Regulation Of Endoplasmic Reticulum Stress Research Articles

GRINA alleviates hepatic ischemia‒reperfusion injury-induced apoptosis and ER-phagy by enhancing HRD1-mediated ATF6 ubiquitination.

Hepatic ischemia‒reperfusion injury (HIRI) is a critical complication of liver surgery and transplantation that contributes significantly to severe organ failure. GRINA, a calcium-regulating endoplasmic reticulum (ER) protein, plays an essential role in controlling the unfolded protein response; however, its role in HIRI remains unclear. The aim of this study was to investigate the function of GRINA in HIRI and explore its potential as a therapeutic target. Liver tissues from patients undergoing hepatectomy, alongside a mouse model of partial HIRI, were used to assess GRINA expression levels. Hepatocyte-specific Grina knockout and transgenic mouse models were generated to explore the effects of GRINA on HIRI. Key markers of inflammation, apoptosis, ER stress, and autophagy were evaluated via real-time PCR, Western blotting, immunohistochemistry, immunofluorescence, and ELISA. RNA sequencing, mass spectrometry, coimmunoprecipitation and ubiquitination assays were used to elucidate the underlying molecular mechanisms. GRINA expression was markedly reduced in hepatocytes from both patients and mice with HIRI, and its expression was inversely correlated with the severity of liver damage. Hepatocyte-specific Grina overexpression mitigated liver injury, the inflammatory response, and hepatocyte apoptosis following HIRI, whereas GRINA deficiency exacerbated these outcomes. Mechanistically, GRINA interacted directly with ATF6 and recruited HRD1 to form a multiprotein complex that catalyzed ATF6 polyubiquitination, thereby promoting its degradation. This process suppressed ER autophagy (ER-phagy), providing cellular protection following HIRI. The inhibition of ATF6 degradation attenuated the protective effects of GRINA in HIRI. Our study highlights the critical role of the GRINA-HRD1-ATF6 complex in regulating ER stress and autophagy during HIRI. These findings provide new insights into therapeutic strategies to alleviate HIRI. HIRI represents a multifaceted pathophysiological challenge commonly encountered during liver surgeries, yet its underlying molecular mechanisms remain inadequately understood. In this study, we revealed a significant negative correlation between GRINA levels and the severity of liver damage in patients with HIRI. Our findings demonstrate that GRINA alleviates endoplasmic reticulum stress by enhancing HRD1-mediated ubiquitination of ATF6, thereby maintaining calcium homeostasis and inhibiting ER-phagy. This study provides novel insights into the role of GRINA in protecting liver cells under HIRI, offering fresh perspectives for clinical prevention and management strategies for HIRI.

CHAC1 Mediates Endoplasmic Reticulum Stress-Dependent Ferroptosis in Calcium Oxalate Kidney Stone Formation.

The initiation of calcium oxalate (CaOx) kidney stone formation is highly likely to stem from injury to the renal tubular epithelial cells (RTECs) induced by stimulation from an aberrant urinary environment. CHAC1 plays a critical role in stress response mechanisms by regulating glutathione metabolism. Endoplasmic reticulum (ER) stress and ferroptosis are demonstrated to be involved in stone formation. This study attempted to elucidate the mechanism of ER stress-dependent ferroptosis and the role of CHAC1 in CaOx kidney stones. Here, regulating ER stress and CHAC1 expression are performed in in vivo and in vitro stone models. These findings indicated that 4-Phenylbutyric acid (4-PBA)treatment and CHAC1 deficiency alleviated the ferroptotic status, including restoring GSH content, suppressing Fe2+ and lipid peroxidation accumulation, as well as regulating ferroptosis-related proteins. Notably, 4-PBA treatment and CHAC1 deficiency both attenuated oxidative damage, improved renal function, importantly, decreased crystal deposition. Additionally, ChIP-seq and ChIP-qPCR analyses demonstrated that CHAC1 is the vital downstream target gene of ATF4. The results indicated that ATF4 depletion inhibited the upregulation of CHAC1 and pro-ferroptotic response induced by Ox stimulation. Overall, ATF4/CHAC1 axis mediating ER stress-dependent ferroptosis may be a promising research direction for identifying potential strategy to prevent and treat CaOx kidney stones.

LRP1 mitigates intervertebral disc degeneration by inhibiting endoplasmic reticulum stress through stabilizing the PPARγ.

Intervertebral disc degeneration (IDD) is a significant cause of lower back pain, characterized by inflammation-mediated extracellular matrix (ECM) degradation, apoptosis, and aging of nucleus pulposus (NP) cells. Identifying key regulatory targets for these processes is crucial for IDD treatment. Previous research has highlighted the role of low-density lipoprotein receptor-related protein 1 (LRP1) in regulating ECM levels and cell fate, but its role in IDD remains under-explored. This study aims to elucidate the function and mechanism of LRP1 in the progression of IDD. LRP1 expression was assessed in clinical tissue samples from patients diagnosed with IDD and in a rat IDD model established using needle puncture injuries. The effects of LRP1 knockdown and treatment with the LRP1 activator SP16 on apoptosis and ECM metabolism in NP cells were analyzed, with a focus on their relationship with endoplasmic reticulum (ER) stress. The interaction and regulatory mechanism between LRP1 and peroxisome proliferator-activated receptor gamma (PPARγ) were further explored to clarify how LRP1 regulates ER stress. Finally, the in vivo therapeutic effect of SP16 was investigated using a rat tail IDD model. We found that LRP1 expression was significantly downregulated in IDD. In NP cells with LRP1 knockdown, there was a marked increase in apoptosis and detrimental ECM remodeling, which were associated with the activation of ER stress. Our research further revealed that LRP1 interacts with PPARγ, stabilizing the PPARγ protein and preventing its lysosomal degradation, thereby mitigating ER stress. Activation of LRP1 in our models significantly reduced ER stress, matrix degradation, and apoptosis, thereby attenuating IDD both in vitro and in vivo. This study systematically investigated the role and mechanisms of the LRP1/PPARγ/ER stress signaling axis in IDD. Our findings suggest that targeting LRP1 to modulate this signaling pathway could provide a promising therapeutic approach for the treatment of IDD. Our study demonstrated that LRP1 can reduce apoptosis and ECM degradation by inhibiting ER stress through stabilizing PPARγ, indicating that targeting LRP1 may be a novel therapeutic strategy for IDD.

Comprehensive analysis of the xbp1 gene in Pacific abalone Haliotis discus hannai: Structure, expression, and role in heat stress response.

The present study explores the x-box binding protein 1 (xbp1) gene in Haliotis discus hannai (Pacific abalone), focusing on its structure, expression, and functional role under heat stress. Southern blot revealed two copies of xbp1 in the intestine and mantle, one in the gill and muscle, and no detection in the digestive gland. mRNA expression levels were highest in the gill, followed by the mantle, intestine, and muscle, with the digestive gland showing the lowest expression. Actinomycin D treatment demonstrated that xbp1 mRNA stability varied among tissues, with slower degradation in the gill and mantle, while rapid degradation was observed in the digestive gland. Heat stress caused a 20 bp fragment removal from xbp1 mRNA, producing spliced xbp1 (xbp1s), with a conserved inositol-requiring enzyme 1α (IRE1α) cleavage motif (5'-CAGCACCUGCUGAUCCUCUG-3'). Genome walking was used to obtain the promoter sequences of downstream genes regulated by xbp1s. Through sequence conservation analysis, the binding sites of xbp1s on these promoters were identified in Pacific abalone. Yeast one-hybrid (Y1H) assays confirmed xbp1s binding to these sites, and morpholino oligonucleotides (MO) treatment effectively suppressed xbp1s production. Western blot analysis demonstrated that heat stress induced the expression of HDEL-related proteins, while MO injection significantly reduced their expression under both basal and heat stress conditions. Immunofluorescence analysis revealed decreased endoplasmic reticulum (ER) chaperone glucose-regulated protein 78 (GRP78) levels and increased apoptosis in MO-treated abalone under heat stress, suggesting a compromised ER stress response. These findings underscore XBP1's crucial role in regulating ER stress management and apoptotic processes, providing new insights into the functional significance of xbp1 in abalone's response to thermal stress.

Osthole ameliorates wear particle-induced osteogenic impairment by mitigating endoplasmic reticulum stress via PERK signaling cascade

BackgroundPeriprosthetic osteolysis and subsequent aseptic loosening are the leading causes of failure following total joint arthroplasty. Osteogenic impairment induced by wear particles is regarded as a crucial contributing factor in the development of osteolysis, with endoplasmic reticulum (ER) stress identified as a key underlying mechanism. Therefore, identifying potential therapeutic targets and agents that can regulate ER stress adaption in osteoblasts is necessary for arresting aseptic loosening. Osthole (OST), a natural coumarin derivative, has demonstrated promising osteogenic properties and the ability to modulate ER stress adaption in various diseases. However, the impact of OST on ER stress-mediated osteogenic impairment caused by wear particles remains unclear.MethodsTiAl6V4 particles (TiPs) were sourced from the prosthesis of patients who underwent revision hip arthroplasty due to aseptic loosening. A mouse calvarial osteolysis model was utilized to explore the effects of OST on TiPs-induced osteogenic impairment in vivo. Primary mouse osteoblasts were employed to investigate the impact of OST on ER stress-mediated osteoblast apoptosis and osteogenic inhibition induced by TiPs in vitro. The mechanisms underlying OST-modulated alleviation of ER stress induced by TiPs were elucidated through Molecular docking, immunochemistry, PCR, and Western blot analysis.ResultsIn this study, we found that OST treatment effectively mitigated TiAl6V4 particles (TiPs)-induced osteolysis by enhancing osteogenesis in a mouse calvarial model. Furthermore, we observed that OST could attenuate ER stress-mediated apoptosis and osteogenic reduction in osteoblasts exposed to TiPs in vitro and in vivo. Mechanistically, we demonstrated that OST exerts bone-sparing effects on stressed osteoblasts upon TiPs exposure by specifically suppressing the ER stress-dependent PERK signaling cascade.ConclusionOsthole ameliorates wear particle-induced osteogenic impairment by mitigating endoplasmic reticulum stress via PERK signaling cascade. These findings suggest that OST may serve as a potential therapeutic agent for combating wear particle-induced osteogenic impairment, offering a novel alternative strategy for managing aseptic prosthesis loosening.

Pentraxin 3 deficiency ameliorates streptozotocin-induced pancreatic toxicity via regulating ER stress and β-cell apoptosis.

The long pentraxin 3 (PTX3), a marker of inflammation, has been associated with cardiovascular disease, obesity, and metabolic syndrome. Recently, elevated serum PTX3 levels have been linked to type 2 diabetes in obese patients with nonalcoholic fatty liver disease. Diabetes mellitus is a metabolic syndrome characterized by hyperglycemia resulting from insufficient insulin secretion or action. However, the precise role of PTX3 in hyperglycemia remains unclear. This study aimed to investigate the physiological roles of PTX3 in vivo. The deformation of pancreatic islets was mitigated in PTX3-deficient mice treated with streptozotocin (STZ) compared to control C57BL/6J mice. In addition, PTX3 deficiency prevented STZ-induced unfolded protein responses and pancreatic β-cell death. Immunoblotting data revealed significant inhibition of IRE1α and CHOP protein expression in PTX3 KO mice administered tunicamycin which is a chemical ER stress inducer. Similarly, tunicamycin-induced Grp78, Grp94, ATF6, and CHOP mRNA levels were reduced in PTX3 KO mice. Moreover, recombinant PTX3 induced CHOP expression and β-cell apoptosis in primary mouse islets. These findings suggest that PTX3 plays a critical role in STZ-induced the deformation of pancreatic islets via regulating ER stress and β-cell apoptosis.

Role of MFN2 in Bovine Embryonic Development and the Mitigation of ER Stress

This study investigated the role of mitochondrial fusion protein-2 (MFN2) in bovine embryonic development and its relationship with endoplasmic reticulum (ER) stress, aiming to increase the efficiency of in vitro embryo culture. Western blot analysis revealed that MFN2 expression peaked at the 2-cell stage, decreased at the 4-cell stage, and gradually increased from the 6-8-cell stage to the blastocyst stage. Inhibiting MFN2 at the zygote stage reduced blastocyst formation and proliferation, and this damage was partially reversed by the ER stress protective agent TUDCA. MFN2 inhibition also led to the decreased formation of the inner cell mass (ICM) and reduced expression of the totipotent genes CDX2 and SOX2. Additionally, reactive oxygen species (ROS) levels increased following MFN2 inhibition but decreased after TUDCA treatment. The expression of antioxidative stress-related genes (SOD and CAT) was downregulated after MFN2 inhibition but upregulated following TUDCA treatment. Furthermore, MFN2 inhibition reduced ER fluorescence intensity and increased the expression of UPR signaling markers (GRP78, XBP1, CHOP, IRE1, and ATF6), indicating increased ER stress. TUDCA administration reversed these effects, restoring MFN2 levels and reducing apoptosis. In conclusion, MFN2 is essential for bovine embryonic development because it regulates ER stress and maintains cell function, with MFN2 deficiency leading to developmental disorders and cell damage. ER stress protectors such as TUDCA can effectively mitigate these negative effects, highlighting a potential strategy for improving in vitro embryo culture efficiency.

Integrative Transcriptomics and Proteomics Analysis Reveals THRSP's Role in Lipid Metabolism.

Background/Objectives: Abnormalities in lipid metabolism and endoplasmic reticulum (ER) stress are strongly associated with the development of a multitude of pathological conditions, including nonalcoholic fatty liver disease (NAFLD), diabetes mellitus, and obesity. Previous studies have indicated a potential connection between thyroid hormone responsive (THRSP) and lipid metabolism and that ER stress may participate in the synthesis of key regulators of adipogenesis. However, the specific mechanisms remain to be investigated. Methods: In this study, we explored the roles of THRSP in lipid metabolism by interfering with THRSP gene expression in mouse mesenchymal stem cells, comparing the effects on adipogenesis between control and interfered groups, and by combining transcriptomic and proteomic analysis. Results: Our results showed that the number of lipid droplets was significantly reduced after interfering with THRSP, and the expression levels of key regulators of adipogenesis, such as LPL, FABP4, PLIN1, and CIDEC, were significantly downregulated. Both transcriptomic and proteomic results showed that the differential genes (proteins) were enriched in the processes of lipolytic regulation, ER stress, cholesterol metabolism, sphingolipid metabolism, PPAR signaling pathway, and glycerophospholipid metabolism. The ER stress marker gene, ATF6, was the most significantly downregulated transcription factor. In addition, RT-qPCR validation indicated that the expression levels of PPAR signaling pathway gene SCD1; key genes of lipid droplet generation including LIPE, DGAT1, and AGPAT2; and ER stress marker gene ATF6 were significantly downregulated. Conclusions: These suggest that THRSP is involved in regulating ER stress and the PPAR signaling pathway, which is closely related to lipid synthesis and metabolism. Interfering with the expression of THRSP may be helpful in ameliorating the occurrence of diseases related to abnormalities in lipid metabolism.

Multiple Myeloma Cells with Increased Proteasomal and ER Stress Are Hypersensitive to ATX-101, an Experimental Peptide Drug Targeting PCNA.

Objectives: To examine the regulatory role of PCNA in MM, we have targeted PCNA with the experimental drug ATX-101 in three commercial cell lines (JJN3, RPMI 1660, AMO) and seven in-house patient-derived cell lines with a more primary cell-like phenotype (TK9, 10, 12, 13, 14, 16 and 18) and measured the systemic molecular effects. Methods: We have used a multi-omics untargeted approach, measuring the gene expression (transcriptomics), a subproteomics approach measuring mainly signalling proteins and proteins in complex with these (signallomics) and quantitative metabolomics. These results are supplemented with traditional analysis, e.g., viability, Western and ELISA analysis. Results: The sensitivity of the cell lines to ATX-101 varied, including between three cell lines derived from the same patient at different times of disease. A trend towards increased sensitivity to ATX-101 during disease progression was detected. Although with different sensitivities, ATX-101 treatment resulted in numerous changes in signalling and metabolite pools in all cell lines. Transcriptomics and signallomics analysis of the TK cell lines revealed that elevated endogenous expression of ribosomal genes, elevated proteasomal and endoplasmic reticulum (ER) stress and low endogenous levels of NAD+ and NADH were associated with ATX-101 hypersensitivity. ATX-101 treatment further enhanced the ER stress, reduced primary metabolism and reduced the levels of the redox pair GSH/GSSG in sensitive cells. Signallome analysis suggested that eleven proteins (TPD52, TNFRS17/BCMA, LILRB4/ILT3, TSG101, ZNRF2, UPF3B, FADS2, C11orf38/SMAP, CGREF1, GAA, COG4) were activated only in the sensitive MM cell lines (TK13, 14 and 16 and JJN3), and not in nine other cancer cell lines or in primary monocytes. These proteins may therefore be biomarkers of cells with activated proteasomal and ER stress even though the gene expression levels of these proteins were not elevated. Interestingly, carfilzomib-resistant cells were at least as sensitive to ATX-101 as the wild-type cells, suggesting both low cross-resistance between ATX-101 and proteasome inhibitors and elevated proteasomal stress in carfilzomib-resistant cells. Conclusions: Our multi-omics approach revealed a vital role of PCNA in regulation of proteasomal and ER stress in MM.

Toosendanin alleviates acute lung injury by reducing pulmonary vascular barrier dysfunction mediated by endoplasmic reticulum stress through mTOR

BackgroundAcute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are severe clinical conditions with limited treatment options. Toosendanin (TSN), a triterpenoid compound with anti-inflammatory effects, has unclear efficacy in ALI. PurposeThis study aimed to evaluate TSN's protective effects on ALI and the related mechanisms. MethodsLipopolysaccharide (LPS)-induced ALI models were developed in vivo and in vitro. Endothelial permeability was measured using Evans Blue dye; lipid reactive oxygen species (ROS) and apoptosis were assessed using flow cytometry. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were determined, and cell viability was measured. mRNA and protein expression were quantified using qRT-PCR and Western blotting. Network pharmacology and surface plasmon resonance were used to identify and validate TSN's targets. ResultsTSN reduced endothelial permeability and LPS-induced ALI. It lowered ROS levels, lipid peroxidation, endoplasmic reticulum (ER) stress, and apoptosis, both in vitro and in vivo. Network pharmacology identified mTOR as a key target of TSN, and surface plasmon resonance analysis confirmed TSN's direct binding to mTOR, underscoring mTOR's role in TSN's protective effects against ALI. Western blotting showed that TSN inhibits mTOR and its phosphorylation. In vitro, the mTOR activator MHY1485 reversed TSN's protective effects, increasing ER stress, apoptosis, and endothelial permeability. In vivo, TSN and rapamycin synergistically protected against ALI. ConclusionThis study is the first to demonstrate that TSN protects against ALI by targeting the mTOR pathway, regulating ER stress and apoptosis and mitigating endothelial damage. These findings suggest a novel approach for ALI treatment and underscore TSN's potential clinical value.

Insulin-degrading enzyme regulates insulin-directed cellular autoimmunity in murine type 1 diabetes.

Type 1 diabetes results from the destruction of pancreatic beta cells by autoreactive T cells. As an autoantigen with extremely high expression in beta cells, insulin triggers and sustains the autoimmune CD4+ and CD8+ T cell responses and islet inflammation. We have previously shown that deficiency for insulin-degrading enzyme (IDE), a ubiquitous cytosolic protease with very high affinity for insulin, induces endoplasmic reticulum (ER) stress and proliferation in islet cells and protects non-obese diabetic mice (NOD) from diabetes. Here we wondered whether IDE deficiency affects autoreactive CD8+ T cell responses to insulin and thereby immune pathogenesis in NOD mice. We find that Ide-/- NOD harbor fewer diabetogenic T cells and reduced numbers of CD8+ T cells recognizing the dominant autoantigen insulin and islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP). Using in vitro digestions and cellular antigen presentation assays, we show that generation of the dominant insulin epitope B15-23 involves both the proteasome and IDE. IDE deficiency attenuates MHC-I presentation of the immunodominant insulin epitope by beta cells to cognate CD8+ T cells. Consequently, Ide-/- islets display reduced susceptibility to autoimmune destruction upon grafting, and to killing by insulin-specific CD8+ T cells. Moreover, Ide-/- mice are partly resistant to disease transfer by CD8+ T cells specific for insulin but not for IGRP. Thus, IDE has a dual role in beta cells, regulating ER stress and proliferation while at the same time promoting insulin-directed autoreactive CD8+ T cell responses.

Impaired inducibility of immune regulatory capacity of peripheral B cells of patients with recurrent pregnancy loss.

The pathogenesis of recurrent pregnancy loss (RPL) is unclear. RPL may have an association with disruption of immune tolerance. The aim of this study is to characterize the inducibility of immune regulatory ability in peripheral naïve B cells of patients with RPL. In this study, blood samples were taken from patients with RPL. B220+ B cells were isolated by flow cytometry cell sorting. The gene profile of B cells was analyzed using RNA sequencing (RNAseq). The results showed that peripheral B220+ B cells of RPL patients had lower expression of IL10 and exacerbated ER stress. The induction of IL10 expression in peripheral B220+ B cells of RPL patients were impaired. High ubiquitination of c-Maf inducing protein (CMIP) was detected in RPL B cells. Exposure to thapsigargin (an ER stress agonist) decreased the amount of CMIP in B cells. The effects of ER stress on reducing CMIP quantity in B cells were mediated by the histone H2B E3 ubiquitin ligase ring finger protein 20 (RNF20). Inhibition of RNF20 or ER stress restored the inducibility of immune regulatory functions of B220+ B cells of RPL patients. In summary, peripheral B cells in patients with RPL show impaired immune regulation capacity, in which exacerbated ER stress plays a crucial role. Regulation of ER stress or inhibition of RNF20 can restore the immune regulatory capacity in the B cells.

Sestrin2 Attenuates Myocardial Endoplasmic Reticulum Stress and Cardiac Dysfunction During Ischemia/Reperfusion Injury.

Sesn2 (Sestrin2) is a stress-induced protein that provides protective effects during myocardial ischemia and reperfusion (I/R) injury, while endoplasmic reticulum (ER) stress may be a pivotal mediator of I/R injury. The goal of this study was to determine whether Sesn2-mTOR (mammalian target of rapamycin) signaling regulates ER stress during myocardial I/R. In vivo cardiac I/R was induced by ligation and subsequent release of the left anterior descending coronary artery in wild-type (WT) and cardiac-specific Sesn2 knockout(Sesn2cKO) mice. At 6 hours and 24 hours after reperfusion, cardiac function was evaluated, and heart samples were collected for analysis. I/R induced cardiac ER stress and upregulated Sesn2 mRNA and protein levels. Inhibiting ER stress with 4-phenylbutyric acid reduced infarct size by 37.5%, improved cardiac systolic function, and mitigated myocardial cell apoptosis post-I/R. Hearts from Sesn2cKO mice displayed increased susceptibility to ER stress during I/R compared with WT. Notably, cardiac mTOR signaling was further increased in Sesn2cKO hearts compared with WT hearts during I/R. In mice with cardiac Sesn2 deficiency, compared with WT, ER lumen was significantly expanded after tunicamycin-induced ER stress, as assessed by transmission electron microscopy. Additionally, pharmacological inhibition of mTOR signaling with rapamycin improved cardiac function after tunicamycin treatment and significantly attenuated the unfolded protein response and apoptosis in WT and Sesn2cKO mice. Sesn2 attenuates cardiac ER stress post-I/R injury via regulation of mTOR signaling. Thus, modulation of the mTOR pathway by Sesn2 could be a critical factor for maintaining cardiac ER homeostasis control during myocardial I/R injury.

The critical role of endoplasmic reticulum stress and the stimulator of interferon genes (STING) pathway in kidney fibrosis

Endoplasmic reticulum (ER) stress is a condition in which the ER is overwhelmed and unable to manage its protein load properly. The precise activation mechanisms and role of ER stress in kidney disease remain unclear. To study this, we performed unbiased transcriptomics analysis to demonstrate ER stress in kidneys of patients with chronic kidney disease and in mouse models of acute and chronic kidney injury (cisplatin and unilateral ureteral obstruction and reanalyzed previously published data on folic acid and mitochondrial transcription factor A(TFAM) knockout mice). Inhibiting the protein kinase RNA-like ER kinase (PERK) arm of ER stress but not activating transcription factor 6 or inositol-requiring enzyme 1, protected mice from kidney fibrosis. The stimulator of interferon genes (STING) was identified as an important upstream activator of ER stress in kidney tubule cells. STING and PERK were found to physically interact, and STING agonists induced PERK activation in kidney tubule cells. Mice with a STING activating mutation presented with ER stress and kidney fibroinflammation. We also generated mice with a tubule specific STING deletion that were resistant to ER stress and kidney fibrosis. Human kidney spatial transcriptomics highlighted a spatial correlation between STING, ER stress and fibrotic gene expression. Thus, our results indicate that STING is an important upstream regulator of PERK and ER stress in tubule cells during kidney fibrosis development.

The role of m5C RNA methylation regulators in the diagnosis and immune microenvironment of osteoarthritis

The 5-methylcytosine (m5C) is a common post-transcriptional RNA methylation modification and is involved in the pathological process of many diseases. However, little is known about the role of m5C in osteoarthritis (OA). OA gene data and the corresponding information were downloaded from the Gene Expression Omnibus database. Based on 36 m5C regulators, we constructed the landscape and diagnostic model for OA. Later, two m5C modification patterns were identified, and functional analyses were performed to evaluate whether these patterns were related to endoplasmic reticulum (ER) stress and mitochondrial autophagy. We further comprehensively analyzed the immune cell infiltration characteristics in different modification patterns in OA. We also established the post-transcriptional regulatory networks and drug-gene networks. Our findings suggested that m5C regulators were differentially expressed between OA and normal samples and could serve as novel biomarkers for the diagnosis of OA. Besides, m5C regulators may be involved in regulating ER stress, mitochondrial autophagy, and immune infiltration in OA. The m5C modification can influence the sensitivity to drugs and the potential post-transcriptional regulatory mechanisms might provide promising targets.

Licochalcone A Ameliorates Cognitive Dysfunction in an Alzheimer's Disease Model by Inhibiting Endoplasmic Reticulum Stress-Mediated Apoptosis.

Endoplasmic reticulum (ER) stress-induced neurodegeneration has been considered an underlying cause of Alzheimer disease (AD). Here, we investigated the beneficial effects of licochalcone A (Lico A), a valuable flavonoid of the root of the Glycyrrhiza species, against cognitive impairment in AD by regulating ER stress. The triple transgenic mouse AD models were used and were administrated 5 or 15mg/kg Lico A. Cognitive deficits, Aβ deposition, ER stress, and neuronal apoptosis were determined using Morris Water Maze test, probe trial, immunofluorescence staining, western blotting, and TUNEL staining. To investigate the mechanisms of how Lico A exerts anti-AD effects, primary hippocampal neurons were isolated from the AD model mice and treated with Lico A, salubrinal, an eIF2α phosphatase inhibitor, ML385, a Nrf2 inhibitor, or LY294002, an inhibitor of PI3K. Pharmacokinetics and toxicity of Lico A (15mg/kg) in AD mice were evaluated. We found that Lico A improved cognitive impairment, decreased Aβ plaques, inhibited ER stress, and reduced neuronal apoptosis in the hippocampus and cortex of AD mice. Treatment with Lico A in primary hippocampal neurons exerted the same effects as it did in vivo. Additionally, cotreatment with ML385 or LY294002 significantly impeded the effects of Lico A against ER stress. Moreover, 15mg/kg Lico A had a good bioavailability and low toxicity in AD mice. Our results demonstrated that Lico A ameliorates ER stress-induced neuronal apoptosis by inhibiting PERK/eIF2α/ATF4/CHOP signaling, suggesting the therapeutic potential of Lico A in AD treatment.

Potential ameliorative effect of Dapagliflozin on systemic inflammation-induced cardiovascular injury via endoplasmic reticulum stress and autophagy pathway.

Dapagliflozin (DPG) is a sodium-glucose cotransporter-2 inhibitor and is used in the treatment of diabetes. In this study, we aimed to investigate the effect of DPG on cardiotoxicity caused by systemic inflammation via endoplasmic reticulum (ER) stress and autophagy. Four groups of thirty-two Wistar Albino rats were created: Control (1ml oral physiological saline for five days and intraperitoneal saline on the 5th day), LPS (1ml oral physiological saline for five days and intraperitoneal 5mg/kg of LPS on the 5th day), LPS + DPG (10mg/kg of DPG orally for five days and 5mg/kg of LPS intraperitoneally on the 5th day), and DPG (10mg/kg of DPG orally for five days and 5mg/kg of SF intraperitoneally on the 5th day). Histopathological and immunohistochemical analyses were performed on heart and aorta tissues. ER stress and autophagy gene markers in heart tissues were evaluated by RT-qPCR. Oxidative stress in heart tissues and serum cardiac enzymes were analyzed by spectrophotometric method. The heart and aortic tissues of the LPS group showed increased expressions of Tumor Necrosis Factor-α (TNF-α) and Caspase-3 (Cas-3), along with mild hyperemia, slight inflammatory cell infiltrations, and myocardial cell damage. The heart tissues also showed genetically increased expressions of include binding immunoglobulin protein (BiP/ GRP78), protein kinase RNA-like ER Kinase (PERK), inositol-requiring enzyme 1 (IRE-1), activating transcription factors 4 (ATF-4), activating transcription factors 4 (ATF6), C/EBP homologous protein (CHOP), and BECLIN 1. Furthermore, Creatine kinase-MB (CK-MB) and Lactate dehydrogenase (LDH) levels in blood tissue significantly increased, according to biochemical analysis. With DPG therapy, all of these findings were reversed. In conclusion, DPG protects against the cardiotoxic effect of systemic inflammation with its antioxidant and anti-inflammatory properties by regulating ER stress and autophagy pathways.

A novel ER stress regulator ARL6IP5 induces reticulophagy to ameliorate the prion burden

ABSTRACT Prion disease is a fatal and infectious neurodegenerative disorder caused by the trans-conformation conversion of PRNP/PrPC to PRNP/PrPSc. Accumulated PRNP/PrPSc-induced ER stress causes chronic unfolded protein response (UPR) activation, which is one of the fundamental steps in prion disease progression. However, the role of various ER-resident proteins in prion-induced ER stress is elusive. This study demonstrated that ARL6IP5 is compensatory upregulated in response to chronically activated UPR in the cellular prion disease model (RML-ScN2a). Furthermore, overexpression of ARL6IP5 overcomes ER stress by lowering the expression of chronically activated UPR pathway proteins. We discovered that ARL6IP5 induces reticulophagy to reduce the PRNP/PrPSc burden by releasing ER stress. Conversely, the knockdown of ARL6IP5 leads to inefficient macroautophagic/autophagic flux and elevated PRNP/PrPSc burden. Our study also uncovered that ARL6IP5-induced reticulophagy depends on Ca2+-mediated AMPK activation and can induce 3 MA-inhibited autophagic flux. The detailed mechanistic study revealed that ARL6IP5-induced reticulophagy involves interaction with soluble reticulophagy receptor CALCOCO1 and lysosomal marker LAMP1, leading to degradation in lysosomes. Here, we delineate the role of ARL6IP5 as a novel ER stress regulator and reticulophagy inducer that can effectively reduce the misfolded PRNP/PrPSc burden. Our research opens up a new avenue of selective autophagy in prion disease and represents a potential therapeutic target. Abbreviations: ARL6IP5: ADP ribosylation factor-like GTPase 6 interacting protein 5; AMPK: adenosine 5’-monophosphate (AMP)-activated protein kinase; CALCOCO1: calcium binding and coiled-coil domain 1; CQ: chloroquine; DAPI: 4’6-diamino-2-phenylindole; ER: endoplasmic reticulum; ERPHS: reticulophagy/ER-phagy sites; KD: knockdown; KD-CON: knockdown control; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3, microtubule-associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; MβCD: methyl beta cyclodextrin; 3 MA: 3-methyladenine; OE: overexpression; OE-CON: empty vector control; PrDs: prion diseases; PRNP/PrPC: cellular prion protein (Kanno blood group); PRNP/PrPSc: infectious scrapie misfolded PRNP; Tm: tunicamycin; UPR: unfolded protein response; UPS: ubiquitin-proteasome system.

Genetic knock-in of EIF2AK3 variants reveals differences in PERK activity in mouse liver and pancreas under endoplasmic reticulum stress

Common single-nucleotide variants (SNVs) of eukaryotic translation initiation factor 2 alpha kinase 3 (EIF2AK3) slightly increase the risk of disorders in the periphery and the central nervous system. EIF2AK3 encodes protein kinase RNA-like endoplasmic reticulum kinase (PERK), a key regulator of ER stress. Three exonic EIF2AK3 SNVs form the PERK-B haplotype, which is present in 28% of the global population. Importantly, the precise impact of these SNVs on PERK activity remains elusive. In this study, we demonstrate that PERK-B SNVs do not alter PERK expression or basal activity in vitro and in the novel triple knock-in mice expressing the exonic PERK-B SNVs in vivo. However, the kinase activity of PERK-B protein is higher than that of PERK-A in a cell-free assay and in mouse liver homogenates. Pancreatic tissue in PERK-B/B mice also exhibit increased susceptibility to apoptosis under acute ER stress. Monocyte-derived macrophages from PERK-B/B mice exhibit higher PERK activity than those from PERK-A/A mice, albeit with minimal functional consequences at acute timepoints. The subtle PERK-B-driven effects observed in liver and pancreas during acute stress implicate PERK as a contributor to disease susceptibility. The novel PERK-B mouse model provides valuable insights into ER stress-induced PERK activity, aiding the understanding of the genetic basis of disorders associated with ER stress.

Methacrylated hyaluronic acid/laponite photosensitive, sustained-release hydrogel loaded with bilobalide for enhancing random flap survival through mitigation of endoplasmic reticulum stress

Random flaps are extensively utilized in plastic surgery due to their flexibility compared to traditional axial vascular system arrangements and their resemblance to injured skin in color, thickness, and texture. Despite these advantages, they are susceptible to ischemia-reperfusion injuries and subsequent necrosis post-transplantation. Bilobalide (BB), a sesquiterpene compound derived from Ginkgo biloba, exhibits notable antioxidant and anti-inflammatory properties and is commonly used to treat ischemiareperfusion injuries. However, its short half-life restricts its sustained efficacy in random flaps. In this study, we synthesized a multi-crosslinked, photosensitive methacryloyl hyaluronic acid(HAMA)/laponite(Lap)/bilobalide (BB) hydrogel. This dualcrosslinked hydrogel demonstrates superior mechanical properties and biocompatibility while providing a stable release of bilobalide. In vitro experiments showed that it significantly reduces edema, promotes angiogenesis, and enhances the survival of random flaps. Further network pharmacology analysis and recovery experiments suggested that the hydrogel's beneficial effects are mediated by the regulation of endoplasmic reticulum stress and specifically identified the regulation of the PERK/TXNIP/NLRP3 signaling pathway as crucial to its anti-inflammatory effects. Therefore, this HAMA/Lap/BB hydrogel promotes the survival of random flaps in rats by alleviating endoplasmic reticulum stress, providing a novel intervention strategy for the treatment of random flaps injuries.