Endoplasmic Reticulum Research Articles

Advancements in autophagy perturbations in Alzheimer's disease: Molecular aspects and therapeutics.

Advancements in autophagy perturbations in Alzheimer's disease: Molecular aspects and therapeutics.

SEL1L-mediated endoplasmic reticulum associated degradation inhibition suppresses proliferation and migration in Huh7 hepatocellular carcinoma cells

BACKGROUND Proteins play a central role in regulating biological functions, and various pathways regulate their synthesis and secretion. Endoplasmic reticulum-associated protein degradation (ERAD) is crucial for monitoring protein synthesis and processing unfolded or misfolded proteins in actively growing tumor cells. However, the role of the multiple ERAD complexes in liver cancer remains unclear. AIM To elucidate the effects of SEL1L -mediated ERAD on Huh7 and explore the underlying mechanisms in vivo and in vitro . METHODS Huh7 cells were treated with ERAD inhibitor to identify ERAD’s role. Cell counting kit-8, 5-ethynyl-2’-deoxyuridine and colony formation experiments were performed. Apoptosis level and migration ability were assessed using fluorescence activated cell sorting and Transwell assay, respectively. Huh7 SEL1L knockout cell line was established via clustered regularly interspaced short palindromic repeats, proliferation, apoptosis, and migration were assessed through previous experiments. The role of SEL1L in vivo and the downstream target of SEL1L were identified using Xenograft and mass spectrometry, respectively. RESULTS The ERAD inhibitor suppressed cell proliferation and migration and promoted apoptosis. SEL1L -HRD1 significantly influenced Huh7 cell growth. SEL1L knockout suppressed tumor cell proliferation and migration and enhanced apoptosis. Mass spectrometry revealed EXT2 is a primary substrate of ERAD. SEL1L knockout significantly increased the protein expression of EXT2 . Furthermore, EXT2 knockdown partially restored the effect of SEL1L knockout. CONCLUSION ERAD inhibition suppressed the proliferation and migration of Huh7 and promoted its apoptosis. EXT2 plays an important role and ERAD might be a potential treatment for Huh7 hepatocellular carcinoma.

Isoguanosine-Induced ER Stress via AMPK Enhances Chemosensitivity in OSCC.

Oral squamous cell carcinoma (OSCC) is the most common malignancy of the head and neck; however, the efficacy of existing treatment is limited and new effective strategies need to be explored. Our previous work demonstrates that isoguanosine (isoG) is a promising nucleoside molecule with superior self-assembly capability and significant anti-OSCC potential. However, the antitumor mechanism of isoG remains unclear. In this study, we reveal that the antiproliferative effect of isoG is mediated by its cellular metabolite, isoguanosine 5'-monophosphate (isoGMP), which induces excessive endoplasmic reticulum (ER) stress and cell death through adenosine monophosphate-activated protein kinase (AMPK) activation. IsoG activates AMPK and induces ER stress at low concentrations, with minimal impact on cell viability at these concentrations. To further explore the therapeutic potential of isoG, we investigated its role in modulating chemosensitivity. Our findings show that AMPK activation enhances the sensitivity of OSCC cells to 5-fluorouracil (5-FU), and the combination of isoG and 5-FU exhibits a synergistic anticancer effect. Building on the self-assembly characteristics of isoG, we developed an innovative treatment platform by introducing dynamic borate ester bonds to form an isoguanosine-phenylenediboronic acid-isoguanosine (isoGPBisoG) structure. When combined with 5-FU, this platform achieved remarkable therapeutic efficacy in 2 OSCC cell-derived xenograft models, with tumor inhibition rates of 71.0% and 56.6%, respectively, compared with control. These findings establish isoG as a potent enhancer of chemotherapeutic efficacy in OSCC via AMPK activation. More importantly, the isoGPBisoG and 5-FU combination represents a significant paradigm of a synergistic therapy platform. This novel approach offers a promising direction for the development of more effective OSCC treatments.

GZ17-6.02 interacts with carboplatin and etoposide to kill neuroblastoma cells.

The biology of GZ17-6.02 alone and more so in combination with either of the standard-of-care agents etoposide or carboplatin killed MYCN overexpressing neuroblastoma (NB) cells is unknown. The methods involved in this study are in-cell immunoblotting, trypan blue exclusion, plasmid and siRNA transfection, assessment of autophagy using a plasmid expressing LC3-GFP-RFP. GZ17-6.02 (602) comprises, by mass, a ratio of curcumin (1.0), harmine (1.3), and isovanillin (7.7). In tumors dosed with 602, the ratio becomes curcumin (1.0), harmine (16), and isovanillin (6.1) (602NR). GZ17-6.02 activated ATM, AMPK, ULK1, ATG13, and PERK and inactivated ERBB1, ERBB2, ERBB3, ERBB4, AKT, mTORC1, mTORC2, SRC, NFκB, YAP, and eIF2α. 602 enhanced autophagosome formation and autophagic flux that was amplified when it was combined with etoposide or carboplatin. Compared with 602, 602NR caused significantly greater autophagosome formation that was also amplified when in combination with chemotherapy and which was reduced ~40% by knockdown of ATM or AMPKα and abolished by knockdown of Beclin1 or ATG5. Knockdown of ATM or AMPKα significantly reduced tumor cell death caused by 602 of 602NR, whereas endoplasmic reticulum stress (eIF2α) and macroautophagy (Beclin1, ATG5) were more effective at maintaining tumor cell survival. Combined knockdown of Beclin1 and the death receptor CD95 almost abolished the antitumor actions of 602 and 602NR. 602, and more so 602NR, kills MYCN NB cells and interacts with standard-of-care chemotherapeutics to cause further killing via autophagy and death receptor signaling.

Genome Mining for Hub Genes Related to Endoplasmic Reticulum Stress in Pancreatitis: A Perspective from In Silico Characterization

Genome Mining for Hub Genes Related to Endoplasmic Reticulum Stress in Pancreatitis: A Perspective from In Silico Characterization

Anti-inflammatory effects of Esomeprazole in septic lung injury by mediating endoplasmic reticulum stress

Anti-inflammatory effects of Esomeprazole in septic lung injury by mediating endoplasmic reticulum stress

Induction of MASH-like pathogenesis in the Nwd1−/− mouse liver

Endoplasmic reticulum (ER) stores Ca2+ and plays crucial roles in protein folding, lipid transfer, and it’s perturbations trigger an ER stress. In the liver, chronic ER stress is involved in the pathogenesis of metabolic dysfunction-associated steatotic liver disease (MASLD) and metabolic dysfunction-associated steatohepatitis (MASH). Dysfunction of sarco/endoplasmic reticulum calcium ATPase (SERCA2), a key regulator of Ca2+ transport from the cytosol to ER, is associated with the induction of ER stress and lipid droplet formation. We previously identified NACHT and WD repeat domain-containing protein 1 (Nwd1) localized at the ER and mitochondria. However, the physiological significance of Nwd1 outside the brain remains unclear. In this study, we revealed that Nwd1−/− mice exhibited pathological manifestations comparable to MASH. Nwd1 interacts with SERCA2 near ER membranes. Nwd1−/− livers exhibited reduced SERCA2 ATPase activity and a smaller Ca2+ pool in the ER, leading to an exacerbated state of ER stress. These findings highlight the importance of SERCA2 activity mediated by Nwd1 in the pathogenesis of MASH.

Endoplasmic Reticulum-Dependent Apoptotic Response to Cellular Stress in Patients with Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic, common autoimmune disease. It is characterized by inflammatory polyarthritis, which can lead to permanent disability in patients. Current treatment is mainly symptom-related, aiming to reduce pain and inflammation, but does not lead to a full recovery. This treatment includes non-steroidal anti-inflammatory drugs (NSAIDs) and disease-modifying anti-rheumatic drugs (DMARDs). It has been shown that, due to chronic inflammation, reduced glucose levels and hypoxia, endoplasmic reticulum (ER) stress is induced in RA patients, leading to the activation of multiple signaling pathways, including the ER-dependent adaptation of the unfolded protein response (UPR) pathway. The aim of this study was to assess the level of apoptosis in patients diagnosed with RA. The study sought to investigate whether UPR response correlated with apoptosis induction could serve as a potential diagnostic marker or therapeutic target. In vitro studies have shown that UPR pathway activity can be observed in patients diagnosed with RA. The study group consisted of PBMC cells from 61 individuals, including a total of 31 rheumatoid arthritis patients and 30 healthy controls. In order to validate UPR activation, we estimated molecular markers of ER stress via RT-qPCR expression analysis. GAPDH expression was used as a standard control. Elevated levels of mRNA for the eIF2α (p-value = 0.001903), the BBC3 (PUMA) (p-value = 0.007457 × 10−7) and the TP53 (p-value = 0.002212) were confirmed in a group of RA patients. Further analysis showed that after the induction of apoptosis the percentage of DNA contained in the tail was 37.78% higher in RA patients than in the control group (p-value = 0.0003) measured by comet assay. The exogenous damage caused by hydrogen peroxide was found to be statistically elevated in RA patients and the caspase-3 level was calculated of 40.17% higher than in controls (p-value = 0.0028). It was also found that PBMC cells from RA patients were more sensitive to apoptotic induction. Our results were confirmed by flow cytometry. The most important finding from our data was the confirmation of elevated sensitivity to apoptosis induction in RA patients; the results showed a 40.23% higher percentage of cells in early apoptosis than in the control group (p-value = 0.0105). Our results may help to assess the feasibility of the application of early diagnosis and targeted therapy in the treatment of RA patients, including the ER signaling pathway via selected UPR-dependent molecular inhibitors.

Targeted Lipidomics and Transcriptomics Unveil Aberrant Lipid Metabolic Remodeling in Visceral and Subcutaneous Adipose Tissue under ER Stress.

Endoplasmic reticulum (ER) stress is known to impair the function of visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT), disrupting lipid metabolism. Despite the crucial role lipid plays in regulating adipose tissue function, the specific lipidomic alterations in VAT and SAT under ER stress remain unclear. In this study, ER stress was induced in VAT and SAT, and targeted lipidomic and transcriptomic approaches were used to analyze lipid metabolism and gene expression profiles. The results revealed that VAT exhibited a stronger ER stress response, characterized by a significant increase in binding immunoglobulin protein (BiP) expression and notable lipidomic disruptions, especially in glycerides and sterols. These disruptions were marked by a decrease in protective polyunsaturated fatty acyl species and the accumulation of lipotoxic molecules. In contrast, SAT displayed less severe lipidomic alterations. Transcriptomic analysis indicated that VAT was more susceptible to immune activation, inflammation, and metabolic dysfunction, while SAT primarily showed alterations in protein folding processes. These findings underscore the tissue-specific mechanisms of ER stress adaptation in VAT and SAT. In conclusion, VAT appears to be a critical target for addressing metabolic dysfunction in obesity and related disorders, with potential therapeutic implications for managing ER stress-induced metabolic diseases.

Advances in Endoplasmic Reticulum Stress Research—Insights from the Special Issue “Endoplasmic Reticulum Stress and Apoptosis”

The endoplasmic reticulum (ER) is pivotal in maintaining the internal homeostasis of the cell [...]

Lysosomes’ fallback strategies: more than just survival or death

Lysosomes are heterogeneous, acidic organelles whose proper functionality is critically dependent on maintaining the integrity of their membranes and the acidity within their lumen. When subjected to stress, the lysosomal membrane can become permeabilized, posing a significant risk to the organelle’s survival and necessitating prompt repair. Although numerous mechanisms for lysosomal repair have been identified in recent years, the progression of lysosome-related diseases is more closely linked to the organelle’s alternative strategies when repair mechanisms fail, particularly in the contexts of aging and pathogen infection. This review explores lysosomal responses to damage, including the secretion of lysosomal contents and the interactions with lysosome-associated organelles in the endolysosomal system. Furthermore, it examines the role of organelles outside this system, such as the endoplasmic reticulum (ER) and Golgi apparatus, as auxiliary organelles of the endolysosomal system. These alternative strategies are crucial to understanding disease progression. For instance, the secretion and spread of misfolded proteins play key roles in neurodegenerative disease advancement, while pathogen escape via lysosomal secretion and lysosomotropic drug expulsion underlie cancer treatment resistance. Reexamining these lysosomal fallback strategies could provide new perspectives on lysosomal biology and their contribution to disease progression.

Lipid Dynamics at Membrane Contact Sites.

In eukaryotes, lipid building blocks for cellular membranes are made largely in the endoplasmic reticulum and then redistributed to other organelles. Lipids are transported between organelles by vesicular trafficking or else by proteins located primarily at sites where different organelles are closely apposed. Here we discuss transport at organelle contact sites mediated by shuttle-like proteins that carry single lipids between membranes to fine-tune their composition and by the more recently discovered bridge-like proteins that tether two organelles and provide a path for bulk lipid movement. Protein-mediated lipid transport is assisted by integral membrane proteins that have roles in (a) lowering the energy barrier for lipid transfer between the membrane and the lipid transfer protein, a key parameter determining the transfer rate, and (b) scrambling lipids to counteract the bilayer asymmetry that would result from such transfer. Advances in this field are shedding new light on a variety of physiological mechanisms.

Proteogenomic characterization reveals tumorigenesis and progression of lung cancer manifested as subsolid nodules

Lung adenocarcinoma (LUAD) radiologically displayed as subsolid nodules (SSNs) is prevalent. Nevertheless, the precise clinical management of SSNs necessitates a profound understanding of their tumorigenesis and progression. Here, we analyze 66 LUAD displayed as SSNs covering 3 histological stages including adenocarcinoma in situ (AIS), minimally invasive adenocarcinoma (MIA) and invasive adenocarcinoma (IAC) by incorporating genomics, proteomics, phosphoproteomics and glycoproteomics. Intriguingly, cholesterol metabolism is aberrantly regulated in the preneoplastic AIS stage. Importantly, target ablation of proprotein convertase subtilisin/kexin type 9 (PCSK9) promotes the initiation of LUAD. Furthermore, sustained endoplasmic reticulum stress is demonstrated to be a hallmark and a reliable biomarker of AIS progression to IAC. Consistently, target promotion of ER stress profoundly retards LUAD progression. Our study provides comprehensive proteogenomic landscape of SSNs, sheds lights on the tumorigenesis and progression of SSNs and suggests preventive and therapeutic strategies for LUAD.

Suppression of stress granule formation is a vulnerability imposed by mutant p53

Missense mutations in the TP53 (p53) gene have been linked to malignant progression. However, our in-silico analyses reveal that hepatocellular carcinoma (HCC) patients with mutant p53 (mutp53) have better overall survival compared to those with p53-null (p53null) HCC, unlike other cancer types. Given the historical use of sorafenib (SOR) monotherapy for advanced HCC, we hypothesize that mutp53 increases sensitivity to SOR, a multikinase inhibitor that induces endoplasmic reticulum (ER) stress. Here we show that mutp53 inhibits stress granule (SG) formation by binding to an ER stress sensor, PKR-like ER kinase (PERK), and a key SG component, GAP SH3 domain-binding protein 1 (G3BP1), contributing to increased sensitivity of SG-competent cells and xenografts to ER stress inducers including SOR. Our study identifies a unique vulnerability imposed by mutp53, suggesting mutp53 as a biomarker for ER stress-inducing agents and highlighting the importance of SG inhibition for cancer treatment.

Patterns of Evolution of the Conceptual Apparatus of Histology and Cytology (17th – 21st Centuries)

ABSTRACT The development of the conceptual apparatus of morphological sciences began to be carried out on a scientific basis starting from the 18th century. To designate many histological structures, they began to use terms already used to designate other (non-histological) structures (tissue, cell, fiber, membrane, plexus, node, membrane, nucleus, colloid, etc.), as well as terms based on a combination of known terms with clarifying additions. At the same time, many new terms were created (centrosome, mitochondria, endoplasmic reticulum, etc.). Many histological and cytological eponyms arose. For many terms, the authorship is unknown or debatable. The modern conceptual apparatus of histology was mainly formed by the middle of the 20th century and was officially formed in the second half of the 20th century. The result of the evolution of the system of histological terminology was the creation of the International Histological Nomenclature (later renamed the International Histological Terminology), in which eponyms were practically absent. Following the creation of this terminology, national histological terminologies were created in a number of countries on its basis, which are periodically revised and supplemented. At present, histology is one of the most systematically organized scientific and educational disciplines.

SH3KBP1 promotes skeletal myofiber formation and functionality through ER/SR architecture integrity

Abstract Dynamic changes in the arrangement of myonuclei and the organization of the sarcoplasmic reticulum are important determinants of myofiber formation and muscle function. To find factors associated with muscle integrity, we perform an siRNA screen and identify SH3KBP1 as a new factor controlling myoblast fusion, myonuclear positioning, and myotube elongation. We find that the N-terminus of SH3KBP1 binds to dynamin-2 while the C-terminus associates with the endoplasmic reticulum through calnexin, which in turn control myonuclei dynamics and ER integrity, respectively. Additionally, in mature muscle fibers, SH3KBP1 contributes to the formation of triads and modulates the Excitation-Contraction Coupling process efficiency. In Dnm2R465W/+ mice, a model for centronuclear myopathy (CNM), depletion of Sh3kbp1 expression aggravates CNM-related atrophic phenotypes and impaired autophagic flux in mutant skeletal muscle fiber. Altogether, our results identify SH3KBP1 as a new regulator of myofiber integrity and function.

Physiological and Transcriptomic Analyses Reveal the Mechanisms of Ilex chinensis Response to Different Types of Simulated Acid Rain

Acid rain has many negative effects on the ecological environment and poses serious abiotic stress onto plants, resulting in substantial ecological and economic impairments annually. Ilex chinensis, a well-known medicinal plant, is sensitive to acid rain, but its response mechanisms are unclear. In this study, we simulated sulfuric acid rain (SAR), mixed acid rain (MIX), and nitric acid rain (NAR) at different pH values to investigate their effects on growth condition, photosynthesis, antioxidants, and nitrogen metabolites. We also explored the metabolic pathways and key genes involved in the response of I. chinensis to acid rain through transcriptome analysis. Physiological analysis showed that I. chinensis suffered the most significant inhibition at pH 3.0, which is manifested in the decrease in height growth rate, specific leaf weight, photosynthetic pigments content, net photosynthetic rate, stomatal conductance, and transpiration rate; the increase in MDA content and SOD activity; and the reduction in glutamine synthetase activity, nitrogen content, and proline content. Transcriptome analysis isolated 314 and 21 shared differentially expressed genes (DEGs) from I. chinensis treated with acid rain at pH 3.0 for 5 d and 15 d, respectively. KEGG enrichment analysis found that different types of acid rain caused changes in multiple metabolic pathways of I. chinensis, and the shared DEGs in 5 d treatment were mainly enriched in ribosomes, oxidative phosphorylation, and glycolysis/glycolysis, etc. The shared DEGs in 115 d treatment were mainly enriched in sulfur metabolism, RNA polymerase, cysteine and methionine metabolism, etc. Further research on gene regulatory networks at the two time points showed that the key pathways of I. chinensis, in response to acid rain stress, include plant–pathogen interaction, MAPK signaling pathway-plant, protein processing in the endoplasmic reticulum, ubiquitin mediated proteolysis, etc., in which 6 hub genes were identified, including TRINITY_DN13584_c0_g1, TRINITY_DN164_c0_g4, TRINITY_DN654_c0_g1, TRINITY_DN13611_c1_g2, TRINITY_DN21290_c0_g2, TRINITY_DN44216_c0_g1. Our findings provide a basis for exploring the regulatory mechanisms of I. chinensis in response to acid rain at the physiological and molecular levels, and for identifying candidate genes with acid tolerance potential.

Transcriptomic Analysis and Identification of Candidate Genes Involved in Rhizome Development in Agropyron michnoi

Agropyron michnoi is a perennial grass with rhizomes in the genus Agropyron. It has a strong tolerance to drought and low temperature, and it is an established species in sandy flat and hilly slope lands, which constitute sandy grassland. So, it is an important forage species in dry grassland and desert steppes. Rhizomes not only enable asexual reproducibility but also confer strong resilience to stresses in A. michnoi. However, during production and utilization, it has been found that there are significant differences in the development of rhizomes among individuals of A. michnoi, yet the regulatory mechanism remains unclear. Therefore, in this study, the A. michnoi ‘Baiyinxile’ was used as the material, and the anatomical structures of the rhizomes, roots, and stems were analyzed using the paraffin sectioning technique. The results showed that the anatomical structure composition of the cross-section of the rhizome was similar to that of the root, while the arrangement of the vascular bundles in the stele was different from that of the root but similar to that of the stem. Subsequently, the Agropyron michnoi plants were classified into two types: plants with rhizomes and plants without rhizomes. Root, stem, and rhizome samples were collected from each type, and RNA sequencing was conducted. De novo transcriptomic analysis was performed to identify the candidate genes involved in rhizome development. From the RNA sequencing, a total of 103.73 Gb clean bases were obtained, from which 215,282 unigenes with an average length of 905.67 bp were assembled. Among these unigenes, 161,175 (74.87%) were functionally annotated based on seven common public databases. From pairwise comparisons of differentially expressed genes between the five samples, 129 candidate genes that are potentially specifically expressed in rhizomes were selected. Pathway enrichment analysis revealed that the rhizome-expressed genes are highly enriched in pathways of phenylpropanoid biosynthesis and starch and sucrose metabolism. The rhizome-specific expression pattern of 10 of the 129 candidate genes was further validated using qRT-PCR. Through the analysis of metabolites, 11 metabolites closely related to rhizome development, such as choline and betaine, were successfully identified. CYP family genes were selected for functional verification, and phylogenetic analysis revealed that CYP86B1 was grouped with CYP 86B1 of species such as Triticum aestivum and Lolium rigidum and was named AmrCYP86B1. The cloning results showed that its size was 1599 bp, and its subcellular localization was in the endoplasmic reticulum. Through stable genetic transformation, the study found that AmrCYP86B1 can promote the development of plant roots and stems and increase the dry matter content of the roots. Hormone detection showed that overexpression of AmrCYP 86B1 decreased the content of ABA hormone and increased the content of GA3 hormone in the plants. Combined with previous studies, it was determined that AmrCYP 86B1 promoted rhizome elongation by regulating ABA and GA3 hormones. The selected candidate genes involved in rhizome development, along with the preliminary functional verification, provide a preliminary mechanistic interpretation of rhizome development. This will contribute to in-depth research on the molecular mechanism of rhizome development in A. Michnoi.

Protein disulfide isomerase is essential for osteoblast differentiation in mice

Protein disulfide isomerase (PDI) is an oxidoreductase responsible for the formation, reduction and isomerization of disulfide bonds of nascent proteins in endoplasmic reticulum (ER). So far, the role of PDI in bone biology has never been characterized using genetically-modified animal models. In this study we generated osteoblast- specific PDI-deficient mice by crossing PDI-floxed (PDIfl/fl) mice with Osx-Cre mice. Compared with their littermate control PDIfl/fl mice, homozygous osteoblast-knockout mice (Osx-Cre/PDIfl/fl) were embryonically lethal, but heterozygous knockout mice (Osx-Cre/PDIfl/wt) displayed significantly pronounced growth retardation and reduced bone length. Besides, the decreases in bone density, osteoblast and osteoclast numbers, collagen fiber content and bone formation rate were observed in Osx-Cre/PDIfl/wt mice. Osteoblast precursors isolated from PDIfl/fl mice were infected with Cre recombinant adenovirus to produce PDI-deficient osteoblasts, followed by induction of differentiation. Osteoblasts deficient of PDI had decreased alkaline phosphatase activity, mineralizing capacity, and differentiation. Quantitative protein mass spectrometry analysis and immunoblotting showed that PDI deficiency markedly decreased the expression of the α-subunits of collagen prolyl 4-hydroxylase (C-P4H), including P4HA1, P4HA2 and P4HA3. These results demonstrate that PDI plays an essential role in osteoblast differentiation and bone formation and is required for the expression of the α-subunit of C-P4H in osteoblasts.

Cannabinoids Activate Endoplasmic Reticulum Stress Response and Promote the Death of Avian Retinal Müller Cells in Culture

Background/Objectives: Activation of cannabinoid CB1 or CB2 receptors induces the death of glial progenitors from the chick retina in culture. Here, by using an enriched retinal glial cell culture, we characterized some mechanisms underlying glial death promoted by cannabinoids. Methods and Results: Retinal cultures obtained from 8-day-old (E8) chick embryos and maintained for 12–15 days (C12–15) were used. MTT assays revealed that the CB1/CB2 agonist WIN 55,212-2 (WIN) decreased cell viability in the cultures in a time-dependent manner, with a concomitant increase in extracellular LDH activity, suggesting membrane integrity loss. Cell death was also dose-dependently induced by cannabidiol (CBD), Δ9-tetrahydrocannabinol (THC), and CP55940, another CB1/CB2 agonist. In contrast to WIN-induced cell death that was not blocked by either antagonist, the deleterious effect of CBD was blocked by the CB2 receptor antagonist SR144528, but not by PF514273, a CB1 receptor antagonist. WIN-treated cultures showed glial cells with large vacuoles in cytoplasm that were absent in cultures incubated with WIN plus 4-phenyl-butyrate (PBA), a chemical chaperone. Since cannabinoids induced the phosphorylation of eukaryotic initiation factor 2-alfa (eIF2α), these results suggest a process of endoplasmic reticulum (ER) swelling and stress. Incubation of the cultures with WIN for 4 h induced a ~five-fold increase in the number of cells labeled with the ROS indicator CM-H2DCFDA. WIN induced the phosphorylation of JNK but not of p38 in the cultures, and also induced an increase in the number of glial cells expressing cleaved-caspase 3 (c-CASP3). The decrease in cell viability and the expression of c-CASP3 was blocked by salubrinal, an inhibitor of eIF2α dephosphorylation. Conclusions: These data suggest that cannabinoids induce the apoptosis of glial cells in culture by promoting ROS production, ER stress, JNK phosphorylation, and caspase-3 processing. The graphical abstract was created at Biorender.com.