scholarly journals Endoplasmic reticulum stress is involved in spiral ganglion neuron apoptosis following chronic kanamycin-induced deafness

2019 ◽  
Vol 39 (2) ◽  
Author(s):  
Yaqin Tu ◽  
Guorun Fan ◽  
Haiying Sun ◽  
Xiong Cai ◽  
Wen Kong
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Time Course ◽  
Morphological Changes ◽  
Spiral Ganglion ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Spiral Ganglion Neuron ◽  
Tunel Staining ◽  
Caspase 12 ◽  
Ganglion Neurons

Abstract Aminoglycoside antibiotics-induced hearing loss is a common sensorineural impairment. Spiral ganglion neurons (SGNs) are first-order neurons of the auditory pathway and are critical for the maintenance of normal hearing. In the present study, we investigated the time-course of morphological changes and the degeneration process of spiral ganglion cells (SGCs) following chronic kanamycin-induced deafness and determined whether the endoplasmic reticulum (ER) stress was involved in the degeneration of SGNs. We detected density changes in SGCs and the expressions of Bip, inositol requirement 1 (IRE1)α, activating transcription factor-6α, p-PERK, p-eIF2α, CHOP, and caspase-12 at each time point after kanamycin treatment. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining was also performed. The number of SGC deletions reached ∼50% at the 70th day after kanamycin administration and the ER of most SGCs were dilated. The expression of p-PERK, p-eIF2α, p-IRE1α, Bip, caspase-12, and Chop was significantly unregulated after kanamycin treatment. The number of SGCs that were positive for both TUNEL and caspase-12 increased from day 7 to 28. Taken together, these data demonstrate that ER stress was involved in kanamycin-induced apoptosis of SGNs. Kanamycin-induced SGN apoptosis is mediated, at least in part, by ER stress-induced upregulation of CHOP and caspase-12.

Early activation of endoplasmic reticulum stress is associated with arginine-induced acute pancreatitis

2006 ◽  
Vol 291 (2) ◽  
pp. G238-G245 ◽  
Author(s):  
Constanze H. Kubisch ◽  
Maria Dolors Sans ◽  
Thiruvengadam Arumugam ◽  
Stephen A. Ernst ◽  
John A. Williams ◽  
...  
Keyword(s):  
Acute Pancreatitis ◽  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Acinar Cell ◽  
Serum Amylase ◽  
Cell Damage ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Tunel Staining ◽  
Early Activation ◽  
Caspase 12

Endoplasmic reticulum (ER) stress mechanisms have been found to play critical roles in a number of diseases states, such as diabetes mellitus and Alzheimer disease, but whether they are involved in acute pancreatitis is unknown. Here we show for the first time that all major ER stress sensing and signaling mechanisms are present in exocrine acini and are activated early in the arginine model of experimental acute pancreatitis. Pancreatitis was induced in rats by intraperitoneal injection of 4.0 g/kg body wt arginine. Pancreatitis severity was assessed by analysis of serum amylase, pancreatic trypsin activity, water content, and histology. ER stress-related molecules PERK, eIF2α, ATF6, XBP-1, BiP, CHOP, and caspase-12 were analyzed. Arginine treatment induced rapid and severe pancreatitis, as indicated by increased serum amylase, pancreatic tissue edema, and acinar cell damage within 4 h. Arginine treatment also caused an early activation of ER stress, as indicated by phosphorylation of PERK and its downstream target eIF2α, ATF6 translocation into the nucleus (within 1 h), and upregulation of BiP (within 4 h). XBP-1 splicing and CHOP expression were observed within 8 h. After 24 h, increased activation of the ER stress-related proapoptotic molecule caspase-12 was observed along with an increase in caspase-3 activity and TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labeling (TUNEL) staining in exocrine acini. These results indicate that ER stress is an important early acinar cell event that likely contributes to the development of acute pancreatitis in the arginine model.

scholarly journals Electroacupuncture Regulates Endoplasmic Reticulum Stress and Ameliorates Neuronal Injury in Rats with Acute Ischemic Stroke

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Ya-min Zhang ◽  
Hong Xu ◽  
Su-hui Chen ◽  
Hua Sun
Keyword(s):  
Endoplasmic Reticulum ◽  
Ischemic Stroke ◽  
Endoplasmic Reticulum Stress ◽  
Infarct Volume ◽  
Neuronal Injury ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Tunel Staining ◽  
Model Group ◽  
Expression Levels ◽  
Caspase 12

Ischemic stroke is a common cause of morbidity, mortality, and disability worldwide. Electroacupuncture (EA) is an effective method for alleviating brain damage after ischemic stroke. However, the underlying mechanism has not been fully elucidated. This study aimed to determine whether endoplasmic reticulum stress (ERS) could contribute to the protective effects of EA in cerebral ischemia/reperfusion injury (CIRI) to provide a rationale for the widespread clinical use of EA. Rats were divided into the sham-operated (sham) group, the CIRI (model) group, and the EA group. Rats in the model group were subjected to middle cerebral artery occlusion (MCAO) for 2 h followed by 72 h of reperfusion. Rats with CIRI were treated daily with EA at GV20 and ST36 for a total of 3 days. The Longa scoring system and adhesive removal somatosensory test were applied to evaluate neurological deficits. Then, 2,3,5-triphenyltetrazolium chloride (TTC) staining was performed to measure the infarct volume. Immunofluorescence staining for NeuN and GFAP and terminal deoxynucleotidyl transferase- (TdT-) mediated dUTP nick-end labeling (TUNEL) staining were performed to detect apoptotic cells in brain tissue. Immunohistochemistry, quantitative real-time polymerase chain reaction (qPCR), and western blotting were used to measure the levels of ERS indicators (GRP78, CHOP/GADD153, p-eIF2α, and caspase 12). The results showed that EA significantly reduced the cerebral infarct volume, improved neurological function, and inhibited neuronal apoptosis. In the EA group compared with the model group, the mRNA expression levels of GRP78 were significantly increased, and the expression levels of proapoptotic proteins (CHOP/GADD153, p-eIF2α, and caspase 12) were significantly decreased. These results suggest that the possible mechanism by which EA protects cells against neuronal injury in CIRI may involve inhibiting endoplasmic reticulum stress.

scholarly journals LncRNA-DRSGN (Down-Regulated in Spiral Ganglion Neurons) Competes with miR-27a for Binding to FAM172A, Supervises SGN Degeneration and ER (Endoplasmic Reticulum) Stress-Autophagy in CHARGE Syndrome

2021 ◽  
Author(s):  
Xian-Bao Cao ◽  
Bi-Zhang Lu ◽  
Jia-Hong Pei ◽  
Cun Feng ◽  
Yan-Fei Guan ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Endoplasmic Reticulum Stress ◽  
Er Stress ◽  
Current Model ◽  
Spiral Ganglion ◽  
Noncoding Rnas ◽  
Charge Syndrome ◽  
Calcium Flux ◽  
Knockout Mouse Model ◽  
Ganglion Neurons

Abstract Background: Hearing loss is one of the most common disabilities in the world and brings a heavy burden to society. The current model is not stable enough, and it has caused serious model interference to clarify the pathogenesis of CHARGE syndrome. Methods: The knockout mouse model of FAM172A gene was constructed, and sits phenotype was identified. Besides, the next-genesequencing experiments of noncoding RNAs were performed utilizing the primary SGNs of model mice. The biofunctions of FAM172A in the relationships between ER (Endoplasmic reticulum) stress, autophagy, and intracellular calcium flux were investigated. Moreover, the above role associated with the competitive combination among LncRNA-DRSGN, miR-27a, and FAM172A were studied in the progression of SGN degeneration and autophagy in the model of CHARGE syndrome. Results: FAM172A(-/-) exhibited abnormal hearing, growth retardation, abnormal eye development, and dysgnosia. It was in line with the phenotype of CHARGE syndrome. Moreover, there was degeneration of SGNs in FAM172A(-/-) mice, and the differential expression of noncoding RNAs in primary SGNs were found and identified, including miR-27a and LncRNA-DRSGN. LncRNA-DRSGN regulated miR-27a as a ceRNA, and miR-27a inhibited FAM172A expression, LncRNA-DRSGN competed with miR-27a for binding to FAM172A, which participated in the regulation of ER stress-related calcium flux. LncRNA-DRSGN regulated the autophagy process of neurons by competing with miR-27a for binding to FAM172A. Conclusion: LncRNA-DRSGN competed with miR-27a for binding to FAM172A, participated in regulating ER stress-related calcium flux, then affected neuron degeneration and autophagy process of SGNs in the model of CHARGE syndrome.

Abstract MP120: Ryanodine Receptor 2 Prevents Endoplasmic Reticulum Stress-induced Defects in Cardiomyocyte Maturation

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Yuxuan Guo ◽  
Blake Jardin ◽  
Silvia Guatimosim ◽  
William T Pu
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Ryanodine Receptor ◽  
Heart Development ◽  
Calcium Oscillations ◽  
Basic Knowledge ◽  
Calcium Handling ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Tunel Staining ◽  
Stress Relieving

Cardiomyocyte (CM) maturation is an essential step in heart development that prepares the organ for robust and sustainable contractions. However, very little is known about the contribution of CM maturation to cardiac pathogenesis, partly due to the paucity of basic knowledge about maturation. For example, although calcium plays well-established roles in cardiac stress responses, whether normal calcium oscillations are required for CM maturation has not been examined. To address this question, we created mice with mosaic inactivation of ryanodine receptor 2 (RYR2), the central channel that mediates calcium release from sarcoplasmic/endoplasmic reticulum (SR/ER), in CMs. RYR2 was depleted in a small fraction of neonatal CMs by adeno-associated virus (AAV)-mediated Cas9-based somatic mutagenesis (CASAAV). The majority of CMs intentionally retained RYR2 expression, which circumvented cardiac dysfunction and death that would result from widespread calcium transient ablation. In RYR2-depleted CMs, sarcomere expansion, transverse-tubule formation, mitochondrial metabolism and maturational hypertrophy were impaired. Strikingly, these maturation defects were associated with unfolded protein response, with the activation of three major ER stress effectors: spliced Xbp1 (XBP1s), ATF6 N-terminus (ATF6N), and ATF4. Tauroursodeoxycholic acid, an ER stress-relieving drug, partially rescued the maturation defects in RYR2-depleted CMs. Over expression (OE) of XBP1s, ATF6N or ATF4 each impaired CM maturation, with phenotypic severity ranked as ATF4>XBP1s>ATF6N. ATF4 OE in neonatal CMs triggered both ultrastructural and transcriptomic changes highly correlated with RYR2 depletion phenotypes. Interestingly, ATF4 OE altered distinct transcriptomic programs in neonatal vs. adult CMs and caused ultrastructural defects only at the neonatal stage. No terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was detected upon RYR2 depletion or ATF4 OE, excluding the contribution of apoptosis to maturation defects. Collectively, these data indicate that RYR2-mediated calcium handling prevents an adverse and stage-specific impact of ER stress on CM maturation, which lead to CM dysfunction.

Abstract 276: SR-BI Suppresses Apoptosis by Reducing Endoplasmic Reticulum Stress and Promoting Akt Activity in Macrophages

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Huan Tao ◽  
Patricia G Yancey ◽  
Sean S Davies ◽  
L Jackson Roberts ◽  
John L Blakemore ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Apolipoprotein E ◽  
Er Stress ◽  
Cell Apoptosis ◽  
Free Cholesterol ◽  
Oxidized Ldl ◽  
Tunel Staining ◽  
Type I ◽  
Apoptotic Cells ◽  
Atherosclerotic Lesions

Objective: Macrophage apoptosis contributes to atherosclerotic plaque necrosis, inflammation, development and rupture. Scavenger receptor class B type I (SR-BI) is a key regulator of HDL metabolism and cellular cholesterol homeostasis. Here we examined the hypothesis that macrophage SR-BI modulates lipid-associated cellular stress and apoptosis. Methods and Results: In vitro cell apoptosis assays were performed in primary macrophages, and for in vivo evidence, we examined TUNEL staining of atherosclerotic lesions of LDLR -/- mice that were reconstituted with SR-BI -/- or WT bone marrow after 16 weeks on a Western diet. We found that SR-BI deficiency led to ~64.3% more apoptotic cells induced by oxidized LDL or free cholesterol in primary macrophages, and 6-fold more lesional apoptotic cells in SR-BI -/- →LDLR -/- mice compared to WT recipient mice. In macrophages, SR-BI deficiency caused significant accumulations of cellular free cholesterol and elevated markers of endoplasmic reticulum (ER) stress. These were exacerbated by feeding mice a high-cholesterol diet or inactivating the apolipoprotein E gene. Peroxidation of lipoproteins and cell membranes leads to modification of phosphatidylethanolamine by lipid aldehydes including isolevuglandins (IsoLG-PE). Treatment of macrophages with IsoLG-PE induced 52.6% more apoptotic cells in SR-BI -/- macrophages compared to WT. Transgenic expression of SR-BI by transfection of SR-BI -/- macrophages rescued oxidative stress-induced ER stress and cell apoptosis. SR-BI deficiency inhibited the Akt pathway compromising macrophage survival and increasing lesion necrosis. Moreover, Akt Activator was able to rescue SR-BI deficiency associated apoptosis in macrophages. Apolipoprotein E interacts with SR-BI in macrophages, co-operating for cellular lipid homeostasis and cell survival signaling. Conclusion: SR-BI protects against cell apoptosis induced by lipid stress in macrophages and atherosclerotic lesions. The underlying mechanisms are, at least in part, through reducing lipid-associated ER stress and promoting Akt activity in macrophages. Thus, we identify macrophage SR-BI-mediated apoptosis pathways as molecular targets for the prevention of atherosclerotic cardiovascular events.

scholarly journals P0015SIRT2 REGULATES CISPLATIN-INDUCED ENDOPLASMIC RETICULUM STRESS THROUGH HEAT SHOCK FACTOR 1 DEACETYLATION

2020 ◽  
Vol 35 (Supplement_3) ◽  
Author(s):  
Woong Park ◽  
Hyeongwan Kim ◽  
Yujin Jung ◽  
Kyung Pyo Kang ◽  
Won Kim
Keyword(s):  
Endoplasmic Reticulum ◽  
Heat Shock ◽  
Er Stress ◽  
Knockout Mice ◽  
Malignant Tumors ◽  
Heat Shock Factor ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Heat Shock Factor 1 ◽  
Caspase 12

Abstract Background and Aims Nephrotoxicity is an important cisplatin-induced adverse reaction and restricts the use of cisplatin to treat malignant tumors. Endoplasmic reticulum (ER) stress is caused by the accumulation of misfolded proteins, and is induced by cisplatin in kidneys. SIRT2 nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase is a member of the sirtuin family, but its role in cisplatin-induced ER stress remains unclear. Method To investigate the effect of SIRT2 on cisplatin-induced ER stress using SIRT2 knockout mice and human proximal tubular epithelial cells (HK-2 cells). We treated cisplatin (20 µg/mL) or induced by intraperitoneal injection of cisplatin (20 mg/kg) and evaluated the changes of ER stress and its signal mechanism. Results Cisplatin administration was found to significantly increase the expressions of PRKR-like ER kinase (PERK), phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), and the C/EBP homologous protein (CHOP) and caspase-12 in the kidneys of SIRT2-wild type mice. However, cisplatin-induced increases in the expressions of p-PERK, p-eIF2α, CHOP and, caspase-12 were diminished in kidneys of SIRT2 knockout mice. In vitro, cisplatin significantly increased the expressions of p-PERK, p-eIF2α, CHOP, and caspase-12 in HK-2 cells. When the effect of SIRT2 on cisplatin-induced ER stress was evaluated using SIRT2-siRNA (ON-TARGET plus human SIRT2 siRNA) or the SIRT2 inhibitors, AGK2 and AK1, knockdown or inhibition of SIRT2 significantly attenuated the cisplatin-induced protein expression of p-PERK, p-eIF2α, CHOP, and caspase-12. Immunoprecipitation studies showed SIRT2 bound physically to heat shock factor (HSF)1 and that HSF1 acetylation was significantly increased by cisplatin. In addition, knockdown of SIRT2 increased cisplatin-induced HSF1 acetylation and increased the expression of heat shock protein (HSP)70. Conclusion These observations suggest that suppression of SIRT2 ameliorates cisplatin-induced ER stress by increasing HSF1 acetylation and HSP expression.

scholarly journals Neuroprotective effects of verbascoside against Alzheimer’s disease via the relief of endoplasmic reticulum stress in Aβ-exposed U251 cells and APP/PS1 mice

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Chunyue Wang ◽  
Xueying Cai ◽  
Ruochen Wang ◽  
Siyu Zhai ◽  
Yongfeng Zhang ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Amyloid Β ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Morris Water Maze Test ◽  
Proteomics Analysis ◽  
Neuroprotective Effects ◽  
U251 Cells

Abstract Background Endoplasmic reticulum (ER) stress is involved in the progression of Alzheimer’s disease (AD). Verbascoside (VB), an active phenylethanoid glycoside that was first isolated from Verbascum sinuatum (the wavyleaf mullein), possesses anti-inflammatory, antioxidative, and anti-apoptotic effects. The purpose of this study was to elucidate the beneficial effects of VB in amyloid β (Aβ)1–42-damaged human glioma (U251) cells and in APPswe/PSEN1dE9 transgenic (APP/PS1) mice. Methods U251 cells were co-incubated with 10 μM of Aβ1-42 and treated with VB. The protective effects of VB were investigated by using 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide assay, flow cytometry, fluorescence staining, and transmission electron microscopy. APP/PS1 transgenic mice were treated for 6 weeks with VB. Learning and memory were evaluated using a Morris water maze test. Immunohistochemistry, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling, thioflavin-S staining, and proteomics analysis were performed to study the potential neuroprotective mechanism. Enzyme-linked immunosorbent assays and western blot were performed to analyze altered protein levels of brain lysates in APP/PS1 mice and/or Aβ1-42-damaged U251 cells. Results In Aβ1-42-damaged U251 cells, VB significantly improved cell viability, inhibited apoptosis, reduced calcium accumulation and the intracellular concentrations of reactive oxygen species, and improved the morphology of mitochondria and ER. In APP/PS1 mice, 6-week administration of VB significantly improved memory and cognition. VB inhibited apoptosis, reduced the deposition of Aβ, reduced the formation of neurofibrillary tangles formed by hyperphosphorylated tau protein, and downregulated the expression levels of 4-hydroxynonenal and mesencephalic astrocyte-derived neurotrophic factor in the brains of APP/PS1 mice. Proteomics analysis of mouse hippocampus suggested that the neuroprotective effect of VB may be related to the reduction of ER stress. This was indicated by the fact that VB inhibited the three branches of the unfolded protein response, thereby attenuating ER stress and preventing apoptosis. Conclusions The results confirmed that VB possesses significant neuroprotective effects, which are related to the reduction of ER stress. These findings support the status of VB as a potentially effective treatment for AD and warrant further research.

scholarly journals Pulsed infrared releases Ca2+ from the endoplasmic reticulum of cultured spiral ganglion neurons

2018 ◽  
Vol 120 (2) ◽  
pp. 509-524 ◽  
Author(s):  
John N. Barrett ◽  
Samantha Rincon ◽  
Jayanti Singh ◽  
Cristina Matthewman ◽  
Julio Pasos ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Infrared Radiation ◽  
Transient Receptor Potential ◽  
Trp Channels ◽  
Spiral Ganglion ◽  
Receptor Potential ◽  
Transient Temperature ◽  
Spiral Ganglion Neurons ◽  
Transient Receptor ◽  
Ganglion Neurons

Inner ear spiral ganglion neurons were cultured from day 4 postnatal mice and loaded with a fluorescent Ca2+ indicator (fluo-4, -5F, or -5N). Pulses of infrared radiation (IR; 1,863 nm, 200 µs, 200–250 Hz for 2–5 s, delivered via an optical fiber) produced a rapid, transient temperature increase of 6–12°C (above a baseline of 24–30°C). These IR pulse trains evoked transient increases in both nuclear and cytosolic Ca2+ concentration ([Ca2+]) of 0.20–1.4 µM, with a simultaneous reduction of [Ca2+] in regions containing endoplasmic reticulum (ER). IR-induced increases in cytosolic [Ca2+] continued in medium containing no added Ca2+ (±Ca2+ buffers) and low [Na+], indicating that the [Ca2+] increase was mediated by release from intracellular stores. Consistent with this hypothesis, the IR-induced [Ca2+] response was prolonged and eventually blocked by inhibition of ER Ca2+-ATPase with cyclopiazonic acid, and was also inhibited by a high concentration of ryanodine and by inhibitors of inositol (1,4,5)-trisphosphate (IP3)-mediated Ca2+ release (xestospongin C and 2-aminoethoxydiphenyl borate). The thermal sensitivity of the response suggested involvement of warmth-sensitive transient receptor potential (TRP) channels. The IR-induced [Ca2+] increase was inhibited by TRPV4 inhibitors (HC-067047 and GSK-2193874), and immunostaining of spiral ganglion cultures demonstrated the presence of TRPV4 and TRPM2 that colocalized with ER marker GRP78. These results suggest that the temperature sensitivity of IR-induced [Ca2+] elevations is conferred by TRP channels on ER membranes, which facilitate Ca2+ efflux into the cytosol and thereby contribute to Ca2+-induced Ca2+-release via IP3 and ryanodine receptors. NEW & NOTEWORTHY Infrared radiation-induced photothermal effects release Ca2+ from the endoplasmic reticulum of primary spiral ganglion neurons. This Ca2+ release is mediated by activation of transient receptor potential (TRPV4) channels and involves amplification by Ca2+-induced Ca2+-release. The neurons immunostained for warmth-sensitive channels, TRPV4 and TRPM2, which colocalize with endoplasmic reticulum. Pulsed infrared radiation provides a novel experimental tool for releasing intracellular Ca2+, studying Ca2+ regulatory mechanisms, and influencing neuronal excitability.

scholarly journals Autophagy activated via GRP78 to alleviate endoplasmic reticulum stress for cell survival in blue light-mediated damage of A2E-laden RPEs

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Jingyang Feng ◽  
Yuhong Chen ◽  
Bing Lu ◽  
Xiangjun Sun ◽  
Hong Zhu ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Signaling Pathway ◽  
Blue Light ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Degenerative Diseases ◽  
Caspase 12 ◽  
Age Related ◽  
Retinal Degenerative Diseases

Abstract Background Retinal pigment epithelium cells (RPEs) are critical for maintaining retinal homeostasis. Accumulation of age-related lipofuscin, N-retinylidene-N-retinylethanolamine (A2E), makes RPEs vulnerable to blue light-mediated damage, which represents an initial cause of some retinal degenerative diseases. This study investigated the activation of autophagy and the signaling pathway involved in glucose-related protein 78 (GRP78) induced autophagy in blue light-mediated damage of A2E-laden RPEs. In addition, we explored whether autophagy could play a protective role by alleviating endoplasmic reticulum (ER) stress to promote RPEs survival. Methods RPEs were incubated with 25 μM A2E for 2 h and exposed to blue light for 20 min. The expression of ER stress-related apoptotic proteins, CHOP and caspase-12, as well as autophagy marker LC3 were measured by western blot analysis. Autophagosomes were observed by both transmission electron microscopy and immunofluorescence assays. GRP78 interference performed by short hairpin RNA (shRNA) was used to identify the signaling pathway involved in GRP78 induced autophagy. Cell death was assessed using TUNEL analysis. Results Treatment with A2E and blue light markedly increased the expression of ER stress-related apoptotic molecules CHOP and caspase-12. The activation of autophagy was recognized by observing autophagosomes at ultrastructural level. Additionally, punctate distributions of LC3 immunofluorescence and enhanced conversions of LC3-I to LC3-II were found in A2E and blue light-treated RPEs. Moreover, GRP78 interference reduced AMPK phosphorylation and promoted mTOR activity, thereby downregulating autophagy. In addition, the inhibition of autophagy made RPEs vulnerable to A2E and blue light damage. In contrast, the autophagy inducer rapamycin alleviated ER stress to promote RPEs survival. Conclusions GRP78 activates autophagy via AMPK/mTOR in blue light-mediated damage of A2E-laden RPEs in vitro. Autophagy may be a vital endogenous cytoprotective process to alleviate stress for RPEs survival in retinal degenerative diseases.

scholarly journals Endoplasmic reticulum stress regulates cell injury in lipopolysaccharide-induced nerve cells

2020 ◽  
Vol 48 (9) ◽  
pp. 030006052094976
Author(s):  
Min Li ◽  
Ying Zhang ◽  
Jixing Wang
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Viability ◽  
Er Stress ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Cell Injury ◽  
Cell Counting ◽  
Tunel Staining ◽  
Dependent Manner ◽  
Stress Markers

Objective Sepsis-associated encephalopathy (SAE) is a common complication of sepsis, and excessive endoplasmic reticulum (ER) stress is closely correlated with the cell injury caused by sepsis. This study aimed to analyze the possible role of ER stress in SAE cell models. Methods PC12 and MES23.5 cells were treated with increasing concentrations of lipopolysaccharides (LPS). The Cell Counting Kit-8 assay was used to detect cell viability and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was performed to assess cell apoptosis. In addition, the protein expression levels of ER stress markers [GRP78, CHOP, inositol-requiring enzyme 1 (IRE1), and PKR-like ER kinase (PERK)] and apoptosis-related proteins (Bax, Bcl-2, caspase-3, and cleaved caspase-3) were analyzed using western blotting. Results LPS treatment activated ER stress markers in both the PC12 and MES23.5 cells. The overexpression of GRP78 significantly reduced cell viability and enhanced cell apoptosis in a time-dependent manner. An ER stress inhibitor, 4-PBA, significantly enhanced cell viability and inhibited the cell apoptosis induced by LPS. Therefore, an enhanced unfolded protein response (UPR) and UPR suppression may regulate cell apoptosis. Conclusions UPR was shown to be involved in regulating LPS-induced neuron injury. UPR could be a potential therapeutic target in SAE.

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