Endoplasmic Reticulum Stress-related Factors Research Articles

SERCA2 protects against cisplatin-induced damage of auditory cells: Possible relation with alleviation of ER stress

Aims: SERCA2, one of the P-type pumps encoded by gene ATP2A2, is the only calcium reflux channel of the endoplasmic reticulum (ER) and participates in maintaining calcium homeostasis. The present study was designed to explore SERCA2 expression pattern in auditory hair cells and the possible mechanism underlying the effects of SERCA2 on cisplatin-induced ototoxicity.Main methods: The SERCA2 expression pattern in cochlea hair cells and HEI-OC1 cells was measured by Western blot (WB) and immunofluorescence staining. The apoptosis and its related factors were detected by TUNEL assay and WB. The expression levels of ER stress-related factors, ATF6, PERK, IRE1α, and GRP78, were measured via WB. As for the determination of SERCA2 overexpression and knockdown, plasmids and lentiviral vectors were constructed, respectively.Key findings: We found that SERCA2 was highly expressed in cochlea hair cells and HEI-OC1 cells. Of note, the level of SERCA2 expression in neonatal mice was remarkably higher than that in adult mice. Under the exposure of 30 μM cisplatin, SERCA2 was down-regulated significantly compared with the control group. In addition, cisplatin administration triggered the occurrence of ER stress and apoptosis. Those events were reversed by overexpressing SERCA2. On the contrary, SERCA2 knockdown could aggravate the above processes.Significance: The findings from the present study disclose, for the first time, that SERCA2 is abundantly expressed in cochlea hair cells, and the suppression of SERCA2 caused by cisplatin could trigger ER homeostasis disruption, thereby implying that SERCA2 might be a promising target to prevent cisplatin-induced cytotoxicity of hair cells.

ShenQiWan ameliorates renal injury in type 2 diabetic mice by modulating mitochondrial fusion and endoplasmic reticulum stress.

Background: ShenQiWan is commonly used in traditional Chinese medicine for the treatment of diabetic nephropathy, which is closely related to mitochondrial fusion and endoplasmic reticulum stress. This study aimed to investigate the intervention effect and molecular mechanisms of ShenQiWan on renal injury in KKAy mice. Methods: C57BL/6J mice (11weeks old) were fed a regular diet upon arrival, while KKAy mice (11weeks old) were fed a high-fat diet upon arrival. At 12weeks of age, KKAy mice with random blood glucose ≥13.9mmol/L were identified as diabetic mice and randomly divided into the model group (n = 30) and the treatment group (n = 30), while C57BL/6J mice of 12weeks old (n = 30) served as the control group. The treatment group received daily aqueous decoction of ShenQiWan (13.5g/kg), while the control group and model group received daily equal amounts of saline from 12weeks old to 24weeks old. The general status of mice was observed regularly, and fasting blood glucose and 24-hour urine microalbumin were measured. Ten mice were euthanized in each group at the age of 16, 20, and 24weeks, serum samples were used for biochemical indexes and kidney tissues were used for morphological studies. GRP78, OPA1, MFN1, MFN2 mRNA and protein expression were detected by Real-time PCR, immunohistochemistry and Western blot. Results: The mice in the model group exhibited symptoms of lethargy, slow movement, obesity, polyuria and proteinuria. Morphological observation revealed pathological changes, including thickening of the glomerular basement membrane and interstitial fibrosis. After treatment with ShenQiWan, the fasting blood glucose level of KKAy mice was significantly reduced, urinary albuminuria was decreased, serum biochemical indexes were improved, renal tissue pathological changes were significantly alleviated. The results also showed a significant reduction in the expression of endoplasmic reticulum stress-related factor GRP78 and an increase in the expression of mitochondrial fusion-related factors OPA1, MFN1 and MFN2 after treatment with ShenQiWan. Conclusion: ShenQiWan can protect diabetic mice from renal damage by modulating mitochondrial fusion and alleviating endoplasmic reticulum stress, exerting its protective effects.

KDM1A represses the apoptosis in osteosarcoma cells via demethylating Bcl-2/c-Myc

IntroductionBackground: Considering the poorly understood mechanism of lysine demethylase 1A (KDM1A) in osteosarcoma (OS), we here commence our investigation to fill the blank.Material and methodsMethods: Following the transfection as appropriate, 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) and flow cytometry assays were used to determine the viability and apoptosis of OS cells MG-63, in which the generation of reactive oxygen species (ROS) and the binding between KDM1A and Bcl-2/ cellular Myc (c-Myc) were separately confirmed via DCF-DA method and chromatin immunoprecipitation-PCR. Reverse-transcription quantitative PCR and western blot were finally introduced to quantify the levels of KDM1A/Bcl-2/c-Myc and endoplasmic reticulum (ER) stress-related factors.ResultsResults: Overexpressed KDM1A enhanced the viability (48 hours) yet repressed the apoptosis and ROS generation, with the downregulation on ER stress-related factors (C/EBP homologous protein [CHOP]; proline-rich extensin-like receptor kinase (PERK) and activating transcription factor 4 [ATF4]) yet the elevation of Bcl-2/c-Myc, while its depletion exerted contrary effects. More importantly, KDM1A could act as the demethylase of Bcl-2/c-Myc, as reflected by the results that the depletion of KDM1A decreased the enrichment of Bcl-2/c-Myc promoter using the antibody against KDM1A yet increased the enrichment by the antibody targeting H3K9me2. Bcl-2/c-Myc silencing, conversely, promoted the ROS generation and apoptosis, elevated the levels of ER stress-related factors and abolished the effects of KDM1A on OS cells.ConclusionsConclusion: KDM1A exerts a repressive effect on the apoptosis of OS cells MG-63 by inhibiting the ROS generation and ER stress via demethylation of Bcl-2 and c-Myc.

Effects of Different Spectrum of LEDs on Retinal Degeneration Through Regulating Endoplasmic Reticulum Stress.

To evaluate the impact of full-spectrum light-emitting diodes (LEDs) on albino guinea pigs' retina and investigate the roles of short-wavelength opsin (S-opsin) and endoplasmic reticulum (ER) stress in light-induced retinal degeneration (LIRD). Three-week-old albino guinea pigs (n = 30) were distributed into five groups under 12/12 light/dark cycles with indoor natural light (NC; 300-500 lux, n = 6), full-spectrum LEDs (FL; 300 lux, n = 6; 3000 lux, n = 6), and commercial cold-white LEDs (CL; 300 lux, n = 6; 3000 lux, n = 6) and raised for 28 days. Hematoxylin and eosin staining and transmission electron microscopy evaluated the morphological changes of retinas. The immunofluorescence and real-time quantitative polymerase chain reaction (RT-qPCR) measured the expression and content of S-opsin and ER stress-related genes and proteins. We found that albino guinea pigs exposed to FL at either 300 lux or 3000 lux developed less severe retinal morphological damage than animals exposed to the CL light, which emerged as a significant characteristic of LIRD. Meanwhile, the damage on the ventral retina was more serious, mainly due to its ability to absorb the blue light in the LEDs more easily. Compared to the FL-exposed groups, the CL light increased the aggregation of S-opsin and the expression of ER stress-related factors. Commercial cold-white LEDs can induce ER stress and unfolded protein response in LIRD, and full-spectrum LED attenuates LIRD by regulating ER stress in albino guinea pig retinas in vivo. Full-spectrum LEDs offer specific eye protection and eye adaptability that can well replace commercial cold-white LEDs in both clinical practice and research. It should be further developed for lighting used in health care facilities.

Porcine Epidemic Diarrhea Virus and Its nsp14 Suppress ER Stress Induced GRP78.

Porcine epidemic diarrhea virus (PEDV), a member of the α-coronavirus genus, can cause vomiting, diarrhea, and dehydration in piglets. Neonatal piglets infected with PEDV have a mortality rate as high as 100%. PEDV has caused substantial economic losses to the pork industry. Endoplasmic reticulum (ER) stress, which can alleviate the accumulation of unfolded or misfolded proteins in ER, involves in coronavirus infection. Previous studies have indicated that ER stress could inhibit the replication of human coronaviruses, and some human coronaviruses in turn could suppress ER stress-related factors. In this study, we demonstrated that PEDV could interact with ER stress. We determined that ER stress could potently inhibit the replication of GⅠ, GⅡ-a, and GⅡ-b PEDV strains. Moreover, we found that these PEDV strains can dampen the expression of the 78 kDa glucose-regulated protein (GRP78), an ER stress marker, while GRP78 overexpression showed antiviral activity against PEDV. Among different PEDV proteins, PEDV non-structural protein 14 (nsp14) was revealed to play an essential role in the inhibition of GRP78 by PEDV, and its guanine-N7-methyltransferase domain is necessary for this role. Further studies show that both PEDV and its nsp14 negatively regulated host translation, which could account for their inhibitory effects against GRP78. In addition, we found that PEDV nsp14 could inhibit the activity of GRP78 promotor, helping suppress GRP78 transcription. Our results reveal that PEDV possesses the potential to antagonize ER stress, and suggest that ER stress and PEDV nsp14 could be the targets for developing anti-PEDV drugs.

Genomic and clinical features of endoplasmic reticulum stress factor in digestive system pan-cancer studies.

Digestive system pan-cancer is one of the lethal malignant tumors, which have the propensity for poor prognosis and difficult treatment. Endoplasmic reticulum (ER) stress has served as a pivotal role in the progression of the tumor, while the implication of ER stress on digestive system pan-cancers still needs elucidation, especially from the perspective of clinical outcome and that of genomic features. First, Among the ER STRESS factors from the REACTOME_UNFOLDED_PROTEIN_RESPONSE_UPR (113 genes) and HALLMARK_UNFOLDED_PROTEIN_RESPONSE (92 genes) terms, 153 ER STRESS regulators were identified after removing replicates. The somatic mutation data and copy number variation data of gastrointestinal pan-cancer were downloaded from The Cancer Genome Atlas (TCGA) database. Then, we explored the clinical outcome and genetic mutation of ER stress-related differentially expressed genes (DEGs) by multiple bioinformatics analysis. Subsequently, we analyzed the Spearman correlation between the drug sensitivity of 179 gastrointestinal anticancer drugs and the transcriptional expression of 153 ER stress factors in 769 cancer cell lines of the GDSC2 cohort. Next, ssGSEA method was used to quantify the immune cell infiltration scores in the tumor microenvironment, and Spearman correlation was used to calculate the correlation between ER stress scores and immune cell infiltration. Finally, we analyzed the cellular origin of ER stress factor dysregulation. We analyzed the genomic changes and clinical outcomes of ER stress factors in different tumors of gastrointestinal pan-cancer. Endoplasmic reticulum stress factor (ER) in digestive tract tumors showed high SNV mutation frequency, less methylation dysregulation and was associated with multiple oncogenic pathways. Endoplasmic reticulum stress factor (ER) is a risk factor for many cancers, but the effect on overall survival in rectal adenocarcinoma is opposite to that in other gastrointestinal tumors. And ER stress factors are highly correlated with drugs that target important pathways. Based on the clinical prognosis and genomic analysis of ER stress-related factors in patients with gastrointestinal pan-cancer, this study provides a new direction for further research on gastrointestinal pan-cancer.

Characterization of a lipid droplet and endoplasmic reticulum stress related gene risk signature to evaluate the clinical and biological value in hepatocellular carcinoma

IntroductionLipid metabolism and endoplasmic reticulum (ER) stress play an important role in the progression and metastasis of hepatocellular carcinoma (HCC). We aimed to establish lipid droplet (LD)-associated and ER stress-related gene risk signature as prognostic indicators.Materials and methodsLiterature searches for LD-associated proteins was screened and validated in The Cancer Genome Atlas (TCGA) and International Cancer Genome Collaboratory (ICGC) databases. A total of 371 samples were enrolled from the TCGA RNA-seq dataset (training cohort) and 240 samples from IGGC RNA-seq dataset (validation cohort). A 10-gene risk signature was established by the last absolute shrinkage and selection operator (LASSO) regression analysis. The prognostic value of the risk signature was evaluated by Cox regression, Kaplan–Meier and ROC Curve analyses. Biological features associated with LD and ER stress-related factors were explored by functional analysis and in vitro experiment.ResultsBased on the medical literatures, 124 lipid droplet-associated proteins were retrieved, and three genes failed to establish a valid prognostic model. ER stress was considered as an important component by functional analysis. A 10-gene risk signature compared the clinicopathology characteristics, immunosuppressive events and a nomogram in HCC patients.ConclusionLD-associated and ER stress-related gene risk signatures highlighted poor prognosis for clinicopathological features, positively correlate with macrophages and T cell immunoglobulin and mucin-3 (TIM-3) expression in the tumor microenvironment, and might act as independent prognostic factors.

Lycium barbarum L. and Salvia miltiorrhiza Bunge protect retinal pigment epithelial cells through endoplasmic reticulum stress

Ethnopharmacological relevanceLycium barbarum L. and Salvia miltiorrhiza Bunge (Gouqi and Danshen, LS) are traditional herbs for the treatment of retinal degeneration in China. LS have been integrated into pharmacopoeia and health care system of many countries around the world. However, the mechanisms by which LS protect retina are not fully clarified. Aim of the studyWe aimed at exploration of the effect of LS on retinal pigment epithelium (RPE) cells apoptosis as well as the endoplasmic reticulum (ER) stress mechanisms. Material and methodsARPE-19 cells were exposed to tunicamycin to induce ER stress, followed by LS treatment for 24 h. The cell morphology was photographed using the Incucyte S3 instrument, and the potential cytotoxic effect and viability were evaluated by CCK-8 assays. The Annexin V-FITC/PI staining and TUNEL assay were conducted to detect cells apoptotic. Western blot and digital PCR were used to detected related protein and gene expression. ResultsThe ARPE-19 cells are increased in number and aligned after treating with LS. 1 mg/ml is the LS high dose group dose and treatment with LS increased cell vitality. LS significantly inhibit ARPE-19 cells apoptosis. Moreover, LS were markedly decreased the expression levels of ER stress-related factors in the ARPE-19 cells. ConclusionsThis study reveals that LS relieve ARPE-19 cells apoptosis by inhibiting ER stress, and here we can speculate that LS have a certain protective effect on retina.

Effect of enalapril maleate-folic acid tablets on inflammatory response and myocardial endoplasmic reticulum stress-related factors in hypertensive rats

Purpose: To determine the effect of enalapril maleate folate on inflammatory reaction and myocardial endoplasmic reticulum stress-related factors in hypertensive rats.Methods: Eighty (80) hypertensive rats with SBP > 140 mmHg were equally assigned to control and study groups. Rats in control group were given normal saline by gavage, while the observation group was given enalapril maleate powder (10 mg/kg/day). After 4 weeks of treatment, 2 mL of inferior vena cava blood was collected from each of the two rat groups. The level of homocysteine (Hcy) was determined with Hcy assay kit, while C-reactive protein (CRP) levels in patients were determined by latex immunoturbidimetry. Serum FBG was assessed using automatic biochemical analyzer. The expression levels of GRP78, CRP94, chop and caspase-12 in aortic smooth muscle cells were assayed immunohistochemically.Results: Central arterial pressure (MAP), aortic media thickness, lvwi, HWI FBG and CRP were significantly higher in the study rats, while Hcy level was lower, than in controls (p < 0.05). There were significantly lower levels of glucose regulatory protein 78 (GRP78), CRP94, CHOP and caspase-12 in study rats than in control rats (p < 0.05).Conclusion: Enalapril maleate-folate slows down inflammatory reaction by reducing the levels of Hcy, CRP and FBG. It inhibits the expressions of GRP78, CRP94, CHOP and caspase-12 in myocardium, reduces damage to myocardial cells, and alleviates or reverses left ventricular hypertrophy in rats with high blood pressure. This provides new insight into the research and development of other drugs.

Effect of acupuncture on endoplasmic reticulum stress-related factors in hippocampus of post-traumatic stress disorder rats

To observe the effect of acupuncture on endoplasmic reticulum stress-related molecules glucose regulated protein 78 kD (GRP78), C/EBP homologous protein (CHOP), cysteinyl aspartate specific proteinase-12 (Caspase-12) and cysteinyl aspartate specific proteinase-3 (Caspase-3)in the hippocampus of rats with post-traumatic stress disorder, so as to explore the possible mechanism of acupuncture in treating post-traumatic stress disorder (PTSD). Twenty-eight SD rats were randomly divided into normal control, model, acupuncture and sertraline groups, with 7 rats in each group. The PTSD rat model was established by single prolonged stress. After modeling, acupuncture was applied to "Baihui" (GV20) and "Dazhui" (GV14) for rats of the acupuncture group for 10 min, once a day for 7 days. Sertraline (10 mg/kg) was given by gavage to rats of the sertraline group daily for 7 days. Rats' behavior was assessed by open field test and novelty-suppressed test. The mRNA expression levels of GRP78 and CHOP in the hippocampus were detected by real-time PCR. The expression le-vels of Caspase-12 and Caspase-3 in the hippocampus were detected by Western blot. Compared with the normal control group, the rearing and crossing times were decreased (P<0.05), the time remaining in the central zone and the total distance of movement were significantly reduced (P<0.01, P<0.05), the time of entering the central area for the first time was significantly increased (P<0.01), the latency of the novelty-suppressed feeding was significantly increased (P<0.05) in the model group, meanwhile the expression level of GRP78 and CHOP mRNAs, Caspase-12 and Caspase-3 proteins in the hippocampus were increased (P<0.05, P<0.01). In comparison with the model group, the crossing times, the time remaining in the central zone and total distance of movement were increased significantly (P<0.05, P<0.01), while the time of entering the central area for the first time, the expression levels of GRP78 and CHOP mRNAs, and Caspase-12 protein in the hippocampus were obviously decreased (P<0.05, P<0.01) in the acupuncture and sertraline groups. In addition, the rearing times were increased significantly (P<0.05), the latency of the novelty-suppressed feeding and the expression of Caspase-3 were decreased significantly (P<0.05) in the sertraline group than in the model group. Acupuncture can significantly down-regulate the expression of endoplasmic reticulum stress-related molecules GRP78, CHOP and Caspase-12 in PTSD rats, which may be one of the mechanisms of acupuncture in treating PTSD.

Comparison of the Drug-Induced Efficacies between Omidenepag Isopropyl, an EP2 Agonist and PGF2α toward TGF-β2-Modulated Human Trabecular Meshwork (HTM) Cells.

To compare the drug-induced efficacies between omidenepag (OMD), an EP2 agonist, and prostaglandin F2α (PGF2α) on glaucomatous trabecular meshwork (TM) cells, two- and three-dimensional (2D and 3D) cultures of TGF-β2-modulated human trabecular meshwork (HTM) cells were used. The following analyses were performed: (1) transendothelial electrical resistance (TEER) and FITC-dextran permeability measurements (2D), (2) the size and stiffness of the 3D spheroids, and (3) the expression (both 2D and 3D) by several extracellular matrix (ECM) molecules including collagen (COL) 1, 4 and 6, and fibronectin (FN), and α smooth muscle actin (αSMA), tight junction (TJ)-related molecules, claudin11 (Cldn11) and ZO1, the tissue inhibitor of metalloproteinase (TIMP) 1–4, matrix metalloproteinase (MMP) 2, 9 and 14, connective tissue growth factor (CTGF), and several endoplasmic reticulum (ER) stress-related factors. TGF-β2 significantly increased the TEER values and decreased FITC-dextran permeability, respectively, in the 2D HTM monolayers, and induced the formation of downsized and stiffer 3D HTM spheroids. TGF-β2-induced changes in TEER levels and FITC-dextran permeability were remarkably inhibited by PGF2α. PGF2α induced increases in the sizes and stiffness of the TGF-β2-treated 3D spheroids, but OMD enhanced only spheroid size. Upon exposure to TGF-β2, the expression of most of the molecules that were evaluated were significantly up-regulated, except some of ER stress-related factors were down-regulated. TJ-related molecules or ER stress-related factors were significantly up-regulated (2D) or down-regulated (3D), and down-regulated (2D) by PGF2α and OMD, while both drugs altered the expression of some of the other genes in the 3D spheroids in a different manner. The findings presented herein suggest that PGF2α and OMD differently modulate the permeability of the TGFβ2-modulated 2D monolayers and the physical properties of the 3D HTM spheroids.

High glucose inhibits the osteogenic differentiation of periodontal ligament stem cells in periodontitis by activating endoplasmic reticulum stress.

BackgroundPeriodontitis is a highly prevalent dental disease which is associated with diabetes and is challenging to cure in diabetic patients. However, the mechanism of comorbid diabetes and periodontitis is still unclear. This study aimed to uncover the role of endoplasmic reticulum (ER) stress in high glucose-associated periodontitis.MethodsPeriodontal tissues were obtained from diabetic patients with periodontitis, periodontitis patients without systemic disease, and healthy teeth. The expressions of ER stress-related factors GRP78, ATF6, PERK and XBP1 were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and immumohistochemical staining. Periodontal ligament stem cells (PDLSCs) from three states of periodontal tissues were isolated and cultured as diabetic PDLSCs (dPDLSCs), inflamed PDLSCs (iPDLSCs) and healthy PDLSCs (hPDLSCs), and the cell stemness was assayed. Different concentrations (8, 11, and 25 mmol/L) of D-glucose were used on hPDLSCs to simulate high glucose microenvironment. The changes of osteogenic ability of PDLSCs were observed, and the expressions of ER stress-related factors in different time point (6, 12, 24, and 72 h) were detected. Finally, GRP78 shRNA lentivirus was used to block ER stress on PDLSCs in the 25 mmol/L D-glucose microenvironment, and the osteogenic ability of PDLSCs was observed.ResultsThe results showed that the expressions of GRP78, ATF6, PERK, and XBP1 were highest in the diabetic periodontitis group and lowest in the healthy periodontal tissue group (P<0.05). The clone formation, osteogenic and lipogenic differentiation abilities were lowest in dPDLSCs and highest in hPDLSCs. With the increase of glucose concentration, the osteogensis ability of PDLSCs decreased. After 6 hours of stimulation with D-glucose 25 mmol/L, the ER stress pathways in PDLSCs were effectively activated, and the peak value was reached at 12 hours. The decrease in the osteogensis ability of PDLSCs in a high glucose microenvironment reversed when ER stress was blocked.ConclusionsThe osteogenic differentiation ability of PDLSCs cells is inhibited in a high glucose microenvironment, and this effect is realized by ER stress activation. Blocking ER stress can partially restore the reduced osteogenic ability of PDLSCs. These results suggest that high glucose inhibits the osteogenic differentiation ability of PDLSCs by activating ER stress, which ultimately exacerbates periodontitis.

Effects of exogenous vitamin D on endoplasmic reticulum stress-related factors in the vitamin D deficiency rats with cerebral ischemia

We observed the potential neuroprotective effect of exogenous vitamin D on cerebral ischemia in vitamin D deficiency rats and explored its mechanism. Sixty Sprague Dawley rats were randomly divided into three groups including control (CON) group, vitamin D deficiency (VDD) group and vitamin D deficiency combined with exogenous vitamin D (VDD + VD) group, with 20 rats in each group. The rats in the CON group were fed with normal diet, while the rats in the VDD and VDD + VD groups with vitamin D-deficient diet for 8 weeks, and then underwent middle cerebral artery occlusion (MCAO). Six hours later, exogenous 25-hydroxyvitaminD (25-VD3) was intravenously injected into the rats of the VDD + VD group every 24 h for 4 times. Motor behaviors were significantly better and 25-VD3 level in the left brain tissue was significantly higher in the CON group than in the VDD and VDD + VD groups 102 h after MCAO (all P < 0.05). Although the motor behaviors, infarct volume and 25-VD3 level in brain tissue showed a good tendency in the VDD + VD group as compared with VDD group, the two groups were not significantly different (all P > 0.05). GPR78 was significantly higher, but apoptosis and CHOP significantly lower in the VDD + VD group than in the VDD group (all P < 0.05). The aggravation of cerebral ischemia caused by vitamin D deficiency is possibly due to up-regulating GRP78 and down-regulating CHOP in brain tissue.

Inhibition of ASIC1a-Mediated ERS Improves the Activation of HSCs and Copper Transport Under Copper Load.

Hepatolenticular degeneration (HLD) is an autosomal recessive genetic disease caused by the toxic accumulation of copper in the liver. Excessive copper will disrupt the redox balance in cells and tissues, causing ischemia, hypoxia, and inflammation. Acid-sensitive ion channel 1a is a cationic channel activated by extracellular acid and allowing Ca2+ and Na+ to flow into cells. Its expression appears in inflammation, arthritis, fibrotic tissue, and damaged environment, but its role in hepatolenticular degeneration has not been studied. This study established a Wistar rat model of high copper accumulation and used CuSO4 to induce the activation of HSC-T6 in an in vitro experiment. In vivo, Wistar rats were examined to determine the serum copper concentration, serum ALT and AST activities, and liver copper accumulation, and liver tissue HE staining and immunohistochemical analyses were conducted. The expression of ASIC1a, α-SMA, Collagen-Ι, GRP78, XBP1, ATP7B, and CCS were detected. Besides, immunofluorescence technology can detect the expression of the phosphorylated protein in vitro. It is suggested that ASIC1a is involved in the quality control of the endoplasmic reticulum, which degrades mutant ATP7B and increases the accumulation of copper. After blocking or silencing the expression of ASIC1a, ELISA can detect the level of inflammatory factors, the expression of endoplasmic reticulum stress-related factors, and ATP7B was improved in a higher copper environment reduction of copper deposition was observed in liver Timm’s staining. Collectively, we conclude that ASIC1a is involved in the HSC activation induced by copper accumulation and promotes the occurrence of hepatolenticular fibrosis.

Inhibition of CACNA1H can alleviate endoplasmic reticulum stress and reduce myocardial cell apoptosis caused by myocardial infarction.

In recent years, coronary heart disease (CHD) has become a disease that cannot be ignored by residents of our country, because CHD will not only endanger people's quality of life, but also threaten their lives. Therefore, this research mainly explores the correlation between myocardial infarction (MI) with endoplasmic reticulum (ER) stress and apoptosis. First, we constructed a model of myocardial ischemia and hypoxia (I/H) in vivo and in vitro, and examined the change of CACNA1H expression. At the same time, in order to research the role of CACNA1H, we chose CACNA1H-specific inhibitor ABT-639 to next research and detect changes in heart injury by detecting changes in creatine kinase (CK) content and lactate dehydrogenase (LDH) activity. Next, we used TUNEL staining and immunofluorescence staining to detect changes in apoptosis and ER stress, and analyzed changes in ER stress and apoptotic pathway expression by Western blotting and quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). At 28 days after MI, the cardiac function of the mice was significantly reduced, the myocardial cell apoptosis rate was dramatically increased, and CACNA1H expression was dramatically increased in vivo and in vitro. In addition, we treated the model group with the ABT-639, and found that ABT-639 can partially protect myocardial function and relieve myocardial cell apoptosis. At the same time, ABT-639 may reduce H9c2 injury after I/H by reducing the degree of ER stress, because we found that the use of ABT-639 can dramatically reduce ER stress-related factors expression, and can inhibit the expression of apoptosis-related factors Caspase-3 and Caspase-9. The CACNA1H inhibitor ABT-639 can alleviate myocardial cell apoptosis caused by MI by reducing the ER stress response.

Endoplasmic reticulum stress related factor IRE1α regulates TXNIP/NLRP3-mediated pyroptosis in diabetic nephropathy

The nod-like receptor protein-3 (NLRP3)-mediated pyroptosis is involved in kidney diseases. Thioredoxin interacting protein (TXNIP) directly interacts with NLRP3. This study aimed to probe the mechanism of TXNIP and NLRP3 pathway in diabetic nephropathy (DN). Marker detection and histological staining indicated that in DN rats, the renal function was destroyed, and the TXNIP/NLRP3 axis was activated to induce inflammatory generation and pyroptosis. The protein levels of TXNIP, NLRP3 inflammatory components and endoplasmic reticulum stress (ERS)-related factors (ATF4, CHOP and IRE1α) were measured. DN rats were injected with LV-TXNIP-shRNA or IRE1α RNase specific inhibitor (STF-083010) to examine ERS- and pyroptosis-related proteins, and renal injury. Silencing TXNIP inhibited the NLRP3 axis and reduced renal damage in DN rats. ERS was activated in DN rats, and miR-200a expression was degraded by IRE1α. miR-200a bound to TXNIP. NRK-52E cells were induced by high glucose (HG) to simulate DN in vitro. The damage and pyroptosis of NRK-52E cells were analyzed. After inhibiting IRE1α, miR-200a expression increased and TXNIP expression decreased. miR-200a inhibition in HG-induced NRK-52E cells partially reversed the reduced pyroptosis by STF-083010. Overall, IRE1α upregulates miR-200a degradation in DN rats, and stimulates the TXINP/NLRP3 pathway-mediated pyroptosis and renal damage.

Seleno-l-methionine suppresses copper-enhanced zinc-induced neuronal cell death via induction of glutathione peroxidase.

Excessive zinc ion (Zn2+) release is induced in pathological situations and causes neuronal cell death. Previously, we have reported that copper ions (Cu2+) markedly exacerbated Zn2+-induced neuronal cell death by potentiating oxidative stress, the endoplasmic reticulum (ER) stress response, and the activation of the c-Jun amino-terminal kinase (JNK) signaling pathway. In contrast, selenium (Se), an essential trace element, and amino acids containing selenium (such as seleno-l-methionine) have been reported to inhibit stress-induced neuronal cell death and oxidative stress. Thus, we investigated the effect of seleno-l-methionine on Cu2+/Zn2+-induced neuronal cell death in GT1-7 cells. Seleno-l-methionine treatment clearly restored the Cu2+/Zn2+-induced decrease in the viable cell number and attenuated the Cu2+/Zn2+-induced cytotoxicity. Accordingly, the levels of ER stress-related factors (especially, CHOP and GADD34) and of phosphorylated JNK increased upon CuCl2 and ZnCl2 co-treatment, whereas pre-treatment with seleno-l-methionine significantly suppressed these upregulations. Analysis of reactive oxygen species (ROS) as upstream factors of these pathways revealed that Cu2+/Zn2+-induced ROS production was clearly suppressed by seleno-l-methionine treatment. Finally, we found that seleno-l-methionine induced the antioxidative protein, glutathione peroxidase. Taken together, our findings suggest that seleno-l-methionine suppresses Cu2+/Zn2+-induced neuronal cell death and oxidative stress via induction of glutathione peroxidase. Thus, we think that seleno-l-methionine may help prevent refractory neurological diseases.

Effects of curcumin on protein expression of glucose regulated protein 78 and caspase-12 of myocardial endoplasmic reticulum stress related factors in type 2 diabetes rats

To study the effect of curcumin on the expression of glucose regulated protein 78 kD(GRP78) and cysteinyl aspartate specific proteinase-12(caspase-12) of myocardial endoplasmic reticulum stress related factors in type 2 diabetes rats. Type 2 diabetes rats model was established by high-fat drink feeding and one-time intraperitoneal injecting streptozotocin(35 mg/kg). After model rats were built, rats was randomly divided into diabetic model group and low dose of curcumin group(200 mg/kg), high dose of curcumin group(400 mg/kg) and captopril group(60 mg/kg) with 10 rats in each group. The rats in each group were ig administered with corresponding drugs once a day. Continuous administration for 12 w. The levels of fasting blood glucose(FBG) and lactate dehydrogenase(LDH), electrocardiogram and heart weight index(HWI) were measured respectively. The myocardial pathological changes were observed by HE staining. The levels of collagen fiber expression in myocardial tissue were performed by Masson staining. The protein expression levels of GRP78 and caspase-12 in myocardium of rats were observed by immunohistochemistry. The result showed that compared with control group, the levels of FBG and LDH of serum were increased obviously, HWI was increased, myocardial cells were hypertrophy, the collagen fibers of intercellular space of cell were increased, the protein expressions of GRP78 and caspase-12 of myocardium were increased in rats, myocardial cell apoptosis was increased in the model group(P&lt;0. 05). Compared with model group, FBG and LDH levels and HWI were reduced, the collagen fiber of intercellular space were decreased, the protein expression levels of GRP78 and caspase-12 were lowered in high dose of curcumin group(P&lt;0. 05). It indicates that Cur defends myocardium tissue in type 2 diabetes rats, which may be related to decreasing the level of blood glucose and the protein expressions of GRP78 and caspase-12, and blocking the ERS-initiated apoptotic.

Induction of Endoplasmic Reticulum Stress by Cadmium and Its Regulation on Nrf2 Signaling Pathway in Kidneys of Rats

ObjectiveThis study was conducted to investigate the regulation of endoplasmic reticulum stress on Nrf2 signaling pathway in the kidneys of rats. MethodsRats were divided into twelve groups of six animals each. Some groups were pre-administered with bacitracin or tauroursodeoxycholic acid (TUDCA), and all of them were treated with 5-20 μmol/kg cadmium (Cd) for 48 h. The oxidative stress levels were analyzed using kits. The mRNA and protein expression levels of endoplasmic reticulum stress-related factors and Nrf2 signaling pathway-related factors were determined using RT-PCR and western blot. ResultsCd exposure resulted in oxidative stress in the kidneys of rats and upregulated the expression of endoplasmic reticulum stress (ERS)-related factors and Nrf2 signaling pathway-related factors, especially at doses of 10 and 20 μmol/kg Cd, and the expression changes were particularly obvious. Moreover, after pretreatment with bacitracin, Cd upregulated the expression of ERS-related factors to a certain extent and, at higher doses, increased the mRNA expression of Nrf2. After pretreatment with TUDCA, Cd reduced the level of ERS to a certain extent; however, at these doses, there were no significant changes in the expression of Nrf2. ConclusionCadmium can result in ERS and oxidative stress in the kidneys of rats, activate Nrf2, and upregulate the transcriptional expression of phase II detoxification enzymes under these experimental conditions. ERS has a positive regulation effect on Nrf2 signaling pathway but has little effect on the negative regulation of Nrf2 signaling pathway in cadmium toxicity.

The role of endoplasmic reticulum stress in pulmonary hypertension in rat induced by chronic hypoxia and hypercapnia

To observe the pulmonary vascular remodeling in rats with pulmonary hypertension induced by hypoxia and hypercapnia, and to explore the role of endoplasmic reticulum stress in pulmonary hypertension. Forty SD rats were random-ly divided into four groups:normoxic control group (N), hypoxia hypercapnia group (HH), ERS inhibitor 4-phenylbutyric acid group (4-PBA), endoplasmic reticulum stress (ERS) pathway agonist tunicamycin group (TM), ten rats in each group.The mean pulmona-ry artery pressure (mPAP), mean carotid artery pressure (mCAP) and right ventricular hypertrophy index of rats in each group were measured.Pulmonary artery smooth muscle cells were identified by immunofluorescence α-smooth muscle actin (α-SMA).Morphologi-cal changes of lung tissue and pulmonary artery were observed by electron microscope.The apoptotic index of pulmonary artery smooth muscle cells in each group was detected by TUNEL.Reverse transcription polymerase chain reaction (RT-PCR) and Western blot were used to detect the expression of glucose-regulated protein (GRP78), C/EBP homologous protein (CHOP), c-Jun N-terminal kinase (JNK) and cysteinyl aspartate specific proteinase-12 (caspase-12) mRNA and protein in each group. ①Compared with the N group, the mPAP, the ratio of right ventricle weight to left ventricle plus ventricular septum weight[RV/(LV+S)]and the ratio of pulmonary artery wall area to total tube area (WA/TA) were increased (P<0.01), and the ratio of pulmonary artery luminal area to total tube area (LA/TA) were decreased (P<0.01), pulmonary artery smooth muscle cell apoptosis index were decreased (P<0.05 or P<0.01) in HH group, 4-PBA group and TM group.ERS related protein and mRNA expressions were increased, the differences were statistically significant.②Compared with the HH group, the mPAP, [RV/(LV+S)]and WA/TA of 4-PBA group were decreased (P <0.01), LA/TA and pulmonary artery smooth muscle cell apoptosis index were increased (P<0.01, P<0.05).The expressions of ERS related protein and mRNA were all decreased (P<0.05 or P<0.01).③Compared with the HH group, the mPAP, [RV/(LV+S)]and WA/TA of TM group were increased (P<0.05 or P<0.01), pulmonary artery middle layer thickened, LA/TA and pulmonary artery smooth muscle cell apoptotic index were decreased (P<0.01).ERS related protein and mRNA expressions were increased with statistical significance except GRP78 protein. Pulmonary vascular remodeling in rats with pulmonary hypertension induced by hypoxia and hypercapnia may be related to the excessive proliferation of pulmonary artery smooth muscle cells and too little apopto-sis;ERS related factors (JNK, caspase-12 and CHOP) are involved in the regulation of pulmonary hypertension induced by hypoxia hypercapnia.