1β (p < 0.01) were increased in 27-OHC treated neurons. Additionally, CTSB was leaked through LMP into the cytosol and induced pyroptosis. Results from the present study also suggested that the CTSB is involved in activation of pyroptosis.Conclusion: Our data indicate that 27-OHC contributes to the pathogenesis of cell death by inducing LMP and pyroptosis in neurons.