Enhanced Endoplasmic Reticulum Stress Research Articles

CD44 correlates with longevity and enhances basal ATF6 activity and ER stress resistance

The naked mole rat (NMR) is the longest-lived rodent, resistant to multiple age-related diseases including neurodegeneration. However, the mechanisms underlying the NMR's resistance to neurodegenerative diseases remain elusive. Here, we isolated oligodendrocyte progenitor cells (OPCs) from NMRs and compared their transcriptome with that of other mammals. Extracellular matrix (ECM) genes best distinguish OPCs of long- and short-lived species. Notably, expression levels of CD44, an ECM-binding protein that has been suggested to contribute to NMR longevity by mediating the effect of hyaluronan (HA), are not only high in OPCs of long-lived species but also positively correlate with longevity in multiple cell types/tissues. We found that CD44 localizes to the endoplasmic reticulum (ER) and enhances basal ATF6 activity. CD44 modifies proteome and membrane properties of the ER and enhances ER stress resistance in a manner dependent on unfolded protein response regulators without the requirement of HA. HA-independent role of CD44 in proteostasis regulation may contribute to mammalian longevity.

Inhibition of TRPA1, Endoplasmic Reticulum Stress, Human Airway Epithelial Cell Damage, and Ectopic MUC5AC Expression by Vasaka (Adhatoda vasica; Malabar Nut) Tea.

This study tested whether a medicinal plant, Vasaka, typically consumed as a tea to treat respiratory malaise, could protect airway epithelial cells (AECs) from wood smoke particle-induced damage and prevent pathological mucus production. Wood/biomass smoke is a pneumotoxic air pollutant. Mucus normally protects the airways, but excessive production can obstruct airflow and cause respiratory distress. Vasaka tea pre- and co-treatment dose-dependently inhibited mucin 5AC (MUC5AC) mRNA induction by AECs treated with wood smoke particles. This correlated with transient receptor potential ankyrin-1 (TRPA1) inhibition, an attenuation of endoplasmic reticulum (ER) stress, and AEC damage/death. Induction of mRNA for anterior gradient 2, an ER chaperone/disulfide isomerase required for MUC5AC production, and TRP vanilloid-3, a gene that suppresses ER stress and wood smoke particle-induced cell death, was also attenuated. Variable inhibition of TRPA1, ER stress, and MUC5AC mRNA induction was observed using selected chemicals identified in Vasaka tea including vasicine, vasicinone, apigenin, vitexin, isovitexin, isoorientin, 9-oxoODE, and 9,10-EpOME. Apigenin and 9,10-EpOME were the most cytoprotective and mucosuppressive. Cytochrome P450 1A1 (CYP1A1) mRNA was also induced by Vasaka tea and wood smoke particles. Inhibition of CYP1A1 enhanced ER stress and MUC5AC mRNA expression, suggesting a possible role in producing protective oxylipins in stressed cells. The results provide mechanistic insights and support for the purported benefits of Vasaka tea in treating lung inflammatory conditions, raising the possibility of further development as a preventative and/or restorative therapy.

Pseudophosphatase STYXL1 depletion enhances glucocerebrosidase trafficking to lysosomes via ER stress.

Pseudophosphatases are catalytically inactive but share sequence and structural similarities with classical phosphatases. STYXL1 is a pseudophosphatase that belongs to the family of dual-specificity phosphatases and is known to regulate stress granule formation, neurite formation and apoptosis in different cell types. However, the role of STYXL1 in regulating cellular trafficking or the lysosome function has not been elucidated. Here, we show that the knockdown of STYXL1 enhances the trafficking of β-glucocerebrosidase (β-GC) and its lysosomal activity in HeLa cells. Importantly, the STYXL1-depleted cells display enhanced distribution of endoplasmic reticulum (ER), late endosome and lysosome compartments. Further, knockdown of STYXL1 causes the nuclear translocation of unfolded protein response (UPR) and lysosomal biogenesis transcription factors. However, the upregulated β-GC activity in the lysosomes is independent of TFEB/TFE3 nuclear localization in STYXL1 knockdown cells. The treatment of STYXL1 knockdown cells with 4-PBA (ER stress attenuator) significantly reduces the β-GC activity equivalent to control cells but not additive with thapsigargin, an ER stress activator. Additionally, STYXL1-depleted cells show the enhanced contact of lysosomes with ER, possibly via increased UPR. The depletion of STYXL1 in human primary fibroblasts derived from Gaucher patients showed moderately enhanced lysosomal enzyme activity. Overall, these studies illustrated the unique role of pseudophosphatase STYXL1 in modulating the lysosome function both in normal and lysosome-storage disorder cell types. Thus, designing small molecules against STYXL1 possibly can restore the lysosome activity by enhancing ER stress in Gaucher disease.

Astragalus Polysaccharide Promotes Doxorubicin-Induced Apoptosis by Reducing O-GlcNAcylation in Hepatocellular Carcinoma

The toxicity and side effects of chemotherapeutic drugs remain a crucial obstacle to the clinical treatment of hepatocellular carcinoma (HCC). Identifying combination therapy from Chinese herbs to enhance the sensitivity of tumors to chemotherapeutic drugs is of particular interest. Astragalus polysaccharide (APS), one of the natural active components in Astragalus membranaceus, has been reported to exhibit anti-tumor properties in diverse cancer cell lines. The aim of this study was to determine the effect of APS on Doxorubicin (Dox)-induced apoptosis in HCC and the underlying mechanism. The results showed that APS dose-dependently promoted Dox-induced apoptosis and enhanced endoplasmic reticulum (ER) stress. Additionally, APS decreased the mRNA level and protein stability of O-GlcNAc transferase (OGT), and increased the O-GlcNAcase (OGA) expression. Furthermore, OGT lentiviral transfection or PugNAc (OGA inhibitor) treatment reversed the ER stress and apoptosis induced by the combination of Dox and APS. A xenograft tumor mouse model confirmed that the combination of APS and Dox showed an advantage in inhibiting tumor growth in vivo. These findings suggested that APS promoted Dox-induced apoptosis in HCC cells through reducing the O-GlcNAcylation, which led to the exacerbation of ER stress and activation of apoptotic pathways.

Role of NAT10-mediated ac4C-modified HSP90AA1 RNA acetylation in ER stress-mediated metastasis and lenvatinib resistance in hepatocellular carcinoma

Emerging evidence showed that epigenetic regulation plays important role in the pathogenesis of HCC. N4-acetocytidine (ac4C) was an acetylation chemical modification of mRNA, and NAT10 is reported to regulate ac4C modification and enhance endoplasmic reticulum stress (ERS) in tumor metastasis. Here, we report a novel mechanism by which NAT10-mediated mRNA ac4C-modified HSP90AA1 regulates metastasis and tumor resistance in ERS of HCC. Immunohistochemical, bioinformatics analyses, and in vitro and in vivo experiments, e.g., acRIP-Seq, RNA-Seq, and double luciferase reporter experiment, were employed to investigate the effect of NAT10 on metastasis and drug resistance in HCC. The increased expression of NAT10 was associated with HCC risk and poor prognosis. Cell and animal experiments showed that NAT10 enhanced the metastasis ability and apoptosis resistance of HCC cells in ERS and ERS state. NAT10 could upregulate the modification level of HSP90AA1 mRNA ac4C, maintain the stability of HSP90AA1, and upregulate the expression of HSP90AA1, which further promotes the metastasis of ERS hepatoma cells and the resistance to apoptosis of Lenvatinib. This study proposes a novel mechanism by which NAT10-mediated mRNA ac4C modification regulates tumor metastasis. In addition, we demonstrated the regulatory effect of NAT10-HSP90AA1 on metastasis and drug resistance of ERS in HCC cells.

Dihydroartemisinin elicits immunogenic death through ferroptosis-triggered ER stress and DNA damage for lung cancer immunotherapy

BackgroundThe immunosuppressive microenvironment of lung cancer serves as an important endogenous contributor to treatment failure. The present study aimed to demonstrate the promotive effect of DHA on immunogenic cell death (ICD) in lung cancer as well as the mechanism. MethodsThe lewis lung cancer cells (LLC), A549 cells and LLC-bearing mice were applied as the lung cancer model. The apoptosis, ferroptosis assay, western blotting, immunofluorescent staining, qPCR, comet assay, flow cytometry, confocal microscopy, transmission electron microscopy and immunohistochemistry were conducted to analyze the functions and the underlying mechanism. ResultsAn increased apoptosis rate and immunogenicity were detected in DHA-treated LLC and tumor grafts. Further findings showed DHA caused lipid peroxide (LPO) accumulation, thereby initiating ferroptosis. DHA stimulated cellular endoplasmic reticulum (ER) stress and DNA damage simultaneously. However, the ER stress and DNA damage induced by DHA could be abolished by ferroptosis inhibitors, whose immunogenicity enhancement was synchronously attenuated. In contrast, the addition of exogenous iron ions further improved the immunogenicity induced by DHA accompanied by enhanced ER stress and DNA damage. The enhanced immunogenicity could be abated by ER stress and DNA damage inhibitors as well. Finally, DHA activated immunocytes and exhibited excellent anti-cancer efficacy in LLC-bearing mice. ConclusionsIn summary, the current study demonstrates that DHA triggers ferroptosis, facilitating the ICD of lung cancer thereupon. This work reveals for the first time the effect and underlying mechanism by which DHA induces ICD of cancer cells, providing novel insights into the regulation of the immune microenvironment for cancer immunotherapy by Chinese medicine phytopharmaceuticals.

NEAT1-SOD2 Axis Confers Sorafenib and Lenvatinib Resistance by Activating AKT in Liver Cancer Cell Lines.

This study investigated the effects of a long noncoding RNA, nuclear paraspeckle assembly transcript 1 (NEAT1) variant 1 (NEAT1v1) on drug resistance in liver cancer cell lines. NEAT1 knockdown activated mitogen-activated protein kinase (MAPK) signaling pathways, including MAPK kinase (MEK)/extracellular signal-regulated kinase (ERK), but suppressed AKT. Moreover, NEAT1 knockdown sensitized liver cancer cells to sorafenib and lenvatinib, both clinically used for treating hepatocellular carcinoma, whereas it conferred resistance to an AKT-targeted drug, capivasertib. NEAT1v1 overexpression suppressed MEK/ERK and activated AKT, resulting in resistance to sorafenib and lenvatinib and sensitization to capivasertib. Superoxide dismutase 2 (SOD2) knockdown reverted the effects of NEAT1v1 overexpression on the sensitivity to the molecular-targeted drugs. Although NEAT1 or SOD2 knockdown enhanced endoplasmic reticulum (ER) stress, concomitant with the suppression of AKT, taurodeoxycholate, an ER stress suppressor, did not restore AKT activity. Although further in vivo and clinical studies are needed, these results suggested that NEAT1v1 switches the growth modality of liver cancer cell lines from MEK/ERK-dependent to AKT-dependent mode via SOD2 and regulates sensitivity to the molecular-targeted drugs independent of ER stress.

Dysregulated UPR and ER Stress Related to a Mutation in the Sdf2l1 Gene Are Involved in the Pathophysiology of Diet-Induced Diabetes in the Cohen Diabetic Rat.

The Cohen Diabetic rat is a model of type 2 diabetes mellitus that consists of the susceptible (CDs/y) and resistant (CDr/y) strains. Diabetes develops in CDs/y provided diabetogenic diet (DD) but not when fed regular diet (RD) nor in CDr/y given either diet. We recently identified in CDs/y a deletion in Sdf2l1, a gene that has been attributed a role in the unfolded protein response (UPR) and in the prevention of endoplasmic reticulum (ER) stress. We hypothesized that this deletion prevents expression of SDF2L1 and contributes to the pathophysiology of diabetes in CDs/y by impairing UPR, enhancing ER stress, and preventing CDs/y from secreting sufficient insulin upon demand. We studied SDF2L1 expression in CDs/y and CDr/y. We evaluated UPR by examining expression of key proteins involved in both strains fed either RD or DD. We assessed the ability of all groups of animals to secrete insulin during an oral glucose tolerance test (OGTT) over 4 weeks, and after overnight feeding (postprandial) over 4 months. We found that SDF2L1 was expressed in CDr/y but not in CDs/y. The pattern of expression of proteins involved in UPR, namely the PERK (EIF2α, ATF4 and CHOP) and IRE1 (XBP-1) pathways, was different in CDs/y DD from all other groups, with consistently lower levels of expression at 4 weeks after initiation of DD and coinciding with the development of diabetes. In CDs/y RD, insulin secretion was mildly impaired, whereas in CDs/y DD, the ability to secrete insulin decreased over time, leading to the development of the diabetic phenotype. We conclude that in CDs/y DD, UPR participating proteins were dysregulated and under-expressed at the time point when the diabetic phenotype became overt. In parallel, insulin secretion in CDs/y DD became markedly impaired. Our findings suggest that under conditions of metabolic load with DD and increased demand for insulin secretion, the lack of SDF2L1 expression in CDs/y is associated with UPR dysregulation and ER stress which, combined with oxidative stress previously attributed to the concurrent Ndufa4 mutation, are highly likely to contribute to the pathophysiology of diabetes in this model.

Brigatinib-repurposed chemo-photodynamic therapy nanoplatform via effective apoptosis against colorectal cancer

Colorectal cancer (CRC), a malignancy with a relatively high fatality rate, still poses a growing global public health challenge due to the lack of well-established and effective treatment strategies. Brigatinib, an ALK inhibitor approved for the treatment of ALK-positive NSCLC, has been repurposed to suppress the growth of ALK-negative CRC cells via sustained endoplasmic reticulum (ER) stress. Moreover, the photosensitizer Chlorine e6 (Ce6) exhibits a high singlet oxygen quantum yield, which promote the generation of reactive oxygen species (ROS) to exacerbate ER stress and function synergistically with chemotherapeutics. However, the clinical application of both brigatinib and Ce6 is limited by their poor water solubility. Therefore, a repurposed chemo-photodynamic therapy nanoplatform (BCT NPs) utilizing d-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) as a nanocarrier for the codelivery of the chemotherapeutics brigatinib and photosensitizer Ce6 is developed, which is capable of disrupting redox homeostasis and enhancing ER stress against CRC. The developed nanoplatform successfully addressed therapeutic drug solubility, improved brigatinib biocompatibility with a lower hemolysis rate, and demonstrated satisfactory CRC suppression effects in vitro and in vivo. Overall, BCT NPs provide a promising strategy to construct repurposed and versatile nanoplatforms against CRC.

The developmental regulator HAND1 inhibits gastric carcinogenesis through enhancing ER stress apoptosis via targeting CHOP and BAK which is augmented by cisplatin.

Epigenetic disruption of tumor suppressor genes, particularly aberrant CpG methylation, plays a crucial role in gastric cancer (GC) pathogenesis. Through CpG methylome and expression profiling, a developmental transcription factor - Hand-And-Neural-crest-Derivative-expressed 1 (HAND1), was identified methylated and downregulated in GC. However, its role and underlying mechanisms in GC progression are poorly understood. Here, we show that HAND1 was frequently downregulated in GC by promoter methylation, and significantly correlated with tumor progression and poor prognosis of GC patients. High expression of HAND1 in GC patients was associated with significantly higher 5-year overall survival rates. Ectopic expression of HAND1 inhibited GC cell growth and migration in vitro and in vivo. HAND1 expression increased ROS levels and cytosolic Ca2+ concentration, enhanced cisplatin-induced apoptosis through endoplasmic reticulum (ER) stress/mitochondria-mediated apoptosis. Knockdown of CHOP and BAK attenuated HAND1-induced cell apoptosis. Overexpression of CHOP increased BAK expression. HAND1 interacts with CHOP, also directly binds to CHOP and BAK promoters and positively regulates BAK transcription. Thus, the present study demonstrates that HAND1 is a tumor suppressor gene methylated in GC, induces ER stress and apoptosis via CHOP and BAK, which is augmented by cisplatin. Low HAND1 expression is an independent poor prognostic factor for GC. The tumor-specific methylation of HAND1 promoter could be a candidate biomarker for GC.

Remdesivir inhibits the progression of glioblastoma by enhancing endoplasmic reticulum stress

Glioblastoma (GBM) is one of the most aggressive primary malignant brain tumors. The major challenge is the lack of effective therapeutic drugs due to the blood–brain barrier (BBB) and tumor heterogeneity. Remdesivir (RDV), a new member of the nucleotide analog family, has previously been shown to have excellent antiviral effects and BBB penetration, and was predicted here to have anti-GBM effects. In vitro experiments, RDV significantly inhibited the growth of GBM cells, with IC50 values markedly lower than those of normal cell lines or the same cell lines treated with temozolomide. Moreover, in multiple mouse models, RDV not only distinctly inhibited the progression and improved the prognosis of GBM but also exhibited a promising biosafety profile, as manifested by the lack of significant body weight loss, liver or kidney dysfunction or organ structural damage after administration. Furthermore, we investigated the anti-GBM mechanism by RNA-seq and identified that RDV might induce apoptosis of GBM cells by enhancing endoplasmic reticulum (ER) stress and activating the PERK-mediated unfolded protein response. In conclusion, our results indicated that RDV might serve as a novel agent for GBM treatment by increasing ER stress and inducing apoptosis in GBM cells.

GZ17-6.02 kills prostate cancer cells in vitro and in vivo.

GZ17-6.02 is undergoing clinical evaluation in solid tumors and lymphoma. We defined the biology of GZ17-6.02 in prostate cancer cells and determined whether it interacted with the PARP1 inhibitor olaparib to enhance tumor cell killing. GZ17-6.02 interacted in a greater than additive fashion with olaparib to kill prostate cancer cells, regardless of androgen receptor expression or loss of PTEN function. Mechanistically, GZ17-6.02 initially caused peri-nuclear activation of ataxia-telangiectasia mutated (ATM) that was followed after several hours by activation of nuclear ATM, and which at this time point was associated with increased levels of DNA damage. Directly downstream of ATM, GZ17-6.02 and olaparib cooperated to activate the AMP-dependent protein kinase (AMPK) which then activated the kinase ULK1, resulting in autophagosome formation that was followed by autophagic flux. Knock down of ATM, AMPKα or the autophagy-regulatory proteins Beclin1 or ATG5 significantly reduced tumor cell killing. GZ17-6.02 and olaparib cooperated to activate protein kinase R which phosphorylated and inactivated eIF2α, i.e., enhanced endoplasmic reticulum (ER) stress signaling. Knock down of eIF2α also significantly reduced autophagosome formation and tumor cell killing. We conclude that GZ17-6.02 and olaparib interact to kill prostate cancer cells in vitro by increasing autophagy and by enhancing ER stress signaling. In vivo, GZ17-6.02 as a single agent profoundly reduced tumor growth and significantly prolonged animal survival. GZ17-6.02 interacted with olaparib to further suppress the growth of LNCaP tumors without ultimately enhancing animal survival. Our data support the consideration of GZ17-6.02 as a possible therapeutic agent in patients with AR+ prostate cancer.

Jatrorrhizine Improves Endothelial Function in Diabetes and Obesity through Suppression of Endoplasmic Reticulum Stress.

Jatrorrhizine (JAT) is one of the major bioactive protoberberine alkaloids found in rhizoma coptidis, which has hypoglycemic and hypolipidemic potential. This study aimed to evaluate the vasoprotective effects of JAT in diabetes and obesity and the underlying mechanism involved. Mouse aortas, carotid arteries and human umbilical cord vein endothelial cells (HUVECs) were treated with risk factors (high glucose or tunicamycin) with and without JAT ex vivo and in vitro. Furthermore, aortas were obtained from mice with chronic treatment: (1) control; (2) diet-induced obese (DIO) mice fed a high-fat diet (45% kcal% fat) for 15 weeks; and (3) DIO mice orally administered JAT at 50 mg/kg/day for the last 5 weeks. High glucose or endoplasmic reticulum (ER) stress inducer tunicamycin impaired acetylcholine-induced endothelium-dependent relaxations (EDRs) in mouse aortas, induced oxidative stress in carotid arteries and HUVECs, downregulated phosphorylations of Akt at Ser473 and eNOS at Ser1177 and enhanced ER stress in mouse aortas and HUVECs, and these impairments were reversed by cotreatment with JAT. JAT increased NO release in high-glucose-treated mouse aortas and HUVECs. In addition, chronic JAT treatment restored endothelial function with EDRs comparable to the control, increased Akt/eNOS phosphorylation, and attenuated ER stress and oxidative stress in aortas from DIO mice. Blood pressure, glucose sensitivity, fatty liver and its morphological change, as well as plasma levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and plasma lipid profile, were also normalized by JAT treatment. Collectively, our data may be the first to reveal the vasoprotective effect of JAT that ameliorates endothelial dysfunction in diabetes and obesity through enhancement of the Akt/eNOS pathway and NO bioavailability, as well as suppression of ER stress and oxidative stress.

Dysfunction of Cytotoxic T Lymphocyte Induced by Hepatoma Cells through the Gln-GLS2-Endoplasmic Reticulum Stress Pathway.

Metabolic activities of tumor cells lead to a depletion of nutrients within the tumor microenvironment, which results in the dysfunction of infiltrating T cells. Here, we explored how glutamine (gln) metabolism, which is essential for biosynthesis and cellular function, can affect the functions of cytotoxic T lymphocytes (CTLs). Activated CTLs were co-cultured with hepatoma cells. Western blot was used to analyze changes of proteins and ELISA was used to analyze changes of effector. RNA-sequencing was used to detect differentially expressed genes in CTLs. The status of the endoplasmic reticulum (ER) was investigated using transmission electron microscopy experiments. Co-culturing CTLs and hepatoma cells revealed that CTLL-2 cells in the co-culture group expressed high levels of PD-1 (Programmed cell death protein 1), TIM-3 (T cell immunoglobulin and mucin domain-containing protein-3), GRP78 (Glucose regulated protein 78), and P-PERK (phosphorylated protein kinase RNA-activated-like endoplasmic reticulum kinase) and secreted low levels of Granzyme B and perforin. Additionally, the substructure of the ER was severely damaged. When CTLs were treated with an inhibitor of ER stress, their functions were restored. Next, complete medium without Gln was used to culture cells, causing CTLs to display dysfunction and ER stress. WB results revealed decreased expression levels of GLS2 and SLC1A5 (Solute carrier family 1 member 5) in CTLs in the co-culture group. Subsequently, glutaminase (GLS) inhibitors were added to the cultures. As expected, CTLs treated with a GLS2 inhibitor had increased protein content of PD-1 and TIM-3, decreased secretion of Granzyme B and perforin, and an enhanced ER stress response. In summary, CTLs are functionally downregulated induced by hepatoma cells through the Gln-GLS2-ERS pathway.

Effect of hyperhomocysteinemia on a murine model of smoke-induced pulmonary emphysema

Hyperhomocysteinemia was reported to enhance endoplasmic reticulum (ER) stress and subsequent apoptosis in several cells. However, the precise mechanisms of smoking susceptibility associated with hyperhomocysteinemia has not been fully elucidated. This study included 7- to 9-week-old C57BL6 male mice induced with hyperhomocysteinemia and were exposed to cigarette smoke (CS). A549 cells (human alveolar epithelial cell line) were cultured with homocysteine and were exposed to cigarette smoke extract (CSE) to observe cell viability and expression of proteins related to the ER stress. After 6 months of CS exposure, pulmonary emphysema was more severely induced in the group under the condition of hyperhomocysteinemia compared to that in the control group. The apoptotic A549 cells increased as homocysteine concentration increased and that was enhanced by CSE. Protein expression levels of ER stress markers were significantly increased after simultaneous stimulation. Notably, vitamin B12 and folate supplementation improved ER stress after simultaneous stimulation of A549 cells. In this study, we showed that hyperhomocysteinemia exacerbates CS exposure-induced emphysema in mice, suggesting that hyperhomocysteinemia and CS stimulation enhance ER stress and subsequent induced apoptosis in alveolar epithelial cells. It was suggested that there is a synergistic effect between homocysteine and CS.

Morusin Enhances Temozolomide Efficiency in GBM by Inducing Cytoplasmic Vacuolization and Endoplasmic Reticulum Stress.

Glioblastoma multiforme (GBM) is an aggressive brain tumor with high risks of recurrence and mortality. Chemoradiotherapy resistance has been considered a major factor contributing to the extremely poor prognosis of GBM patients. Therefore, there is an urgent need to develop highly effective therapeutic agents. Here, we demonstrate the anti-tumor effect of morusin, a typical prenylated flavonoid, in GBM through in vivo and in vitro models. Morusin showed selective cytotoxicity toward GBM cell lines without harming normal human astrocytes when the concentration was less than 20 µM. Morusin treatment significantly induced apoptosis of GBM cells, accompanied by the activation of endoplasmic reticulum (ER) stress, and the appearance of cytoplasmic vacuolation and autophagosomes in cells. Then, we found the ER stress activation and cytotoxicity of morusin were rescued by ER stress inhibitor 4-PBA. Furthermore, morusin arrested cell cycle at the G1 phase and inhibited cell proliferation of GBM cells through the Akt–mTOR–p70S6K pathway. Dysregulation of ERs and cell cycle in morusin exposed GBM cells were confirmed by RNA-seq analysis. Finally, we demonstrated the combination of morusin and TMZ remarkably enhanced ER stress and displayed a synergistic effect in GBM cells, and suppressed tumor progression in an orthotopic xenograft model. In conclusion, these findings reveal the toxicity of morusin to GBM cells and its ability to enhance drug sensitivity to TMZ, suggesting the potential application value of morusin in the development of therapeutic strategies for human GBM.

On the origin of metastases: Induction of pro-metastatic states after impending cell death via ER stress, reprogramming, and a cytokine storm.

How metastatic cells arise is unclear. Here, we search for the induction of recently characterized pro-metastatic states as a surrogate for the origin of metastasis. Since cell-death-inducing therapies can paradoxically promote metastasis, we ask if such treatments induce pro-metastatic states in human colon cancer cells. We find that post-near-death cells acquire pro-metastatic states (PAMEs) and form distant metastases invivo. These PAME ("let's go" in Greek) cells exhibit a multifactorial cytokine storm as well as signs of enhanced endoplasmic reticulum (ER) stress and nuclear reprogramming, requiring CXCL8, INSL4, IL32, PERK-CHOP, and NANOG. PAMEs induce neighboring tumor cells to become PAME-induced migratory cells (PIMs): highly migratory cells that re-enact the storm and enhance PAME migration. Metastases are thus proposed to originate from the induction of pro-metastatic states through intrinsic and extrinsic cues in a pro-metastatic tumoral ecosystem, driven by an impending cell-death experience involving ER stress modulation, metastatic reprogramming, and paracrine recruitment via a cytokine storm.

Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) overexpression alleviates endoplasmic reticulum stress after acute kidney injury

Background Mitochondrial biogenesis dysregulation and enhanced endoplasmic reticulum (ER) stress have been implicated in the progression of acute kidney injury (AKI). However, the interaction between these two events remains poorly understood. This study was designed to investigate the role of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) expression, a key factor in mitochondrial biogenesis, in renal ER stress at 24 h after AKI and the underlying mechanisms. Methods Mice were administered recombinant adenovirus encoding murine PGC-1α (100 μl, 1.0 × 109PFU/ml) or vehicle five days before renal ischemia reperfusion (I/R) or sham operation. Twenty-four hours after the operation, kidney and serum samples were collected for evaluation. Results We first confirmed that PGC-1α transfection elevated the PGC-1α levels and mitochondrial transcripts in the kidney 24 h after AKI. Then, we found PGC-1α overexpression improved renal function. PGC-1α transfection inhibited AKI-induced ER stress through the unfolded protein response (UPR) pathway, resulting in the suppression of apoptosis via both mitochondrial and ER pathways. Further study showed that the expression of mitofusin 2 (Mfn2), an interaction protein between mitochondria and ER, was increased after PGC-1α overexpression. We also found the expression of a novel ER stress regulator, hairy and enhancer of split 1 (Hes1), was decreased after PGC-1α transfection. Conclusions Our findings reveal that mitochondrial biogenesis plays an important role in the progression of AKI-induced ER stress and provide useful evidence for research on organelle crosstalk during AKI.

BRAF inhibition promotes ER stress-mediated cell death in uveal melanoma.

Melanoma with a BRAF mutation is more common to develop into a fatal disease. BRAF mutation inhibitor-induced autophagy affects the drug efficacy in many cancer types. The role of autophagy during BRAF inhibition in uveal melanoma (UM) remains unclear. In this study, we examined the autophagic flux and compared the number of autophagic vacuoles during the BRAF inhibition in UM. The PKR-like endoplasmic reticulum (ER) kinase (PERK) arm was studied to test whether the ER stress was involved. The effects of downregulation of ER stress by targeting the PERK arm (pharmacologically and genetically) were also assessed. We found a dose-dependent increase of autophagic flux in OCM1A cells during the BRAF inhibition. This phenomenon was further verified by an enhanced number of GFP-LC3 puncta and was finally confirmed by raised autophagic index examined by transmission electron microscopy. Pathway analysis revealed that the vemurafenib (the BRAF inhibitor)-induced autophagy was independent of the MAPK signaling pathway. Instead, it was possibly regulated via the enhanced ER stress response. We further found that the inhibition of ER stress response rescued cell death. Therefore, our results suggest BRAF inhibition promotes ER stress response-induced autophagy in UM. Targeting ER stress response can partially revert autophagy and rescue cell death, which may impair the anti-tumor effect of BRAF inhibitor in UM.

Konjac glucomannan enhances 5-FU-induced cytotoxicity of hepatocellular carcinoma cells via TLR4/PERK/CHOP signaling to induce endoplasmic reticulum stress.

5-Fluorouracil (5-FU) is a commonly used chemotherapeutic agent for various cancers. However, the drug resistance developed by tumor cells hinders the therapeutic effect. Konjac glucomannan (KGM) is indicated to sensitize 5-FU-resistant hepatocellular carcinoma (HCC) cells to 5-FU. In our study, we found that KGM or 5-FU treatment alone did not affect the malignant cell behaviors and endoplasmic reticulum (ER) stress of 5-FU-resistant HCC cells or HepG2/5-FU and Bel-7402/5-FU cells, while cotreatment with KGM and 5-FU significantly facilitated HCC cell apoptosis and ER stress and suppressed cell proliferation potential and migration abilities. Moreover, we explored the underlying mechanism by which KGM induces 5-FU cytotoxicity in HCC cells. We found that Toll-like receptor 4 (TLR4) was downregulated in KGM- and 5-FU-treated HCC cells. TLR4 overexpression reversed the KGM and 5-FU cotreatment-induced inhibition of the malignant behaviors of 5-FU-resistant HCC cells. Furthermore, KGM enhanced 5-FU-induced ER stress by inhibiting TLR4 to activate PERK/ATF4/CHOP signaling. Xenograft mouse models were established using HepG2/5-FU cells, and KGM was demonstrated to reverse 5-FU resistance in HCC tumors in vivo by suppressing TLR4 to enhance ER stress and activate PERK/ATF4/CHOP signaling. In conclusion, KGM combined with 5-FU treatment significantly promoted apoptosis and reduced cell proliferation, migration and ER stress in 5-FU-resistant HCC cells compared with KGM or 5-FU treatment alone by downregulating TLR4 to activate PERK/ATF4/CHOP signaling.