Endoplasmic Reticulum Stress-induced Apoptosis Research Articles

Anticancer Effect of Medicinal Mushroom Extract on Renal Cell Carcinoma: Alternative Therapeutic Implication

Background: To investigate if medicinal Poria mushroom extract (PE) would be a potential therapeutic agent against renal cell carcinoma (RCC) as it has been shown to have anticancer/antitumor activity with few side effects. We examined its anticancer effect on RCC as well as its potential anticancer mechanism in vitro . Methods: Anticancer effect of PE was first assessed by the dose-dependent effects (0 - 200 µg/mL) on human RCC, ACHN cells. Its anticancer mechanism was then explored focusing on specific biochemical and molecular parameters Results: PE concentrations >= 100 µg/mL led to 50-90% reduction in cell viability, accompanied by a significant decrease in hexokinase activity and cellular adenosine triphosphate (ATP) level, implying the inhibition of glycolysis. The key metabolic regulators, adenosine monophosphate (AMP)-activated protein kinase (AMPK), protein kinase B (Akt), and mammalian target of rapamycin (mTOR), were specifically modulated by PE. These results were indicative of subsequent growth cessation and cell death. Cell cycle analysis also revealed a G 1 cell cycle arrest and the upregulation of activating transcription factor 4 (ATF4), C/EBP homologous protein (CHOP), and glucose-regulated protein 78 (GRP78), indicated induction of endoplasmic reticulum (ER) stress. Ultimately, such ER stress led to apoptosis, evidenced by proteolytic activation of caspase-3 (Csp-3) and poly-ADP ribose polymerase (PARP). Conclusions: PE has anticancer effect on ACHN cells, resulting in the significant reduction in cell viability. Its anticancer mechanism appears to be linked to glycolysis inhibition, modulations of metabolic pathways, and ER stress-induced apoptosis. World J Nephrol Urol. 2022;11(1):1-9 doi: https://doi.org/10.14740/wjnu433

Estradiol Inhibits ER Stress-Induced Apoptosis in Chondrocytes and Contributes to a Reduced Osteoarthritic Cartilage Degeneration in Female Mice

Gender differences are a common finding in osteoarthritis (OA). This may result from a differential response of males and females to endoplasmic reticulum (ER) stress in articular chondrocytes. We have previously described that ER stress in cartilage-specific ERp57 KO mice (ERp57 cKO) favors the development of knee OA, since this stress condition cannot be adequately compensated in articular chondrocytes with increasing age leading to the induction of apoptotic cell death and subsequent cartilage degeneration. The aim of this study was to enlighten gender-specific differences in ER stress, apoptosis, and OA development in ERp57 cKO mice. The analyses were extended by in vitro studies on the influence of estradiol in CRISPR/Cas9-generated C28/I2 ERp57 knock out (KO) and WT cells. ER stress was evaluated by immunofluorescence analysis of the ER stress markers calnexin (Cnx) and binding-immunoglobulin protein (BiP), also referred to as glucose-regulating protein 78 (GRP78) in vivo and in vitro. Apoptotic cell death was investigated by a commercially available cell death detection ELISA and TUNEL assay. OA development in mice was analyzed by toluidine blue staining of paraffin-embedded knee cartilage sections and quantified by OARSI-Scoring. Cell culture studies exhibited a reduction of ER stress and ER stress-induced apoptosis in C28/I2 cells in presence of physiological estradiol concentrations. This is consistent with a slower increase in age-related ER stress and a reduced number of apoptotic chondrocytes in female mice compared to male littermates contributing to a reduced osteoarthritic cartilage degeneration in female mice. Taken together, this study demonstrates that the female sex hormone estradiol can reduce ER stress and ER stress-induced apoptosis in articular chondrocytes, thus minimizing critical events favoring osteoarthritic cartilage degeneration. Therefore, the inhibition of ER stress through a modulation of effects induced by female sex hormones appears to be attractive for OA therapy.

Irisin protects cardiomyocytes against hypoxia/reoxygenation injury via attenuating AMPK mediated endoplasmic reticulum stress.

Endoplasmic reticulum (ER) stress plays a central role in myocardial ischemia/reperfusion (I/R) injury. Irisin has been reported to have protective properties in ischemia disease. In this study, we aimed at investigating whether irisin could alleviate myocardial I/R injury by ER stress attenuation. The in vitro model of hypoxia/reoxygenation (H/R) was established, which resembles I/R in vivo. Cell viability and apoptosis were estimated. Expressions of cleaved caspase-3, cytochrome c, GRP78, pAMPK, CHOP, and eIF2α were assessed by western blot. Our results revealed that pre-treatment with irisin significantly decreased cytochrome c release from mitochondria and caspase-3 activation caused by H/R. Irsin also reduced apoptosis and increased cell viability. These effects were abolished by AMPK inhibitor compound C pre-treatment. Also, GRP78 and CHOP expressions were up-regulated in the H/R group compared to the control group; however, irisin attenuated their expression. The pAMPK level was significantly decreased compared to the control, and this effect could be partly reversed by metformin pre-treatment. These results suggest that ER stress is associated with cell viability decreasing and cardiomyocytes apoptosis induced by H/R. Irisin could efficiently protect cardiomyocytes from H/R-injury via attenuating ER stress and ER stress-induced apoptosis.

Cyanidin-3-glucoside improves the barrier function of retinal pigment epithelium cells by attenuating endoplasmic reticulum stress-induced apoptosis

Excessive exposure to blue light from smartphones, computers, and other video equipment causes retinal degeneration. Cyanidin-3-glucoside (C3G) exerts protective effects on retinal cells. However, the mechanism by which C3G enhances the barrier function of retinal pigment epithelial (RPE) cells remains unclear. This study investigated the effects of C3G on blue light-irradiated A2E-containing RPE cells and explored whether or not the endoplasmic reticulum (ER) stress and downstream nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathways are involved in the mechanism. Results showed that C3G (10 and 25 μM) observably increased the viability and inhibited the apoptosis of RPE cells. Furthermore, C3G enhanced the barrier function of RPE cells and upregulated the expression of tight junction proteins. Blue light irradiation triggered ER stress, but C3G significantly suppressed the PERK/eIF2α/ATF4/CHOP pathway and maintained normal ER morphology in RPE cells. C3G also activated the Nrf2 pathway to promote RPE survival, which was independent of ER stress modulating Nrf2 activity. This study suggests that C3G promotes the barrier function of RPE cells by regulating ER stress-induced apoptosis, thereby offering a new approach to preventing retinal diseases. Thus, C3G is a potential functional food ingredient to improve visual health.

Cloning and expressions of chop in loach (Misgurnus anguillicaudatus) and its response to hydrogen peroxide (H2O2) stress.

C/EBP [CCAAT/enhancer-binding protein]-homologous protein gene (chop) which plays an important role in endoplasmic reticulum stress-induced apoptosis was investigated here by RACE and qPCR in an aquaculture animal for the first time. The full-length cDNA sequence of loach (Misgurnus anguillicaudatus) chop was 2533 bp, encoding 266 amino acids. The expression level of loach chop changed during different early life stages, with the highest expression at the 8-cell stage. Among different tissues, loach chop predominantly was expressed in gill, spleen, and gonad. We performed a hydrogen peroxide (H2O2, a common-used disinfectant) stress trial to explore the role of loach chop, with three different concentrations (0 μM, 50 μM, and 100 μM) of H2O2. The 100-μM dose was lethal for half the population but the other concentrations did not result in mortality. The activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPX) in loach gill, liver, and spleen decreased with extended stress time and increased H2O2 concentration. The expression levels of gill chop in loaches from the 100-μM group were significantly higher than those from the other two treatment groups at 12 and 24 h of exposure. atf4 and bax, two proapoptotic genes, were significantly upregulated in gills of loaches from the 100-μM group compared to the other two groups 18 h and 24 h after treatment. bcl2, an antiapoptotic gene, presented an opposite trend. These results indicated a close relationship between H2O2 stress and fish apoptosis with loach chop playing an important role in H2O2 stress response.Supplementary InformationThe online version contains supplementary material available at 10.1007/s10695-022-01067-3.

Tanshinone IIA suppresses endoplasmic reticulum stress-induced apoptosis of high glucose-conditioned vascular endothelial cells by regulating microRNA-133/RAC-1 pathway

Cellular stress caused by abnormal accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) is becoming a possible driver of cardiovascular diseases. The primary aim of our study was to explore Tanshinone IIA (Tan IIA)-induced protection against ER-induced apoptosis of human umbilical vein endothelial cells (HUVECs). HUVECs were treated with high glucose (HG), administrated with Tan IIA (2.5, 5, and 10 μM), and transfected with microRNA-133 (miR-133) mimic or inhibitor. Then cell viability was evaluated by MTT assay, the release of lactate dehydrogenase (LDH) was measured by detection kit, and ER stress was analyzed by Western blot measurement of ER stress-related indicators. Cell apoptosis was observed through flow cytometry and Western blot analysis of apoptosis-related markers. miR-133 and RAC-1 expressions in HUVECs were assessed. For HUVECs, HG inhibited cellular survival, promoted the release of LDH, apoptosis and ER stress, down-regulated miR-133, and up-regulated RAC-1. We demonstrated that Tan IIA reverted the damage of HG to HUVECs in a concentration-dependent manner. miR-133 could negatively regulate RAC-1 expression, and Tan IIA inhibited RAC-1 expression by elevating miR-133, thereby reducing the damage of HG to HUVECs. Tan IIA regulates miR-133–RAC-1 axis to reduce the apoptosis caused by ER stress in HG-induced HUVECs, which could provide new insights for treating cardiovascular diseases.

Nrf2-mediated induction of Cyp2a5 partially protects against reductive endoplasmic reticulum stress in mouse hepatocytes

Cytochrome P450 2a5 (Cyp2a5) is distinct from other P450 enzymes in that it is induced in the endoplasmic reticulum (ER) of mouse hepatocytes in conditions that are injurious to the liver. These conditions cause ER stress eventually resulting in apoptosis if not rectified. We previously showed that mouse hepatic Cyp2a5 is induced during reductive ER stress caused by the intramolecular disulfide form of dithiothreitol, trans-4,5-dihydroxy-1,2-dithiane (DTTox), and that overexpression of Cyp2a5 provides partial protection against apoptosis due to bilirubin (BR), a compound known to cause ER stress. The purpose of this study was to investigate the mechanism of Cyp2a5 gene regulation by DTTox and to determine if Cyp2a5 plays a cytoprotective role during reductive ER stress. Exposure to DTTox (10 mM) and another reductive ER stressor, 2-mercaptoethanol (1 mM), for 48 h markedly increased Cyp2a5 protein levels in primary mouse hepatocytes. In addition, DTTox transactivated Cyp2a5 via a mechanism involving the transcription factor nuclear factor-(erythroid-derived 2)-like 2 (Nrf2). Expression of the BR-conjugating enzyme, UDP glucuronosyl transferase 1A1 (UGT1A1) was also increased after DTTox treatment, however, this was reduced by Cyp2a5 overexpression. Hemin, a porphyrin inducer of Cyp2a5, induced mRNA splicing of X-box binding protein 1 (XBP-1), a transcription factor involved in the ER stress response, however, this was also reduced by Cyp2a5 overexpression. Finally, overexpression of Cyp2a5 partially blocked DTTox-mediated caspase-3 cleavage in Hepa 1–6 cells suggesting a cytoprotective role during ER stress. These findings demonstrate that Nrf2-mediated induction of Cyp2a5 in a reducing ER environment provides partial protection against ER stress-induced apoptosis by decreasing XBP-1 mRNA splicing and caspase-3 cleavage.

Quercetin Alleviates Endoplasmic Reticulum Stress-Induced Apoptosis in Buffalo Ovarian Granulosa Cells.

Simple SummaryGranulosa cells are critical components of the ovary that nurture germ cells and sustain oocyte maturation. Apoptosis of granulosa cells leads to follicular atresia, which in turn leads to female infertility. There are many reasons for the apoptosis of granulosa cells, one of which is apoptosis induced by endoplasmic reticulum stress. We found that quercetin could attenuate the effects of endoplasmic reticulum stress on granulosa cells by the PERK/CHOP signaling pathway. The results provide a novel strategy for inhibiting the apoptosis of granulosa cells.Endoplasmic reticulum (ER) stress plays a crucial role in granulosa cell (GCs) apoptosis, which is the main cause of follicular atresia. Quercetin (QC), a plant-derived flavonoid, has antioxidant, anti-inflammatory, and other biological properties. However, whether QC can alleviate the effects of ER stress on buffalo GCs remains unknown. In this study, we constructed an ER stress model in buffalo GCs by using tunicamycin (TM) and pre-treated with QC to explore the effect of QC on cells under ER stress. Apoptosis was detected by Annexin fluorescein 5 isothiocyanate (V-FITC), and the expressions of mRNA and related proteins involved in ER stress and apoptosis were detected via real-time polymerase chain reaction and Western blot. The results revealed that ER stress can cause apoptosis in GCs, whereas QC pre-treatment can prevent apoptosis caused by ER stress. After pre-treatment with QC, the expression levels of ER stress-related genes and proteins significantly decreased, pro-apoptotic genes were significantly down-regulated, and anti-apoptotic genes were significantly up-regulated. Furthermore, the results of Chop gene overexpression suggested that QC alleviated ER stress via the PERK/CHOP signaling pathway. In this study, we preliminarily elucidated that QC alleviates ER stress-induced apoptosis in buffalo GCs, and the results suggest a novel strategy for delaying follicular atresia by inhibiting GCs apoptosis.

IRhom pseudoproteases regulate ER stress-induced cell death through IP3 receptors and BCL-2

The folding capacity of membrane and secretory proteins in the endoplasmic reticulum (ER) can be challenged by physiological and pathological perturbations, causing ER stress. If unresolved, this leads to cell death. We report a role for iRhom pseudoproteases in controlling apoptosis due to persistent ER stress. Loss of iRhoms causes cells to be resistant to ER stress-induced apoptosis. iRhom1 and iRhom2 interact with IP3 receptors, critical mediators of intracellular Ca2+ signalling, and regulate ER stress-induced transport of Ca2+ into mitochondria, a primary trigger of mitochondrial membrane depolarisation and cell death. iRhoms also bind to the anti-apoptotic regulator BCL-2, attenuating the inhibitory interaction between BCL-2 and IP3 receptors, which promotes ER Ca2+ release. The discovery of the participation of iRhoms in the control of ER stress-induced cell death further extends their potential pathological significance to include diseases dependent on protein misfolding and aggregation.

Unfolded Protein Response Is Activated by Aurora Kinase A in Esophageal Adenocarcinoma.

Simple SummaryEsophageal cancer is the 6th most common cause of cancer-related deaths in 2018 worldwide, with a 5-year survival rate of around 20%. This study reported that esophageal adenocarcinoma cancer cells take advantage of unfolded protein response to survival. Our data using both in vitro cancer cell models and in vivo mice models discovered, for the first time, that Aurora kinase A hijacks pro-survival unfolded protein response in esophageal adenocarcinoma, promoting the survival of cancer cells under reflux-mediated stress conditions.Unfolded protein response (UPR) protects malignant cells from endoplasmic reticulum stress-induced apoptosis. We report that Aurora kinase A (AURKA) promotes cancer cell survival by activating UPR in esophageal adenocarcinoma (EAC). A strong positive correlation between AURKA and binding immunoglobulin protein (BIP) mRNA expression levels was found in EACs. The in vitro assays indicated that AURKA promoted IRE1α protein phosphorylation, activating prosurvival UPR in FLO-1 and OE33 cells. The use of acidic bile salts to mimic reflux conditions in patients induced high AURKA and IRE1α levels. This induction was abrogated by AURKA knockdown in EAC cells. AURKA and p-IRE1α protein colocalization was observed in neoplastic gastroesophageal lesions of the L2-IL1b mouse model of Barrett’s esophageal neoplasia. The combined treatment using AURKA inhibitor and tunicamycin synergistically induced cancer cell death. The use of alisertib for AURKA inhibition in the EAC xenograft model led to a decrease in IRE1α phosphorylation with a significant reduction in tumor growth. These results indicate that AURKA activates UPR, promoting cancer cell survival during ER stress in EAC. Targeting AURKA can significantly reverse prosurvival UPR signaling mechanisms and decrease cancer cell survival, providing a promising approach for the treatment of EAC patients.

Molybdenum and cadmium co-exposure induces endoplasmic reticulum stress-mediated apoptosis by Th1 polarization in Shaoxing duck (Anas platyrhyncha) spleens

Excessive molybdenum (Mo) and cadmium (Cd) are deleterious to animals, but immunotoxicity co-induced by Mo and Cd remains unclear. To ascertain the confederate impacts of Mo and Cd on endoplasmic reticulum (ER) stress-mediated apoptosis by Helper T (Th) cells 1 polarization in the spleen of ducks, we randomly allocated forty 8-day-old Shaoxing ducks (Anas platyrhyncha) into 4 groups and reared them with having different doses of Mo and/or Cd basic diet. At the 16th week of the experiment, serum and spleen tissues were extracted. Data confirmed that Mo and/or Cd strikingly promoted their levels in spleen, caused histological abnormality and trace elements imbalance, and disrupted Th1/Th2 balance to divert toward Th1, then triggered ER stress by increasing three branches PERK/eIF2α/CHOP, IRE1/Caspase-12 and TRAF2/JNK signaling pathways-related genes mRNA and proteins levels, which stimulated apoptosis by elevating Bak-1, Bax, Caspase-9, Caspase-3 mRNA expression, and cleaved-Caspase-9/Caspase-9, cleaved-Caspase-3/Caspase-3 proteins expression as well as apoptosis rate, and decreasing Bcl-xL, Bcl-2 mRNA expression and Bcl-2/Bax ratio. Besides, the variation in combined group was most evident. Briefly, the study indicates that Mo and/or Cd exposure trigger ER stress-induced apoptosis via Th1 polarization in duck spleens, and its mechanism is somehow closely linked with the deposition of Cd and Mo, which may aggravate toxic damage to spleen.

Dysfunctional Endoplasmic Reticulum-Mitochondrion Coupling Is Associated with Endoplasmic Reticulum Stress-Induced Apoptosis and Neurological Deficits in a Rodent Model of Severe Head Injury.

Cellular homeostasis requires critical communications between the endoplasmic reticulum (ER) and mitochondria to maintain the viability of cells. This communication is mediated and maintained by the mitochondria-associated membranes and may be disrupted during acute traumatic brain injury (TBI), leading to structural and functional damage of neurons and supporting cells. To test this hypothesis, we subjected male C57BL/6 mice to severe TBI (sTBI) using a controlled cortical impact device. We analyzed the physical ER-mitochondrion contacts in the perilesional cortex using transmission electron microscopy, Western blot, and immunofluorescence. We specifically measured changes in the production of reactive oxygen species (ROS) in mitochondria, the unfolded protein response (UPR), the neuroinflammatory response, and ER stress-mediated apoptosis in the traumatic injured cerebral tissue. A modified neurological severity score was used to evaluate neurological function in the sTBI mice. We found that sTBI induced significant reorganizations of mitochondria-associated ER membranes (MAMs) in the cerebral cortex within the first 24 h post-injury. This ER-mitochondrion coupling was enhanced, reaching its peak level at 6 h post-sTBI. This enhanced coupling correlated closely with increases in the expression of the Ca2+ regulatory proteins (inositol 1,4,5-trisphosphate receptor type 1 [IP3R1], voltage-dependent anion channel 1 [VDAC1], glucose-regulated protein 75 [GRP75], Sigma 1 receptor [Sigma-1R]), production of ROS, degree of ER stress, levels of UPR, and release of proinflammatory cytokines. Further, the neurological function of sTBI mice was significantly improved by silencing the gene for the ER-mitochondrion tethering factor PACS2, restoring the IP3R1-GRP75-VDAC1 axis of Ca2+ regulation, alleviating mitochondria-derived oxidative stress, suppressing inflammatory response through the PERK/eIF2α/ATF4/CHOP pathway, and inhibiting ER stress and associated apoptosis. These results indicate that dysfunctional ER-mitochondrion coupling might be primarily involved in the neuronal apoptosis and neurological deficits, and modulating the ER-mitochondrion crosstalk might be a novel therapeutic strategy for sTBI.

CFLIP downregulation is an early event required for endoplasmic reticulum stress-induced apoptosis in tumor cells

Protein misfolding or unfolding and the resulting endoplasmic reticulum (ER) stress frequently occur in highly proliferative tumors. How tumor cells escape cell death by apoptosis after chronic ER stress remains poorly understood. We have investigated in both two-dimensional (2D) cultures and multicellular tumor spheroids (MCTSs) the role of caspase-8 inhibitor cFLIP as a regulator of the balance between apoptosis and survival in colon cancer cells undergoing ER stress. We report that downregulation of cFLIP proteins levels is an early event upon treatment of 2D cultures of colon cancer cells with ER stress inducers, preceding TNF-related apoptosis-inducing ligand receptor 2 (TRAIL-R2) upregulation, caspase-8 activation, and apoptosis. Maintaining high cFLIP levels during ER stress by ectopic expression of cFLIP markedly inhibits ER stress-induced caspase-8 activation and apoptosis. Conversely, cFLIP knockdown by RNA interference significantly accelerates caspase-8 activation and apoptosis upon ER stress. Despite activation of the proapoptotic PERK branch of the unfolded protein response (UPR) and upregulation of TRAIL-R2, MCTSs are markedly more resistant to ER stress than 2D cultures of tumor cells. Resistance of MCTSs to ER stress-induced apoptosis correlates with sustained cFLIPL expression. Interestingly, resistance to ER stress-induced apoptosis is abolished in MCTSs generated from cFLIPL knockdown tumor cells. Overall, our results suggest that controlling cFLIP levels in tumors is an adaptive strategy to prevent tumor cell’s demise in the unfavorable conditions of the tumor microenvironment.

MiR-16-5p Suppression Protects Human Cardiomyocytes against Endoplasmic Reticulum and Oxidative Stress-Induced Injury.

Oxidative stress, defined as the excess production of reactive oxygen species (ROS) relative to antioxidant defense, plays a significant role in the development of cardiovascular diseases. Endoplasmic reticulum (ER) stress has emerged as an important source of ROS and its modulation could be cardioprotective. Previously, we demonstrated that miR-16-5p is enriched in the plasma of ischemic dilated cardiomyopathy (ICM) patients and promotes ER stress-induced apoptosis in cardiomyocytes in vitro. Here, we hypothesize that miR-16-5p might contribute to oxidative stress through ER stress induction and that targeting miR-16-5p may exert a cardioprotective role in ER stress-mediated cardiac injury. Analysis of oxidative markers in the plasma of ICM patients demonstrates that oxidative stress is associated with ICM. Moreover, we confirm that miR-16-5p overexpression promotes oxidative stress in AC16 cardiomyoblasts. We also find that, in response to tunicamycin-induced ER stress, miR-16-5p suppression decreases apoptosis, inflammation and cardiac damage via activating the ATF6-mediated cytoprotective pathway. Finally, ATF6 is identified as a direct target gene of miR-16-5p by dual-luciferase reporter assays. Our results indicate that miR-16-5p promotes ER stress and oxidative stress in cardiac cells through regulating ATF6, suggesting that the inhibition of miR-16-5p has potential as a therapeutic approach to protect the heart against ER and oxidative stress-induced injury.

Saponins From Platycodon grandiflorum Reduces Cisplatin-Induced Intestinal Toxicity in Mice through Endoplasmic Reticulum Stress-Activated Apoptosis.

Saponins from the roots of Platycodon grandiflorum, an edible medicinal plant, have shown a wide range of beneficial effects on various biological processes. In this study, an animal model was established by a single intraperitoneal injection of cisplatin (20[Formula: see text]mg/kg) for evaluating the protective effects of saponins from the roots of P. grandiflorum (PGS, 15[Formula: see text]mg/kg and 30[Formula: see text]mg/kg) in mice. The results indicated that PGS treatment for 10 days restored the destroyed intestinal mucosal oxidative system, and the loosened junctions of small intestinal villi was significantly improved. In addition, a significant mitigation of apoptotic effects deteriorated by cisplatin exposure in small intestinal villi was observed by immunohischemical staining. Also, western blot showed that PGS could effectively prevent endoplasmic reticulum (ER) stress-induced apoptosis caused by cisplatin in mice by restoring the activity of PERK (an ER kinase)-eIF2[Formula: see text]-ATF4 signal transduction pathway. Furthermore, molecular docking results of main saponins in PGS suggested a better binding ability with target proteins. In summary, the present work revealed the underlying protective mechanisms of PGS on intestinal injury induced by cisplatin in mice.

Nrf2 loss of function exacerbates endoplasmic reticulum stress-induced apoptosis in TBI mice

Nuclear factor erythroid 2-related factor 2 (Nrf2) plays an important role in neuroprotection and recover. Our studies have showed that endoplasmic reticulum (ER) stress-induced apoptosis aggravates secondary damage following traumatic brain injury (TBI). Whether Nrf2 involved in ER stress and ER stress-mediated apoptosis is not clearly investigated. This present study explored the effect of Nrf2 knockout on ER stress and ER stress-induced apoptosis in TBI mice. A lateral fluid percussion injury (FPI)model of TBI was built based on Nrf2 knockout (Nrf2(−/−)) mice and wild-type (Nrf2(+/+)) mice, and the expressions of marker proteins of ER stress and ER stress-induced apoptosis were checked at 24 h following TBI. We found that Nrf2(−/−) mice presented more severe neurological deficit, brain edema and neuronal cell apoptosis compared with Nrf2(+/+) mice. And, the TBI Nrf2(−/−) mice were significantly increased expression of marker proteins of ER stress and ER stress-induced apoptotic pathway including glucose regulated protein (GRP78), protein kinase RNA-like ER kinase (PERK), inositol requiring enzyme (IRE1), activating transcription factor 6 (ATF6), C/EBP homologous protein (CHOP), caspase-12 and caspase-3, compared with that in WT mice. These results suggest that Nrf2 could ameliorate TBI-induced second brain injury partly through ER stress signal pathway.

CCAAT/Enhancer-Binding Protein Homologous Protein (CHOP) Deficiency Attenuates Heatstroke-Induced Intestinal Injury

The intestine is one of the main target organs involved in the pathological process of heatstroke. CCAAT/enhancer-binding protein homologous protein (CHOP) is involved in endoplasmic reticulum (ER) stress-induced apoptosis. This study aimed to explore the role of CHOP in heatstroke-induced intestinal injury and potential therapy. An in vitro heat stress (HS) model using Caco-2 cells was employed. We observed the role of CHOP in apoptosis-mediated intestinal epithelial cell injury secondary to HS by evaluating cell viability, lactate dehydrogenase release, apoptosis levels, and GRP78, PERK, ATF4, CHOP, Bcl-2, and BAX mRNA and protein expression. To further study the role of CHOP in HS-induced intestinal barrier dysfunction, we assessed transepithelial electrical resistance, paracellular tracer flux, ultrastructure of tight junctions, and protein expression of ZO-1 and occludin. Male wild-type mice and CHOP knockout mice were used for in vivo experiments. We evaluated serum d-lactate and diamine oxidase levels, histopathological changes, intestinal ultrastructure, and ZO-1 and occludin protein expression. HS activated the PERK-CHOP pathway and promoted apoptosis by upregulating BAX and downregulating Bcl-2; these effects were prevented by CHOP silencing. Intestinal epithelial barrier function was disrupted by HS in vitro and in vivo. CHOP silencing prevented intestinal barrier dysfunction in Caco-2 cells, whereas CHOP knockout mice exhibited decreased intestinal mucosal injury. The ER stress inhibitor 4-phenylbutyrate (4-PBA) prevented HS-induced intestinal injury in vitro and in vivo. This study indicated that CHOP deficiency attenuates heatstroke-induced intestinal injury and may contribute to the identification of a novel therapy against heatstroke associated with the ER stress pathway.

Fine particulate matter induces endoplasmic reticulum stress-mediated apoptosis in human SH-SY5Y cells

Exposure to ambient fine particulate matter (PM2.5) may contribute to brain injury, however, the molecular mechanisms have not yet been fully described. In this study, the human SH-SY5Y cells were treated with PM2.5 with different concentrations (0, 25, 100, and 250 μg/mL) for 24 h to investigate the cell apoptosis mediated by endoplasmic reticulum (ER) stress. The ratio of apoptosis, Ca2+ level, biomarkers of ER stress and apoptosis were determined. The results revealed that PM2.5 triggered the increase of apoptosis ratio and cellular Ca2+ levels. Compared with control, the expression of GRP78 and phosphorylation of IER1α and p38 were enhanced significantly in the cells under the conditions of PM2.5 exposure for activating ER stress signals. Besides, the key genes (CHOP/DR5/Caspase8/Caspase12) in ER stress-induced apoptosis signals were up-regulated after the PM2.5 treatment compared to the control. The results suggested PM2.5 induced apoptosis in SH-SY5Y cells by the stimulation of ER stress, which may be the potential mechanism of neurological diseases incurred by PM2.5.

Therapeutic Role of Sirtuins Targeting Unfolded Protein Response, Coagulation, and Inflammation in Hypoxia-Induced Thrombosis.

Thrombosis remains one of the leading causes of morbidity and mortality across the world. Many pathological milieus in the body resulting from multiple risk factors escort thrombosis. Hypoxic condition is one such risk factor that disturbs the integrity of endothelial cells to cause an imbalance between anticoagulant and procoagulant proteins. Hypoxia generates reactive oxygen species (ROS) and triggers inflammatory pathways to augment the coagulation cascade. Hypoxia in cells also activates unfolded protein response (UPR) signaling pathways in the endoplasmic reticulum (ER), which tries to restore ER homeostasis and function. But the sustained UPR linked with inflammation, generation of ROS and apoptosis stimulates the severity of thrombosis in the body. Sirtuins, a group of seven proteins, play a vast role in bringing down inflammation, oxidative and ER stress and apoptosis. As a result, sirtuins might provide a therapeutic approach towards the treatment or prevention of hypoxia-induced thrombosis. Sirtuins modulate hypoxia-inducible factors (HIFs) and counteract ER stress-induced apoptosis by attenuating protein kinase RNA-like endoplasmic reticulum kinase (PERK)/Eukaryotic translation initiation factor 2α (eIF2α) pathway activation. It prevents ER-stress mediated inflammation by targeting X-Box Binding Protein 1 (XBP1) and inhibiting nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κβ) signaling through deacetylation. Sirtuins also obstruct nucleotide-binding domain, leucine-rich-containing family, pyrin domain containing 3 (NLRP3) inflammasome activation to reduce the expression of several pro-inflammatory molecules. It protects cells against oxidative stress by targeting nuclear factor erythroid 2-related factor 2 (Nrf2), glutathione (GSH), forkhead box O3 (FOXO3), superoxide dismutase (SOD), catalase (CAT), peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α), glucose-6-phosphate dehydrogenase (G6PD), phosphoglucomutase-2 (PGAM2), and NF-κB, to name few. This review, thus, discusses the potential role of sirtuins as a new treatment for hypoxia-induced thrombosis that involves an intersection of UPR and inflammatory pathways in its pathological manifestation.

P62/SQSTM1-induced caspase-8 aggresomes are essential for ionizing radiation-mediated apoptosis

The autophagy–lysosome pathway and apoptosis constitute vital determinants of cell fate and engage in a complex interplay in both physiological and pathological conditions. Central to this interplay is the archetypal autophagic cargo adaptor p62/SQSTM1/Sequestosome-1 which mediates both cell survival and endoplasmic reticulum stress-induced apoptosis via aggregation of ubiquitinated caspase-8. Here, we investigated the role of p62-mediated apoptosis in head and neck squamous cell carcinoma (HNSCC), which can be divided into two groups based on human papillomavirus (HPV) infection status. We show that increased autophagic flux and defective apoptosis are associated with radioresistance in HPV(-) HNSCC, whereas HPV(+) HNSCC fail to induce autophagic flux and readily undergo apoptotic cell death upon radiation treatments. The degree of radioresistance and tumor progression of HPV(-) HNSCC respectively correlated with autophagic activity and cytosolic levels of p62. Pharmacological activation of the p62-ZZ domain using small molecule ligands sensitized radioresistant HPV(-) HNSCC cells to ionizing radiation by facilitating p62 self-polymerization and sequestration of cargoes leading to apoptosis. The self-polymerizing activity of p62 was identified as the essential mechanism by which ubiquitinated caspase-8 is sequestered into aggresome-like structures, without which irradiation fails to induce apoptosis in HNSCC. Our results suggest that harnessing p62-dependent sequestration of ubiquitinated caspase-8 provides a novel therapeutic avenue in patients with radioresistant tumors.