Endoplasmic reticulum(ER)stress occurs when ER homeostasis is perturbed with accumulation of unfolded/misfolded protein or calcium depletion.The unfolded protein response(UPR),comprising of inositol-requiring enzyme 1...Endoplasmic reticulum(ER)stress occurs when ER homeostasis is perturbed with accumulation of unfolded/misfolded protein or calcium depletion.The unfolded protein response(UPR),comprising of inositol-requiring enzyme 1 a(IRE1 a),double-stranded RNA-dependent protein kinase(PKR)-like ER kinase(PERK)and activating transcription factor 6(ATF6)signaling pathways,is a protective cellular response activated by ER stress.However,UPR activation can also induce cell death upon persistent ER stress.The liver is susceptible to ER stress given its synthetic and other biological functions.Numerous studies from human liver samples and animal disease models have indicated a crucial role of ER stress and the UPR signaling pathways in the pathogenesis of liver diseases,including non-alcoholic fatty liver disease(NAFLD),alcoholic liver disease(ALD),alpha-1 antitrypsin(AAT)deficiency(AATD),cholestatic liver disease,drug-induced liver injury,ischemia/reperfusion(I/R)injury,viral hepatitis and hepatocel-lular carcinoma(HCC).Extensive investigations have demonstrated the potential underlying mechanisms of the induction of ER stress and the contribution of the UPR pathways during the development of the diseases.Moreover,ER stress and the UPR proteins and genes have become emerging therapeutic targets to treat liver diseases.展开更多
Endoplasmic reticulum(ER)stress,as an emerging hallmark feature of cancer,has a considerable impact on cell proliferation,metastasis,invasion,and chemotherapy resistance.Ovarian cancer(OvCa)is one of the leading cause...Endoplasmic reticulum(ER)stress,as an emerging hallmark feature of cancer,has a considerable impact on cell proliferation,metastasis,invasion,and chemotherapy resistance.Ovarian cancer(OvCa)is one of the leading causes of cancer-related mortality across the world due to the late stage of disease at diagnosis.Studies have explored the influence of ER stress on OvCa in recent years,while the predictive role of ER stress-related genes in OvCa prognosis remains unexplored.Here,we enrolled 552 cases of ER stress-related genes involved in OvCa from The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)cohorts for the screening of prognosis-related genes.The least absolute shrinkage and selection operator(LASSO)regression was applied to establish an ER stress-related risk signature based on the TCGA cohort.A seven-gene signature revealed a favorable predictive efficacy for the TCGA,International Cancer Genome Consortium(ICGC),and another GEO cohort(P<0.001,P<0.001,and P=0.04,respectively).Moreover,functional annotation indicated that this signature was enriched in cellular response and senescence,cytokines interaction,as well as multiple immune-associated terms.The immune infiltration profiles further delineated an immunologic unresponsive status in the high-risk group.In conclusion,ER stress-related genes are vital factors predicting the prognosis of OvCa,and possess great application potential in the clinic.展开更多
This study aimed to elucidate the molecular mechanisms by which berberine protects against cerebral ischemia/reperfusion(I/R)injury.The oxygen-glucose deprivation/reperfusion(OGD/R)PC12 model was established.Cell coun...This study aimed to elucidate the molecular mechanisms by which berberine protects against cerebral ischemia/reperfusion(I/R)injury.The oxygen-glucose deprivation/reperfusion(OGD/R)PC12 model was established.Cell counting kit-8(CCK-8)was used to detect the toxicity of berberine and the viability of PC12 cells.Hoechst 33258 staining and flow cytometry were used to observe the nuclear morphology,and changes of apoptosis and reactive oxygen species(ROS),respectively.Western blotting and immunofluorescence assay were employed to detect autophagy-related proteins[microtubule-associated protein 1A/1B-light chain 3(LC3),P62/SQSTM-1,Beclin-1]and endoplasmic reticulum(ER)stress-related markers[glucose-regulated protein 78(GRP78),C/EBP homologous protein(CHOP),Bcl-2-associated X(Bax)and cleaved caspase-3].The GFP-RFP-LC3 adenovirus was used to assay the change of autophagic flux.Our results showed that berberine could increase the viability of PC12 cells,decrease the concentrations of ROS after OGD/R treatment,and suppress OGD/R-induced ER stress and autophagy.Moreover,the results revealed the involvement of the mammalian target of rapamycin(mTOR)pathway in the induction of autophagy,and berberine could activate the phosphorylation of mTOR and thus mitigate autophagy.In conclusion,our study suggested that berberine may protect against OGD/R-induced apoptosis by regulating ER stress and autophagy,and it holds promises in the treatment of cerebral I/R injury.展开更多
Autophagy is a self-eating process,in which the damaged or excessed cell organelles and misfolded protein aggregates are removed from the cellular microenvironment.Autophagy is generally thought of as a pro-survival m...Autophagy is a self-eating process,in which the damaged or excessed cell organelles and misfolded protein aggregates are removed from the cellular microenvironment.Autophagy is generally thought of as a pro-survival mechanism which is not only important for balancing energy supply at times of nutrient deprivation but also in the removal of various stress stimuli to ensure homeostasis.In addition to the target materials of“self”origin,autophagy can also eliminate intracellular pathogens and acts as a defense mechanism to curb infections.In addition,autophagy is linked to the host cell's innate immune response.However,viruses have evolved various strategies to manipulate and overtake host cell machinery to establish productive replication and maintain infectious process.In fact,replication of many viruses has been found to be autophagy-dependent and suppression of autophagy can potentially affect the viral replication.Thus,autophagy can either serve as an anti-viral defense mechanism or a pro-viral process that supports viral replication.Hepatitis B virus(HBV)and hepatitis C virus(HCV)are known to co-opt cellular autophagy process as a pro-viral tool.Both viruses also induce mitophagy,which contributes to the establishment of chronic hepatitis.This review focuses on the roles of autophagy and mitophagy in the chronic liver disease pathogenesis associated with HBV and HCV infections.展开更多
The endoplasmic reticulum(ER)is an intracellular organelle consisting of a continuous network of membranes.In the liver,the ER is highly active in protein modification,lipid metabolism,and xenobiotic detoxification.Ma...The endoplasmic reticulum(ER)is an intracellular organelle consisting of a continuous network of membranes.In the liver,the ER is highly active in protein modification,lipid metabolism,and xenobiotic detoxification.Maintaining these complicated processes requires elaborate control of the ER lumen environment as well as the ER volume.Increasing evidence suggests that autophagy plays a critical role in regulating the homeostasis of hepatic ER contents and levels of cytochrome P450(CYP)enzymes via selective ER-phagy.This review will provide an overview of ER-phagy,summarizing the possible roles of recently identified ER-phagy receptor proteins in regulating the homeostasis of hepatic ER and CYP enzymes as well as outlining the various implications of ER-phagy in ER-related liver diseases.展开更多
Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", c...Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", commonly known as ‘Diabetes mellitus’ in clinics.Previous studies have shown that SGY-P could reduce insulin resistance and repair damaged pancreas in db/db mice, but the underlying mechanisms were unclear. Here, we investigated whether treatment with SGY-P could protect pancreatic β-cells from apoptosis and uncovered the underlying mechanisms. db/db mice were used to observe the hypoglycemic and islet protective effect in vivo. Apoptosis was induced in mouse insulinoma 6(MIN6) cells by palmitate, following which the cells were treated with SGY-P for elucidating the anti-apoptotic mechanism in vitro. Cell viability and nuclear morphology were detected by CCK-8 assay and Hoechst 33258 staining. The expression levels of apoptosis-, endoplasmic reticulum(ER) stress-, and autophagy-related proteins were measured by western blot. The results showed that SGY-P reduced fasting blood glucose, pancreatic pathological changes, and islet β-cell apoptosis in db/db mice. Palmitate-induced apoptosis in MIN6 cells was decreased by SGY-P treatment. Hence, SGY-P therapy exhibited a protective effect on pancreatic β-cells by decreasing the expression of cleaved caspase-3, cleaved PARP and Bax, and increasing Bcl-2 by suppressing ER stress(Bip/XBP1/IRE1α/CHOP/Caspase-12) and autophagy(LC3/p62/Atg5) pathways.2/Atg5 pathways.展开更多
Clinically,a large proportion of glaucoma patients undergo repeated intraocular pressure(IOP)spike(Spike IOP)attacks during their sleep,which may facilitate retinopathy.In this study,we established a mouse model of re...Clinically,a large proportion of glaucoma patients undergo repeated intraocular pressure(IOP)spike(Spike IOP)attacks during their sleep,which may facilitate retinopathy.In this study,we established a mouse model of repeated transient Spike IOP to investigate the direct damage to the retina following Spike IOP attacks,and elucidated the underlying molecular mechanism.We analyzed the changes in the number of retinal ganglion cells(RGCs)via immunofluorescence.Thereafter,we detected retinal cell apoptosis via terminal deoxynucleotidyl transferase deoxyuridine triphosphate(d UTP)nick-end labeling(TUNEL)staining,and performed RNA sequencing(RNA-seq)to reveal the underlying molecular mechanism.Finally,we validated the expression of key molecules in the endoplasmic reticulum(ER)stress pathway using quantitative real-time polymerase chain reaction(q RT-PCR)and western blot analysis.Results revealed a time-dependent RGC loss in Spike IOP,evidenced by a reduction in the number of Brn3 a-positive RGCs in experimental eyes following a 7-d continuous treatment with Spike IOP.In addition,TUNEL staining indicated that apoptosis of retinal cells started in the outer nuclear layer(ONL),and then spread to the ganglion cell layer(GCL)with time.RNA-seq analysis revealed that ER stress might be involved in Spike IOP-induced retinal injury.This result was corroborated by western blot,which revealed upregulation of ER stress-related proteins including binding immunoglobulin protein/glucose-regulated protein 78(Bi P/GRP78),phosphorylated inositolrequiring enzyme 1(p-IRE1),unspliced X-box-binding protein 1(XBP1-u),spliced X-box-binding protein 1(XBP1-s),phosphorylated c-Jun N-terminal kinase(p-JNK),C/EBP-homologous protein(CHOP),and B-cell lymphoma 2(Bcl-2)-associated X protein(Bax).These findings indicate that repeated IOP transients are detrimental to the retina,while ER stress plays an important role in retinal cell apoptosis in this situation.Notably,repeated Spike IOP among glaucoma patients is a crucial factor for progressive retino展开更多
Objective Autophagy is a highly conserved intracellular degradation pathway. Many picornaviruses induce autophagy to benefit viral replication, but an understanding of how autophagy occurs remains incomplete. In this ...Objective Autophagy is a highly conserved intracellular degradation pathway. Many picornaviruses induce autophagy to benefit viral replication, but an understanding of how autophagy occurs remains incomplete. In this study, we explored whether coxsackievirus B3(CVB3) infection induced autophagy through endoplasmic reticulum(ER) stress. Methods In CVB3-infected HeLa cells, the specific molecules of ER stress and autophagy were detected using Western blotting, reverse transcription polymerase chain reaction(RT-PCR), and confocal microscopy. Then PKR-like ER protein kinase(PERK) inhibitor, inositol-requiring protein-1(IRE1) inhibitor, or activating transcription factor-6(ATF6) inhibitor worked on CVB3-infected cells, their effect on autophagy was assessed by Western blotting for detecting microtubule-associated protein light chain 3(LC3). Results CVB3 infection induced ER stress, and ER stress sensors PERK/eIF2α, IRE1/XBP1, and ATF6 were activated. CVB3 infection increased the accumulation of green fluorescent protein(GFP)-LC3 punctuation and induced the conversion from LC3-Ⅰ to phosphatidylethanolamine-conjugated LC3-1(LC3-Ⅱ). CVB3 infection still decreased the expression of mammalian target of rapamycin(mTOR) and p-mTOR. Inhibition of PERK, IRE1, or ATF6 significantly decreased the ratio of LC3-Ⅱ to LC3-Ⅰ in CVB3-infected HeLa cells. Conclusion CVB3 infection induced autophagy through ER stress in HeL a cells, and PERK, IRE1, and ATF6 a pathways participated in the regulation of autophagy. Our data suggested that ER stress may inhibit mTOR signaling pathway to induce autophagy during CVB3 infection.展开更多
Recently we found that multi-walled carbon nanotube (MWCNT) exposure alters the mRNA levels of endoplasmic reticulum (ER) stress/autophagic genes, but the impact of biological molecules on this response is unclear. He...Recently we found that multi-walled carbon nanotube (MWCNT) exposure alters the mRNA levels of endoplasmic reticulum (ER) stress/autophagic genes, but the impact of biological molecules on this response is unclear. Herein, we compared the different actions of carboxylated MWCNTs (c-MWCNTs) pre-incubated with bovine serum albumin (BSA) or BSA-complexed free fatty acid (denoted as FFA) on macrophages derived from THP-1 monocytes (denoted as THP-1 macrophages). C-MWCNTs exhibited increased diameter and hydrodynamic size as well as decreased absolute zeta potential value after pre-incubation with BSA or FFA, which suggested a coating effect. Cytotoxicity or oxidative stress were not significantly induced after exposure to BSA-or FFA-coated c-MWCNTs. BSA-pre-incubated c-MWCNTs significantly enhanced the expression of the ER stress gene, DDIT3 and the autophagic genes, ATG5, BECN1, and PLIN2, but the mRNA levels of these genes was significantly decreased by FFA-pre-incubated c-MWCNTs. FFA-pre-incubated c-MWCNTs induced significantly higher lipid accumulation and interleukin-6 (IL-6) protein level compared with BSA-pre-incubated c-MWCNTs, which suggested that FFA-pre-incubated c-MWCNTs may more effectively induce the formation of macrophage foam cells. Collectively, our data indicated that pre-incubation with FFA may influence c-MWCNT-induced ER stress/autophagic gene expression and foam cell formation in THP-1 macrophages.展开更多
目的探讨蛋白酶体抑制剂(EPO)诱导前列腺癌DU145细胞凋亡机制是否与内质网应激(Er-stress)有关。方法用不同浓度的EPO处理DU145细胞,MTS检测细胞生长情况;流式细胞仪检测细胞凋亡率;实时定量聚合酶链反应(Real-time PCR)检测Er-stress...目的探讨蛋白酶体抑制剂(EPO)诱导前列腺癌DU145细胞凋亡机制是否与内质网应激(Er-stress)有关。方法用不同浓度的EPO处理DU145细胞,MTS检测细胞生长情况;流式细胞仪检测细胞凋亡率;实时定量聚合酶链反应(Real-time PCR)检测Er-stress相关分子同源蛋白质(CHOP)、葡萄糖调节蛋白78(GRP78)、X盒结合蛋白1(XBP-1)、剪接型X盒结合蛋白1信使核糖核酸(XBP-1s m RNA);Western blot检测CHOP和GRP78的表达。结果 EPO抑制DU145细胞生长,并且呈现浓度梯度依赖性,处理组细胞凋亡率高于未处理组,高剂量组凋亡率高于低剂量组。EPO处理后,CHOP、剪接型X盒结合蛋白1(XBP-1s)、GRP78 m RNA明显升高,72 h最明显。低剂量组与高剂量组48 h CHOP和XBP-1s的表达比较差异均有统计学意义,两组48和72 h GRP78比较差异均有统计学意义。EPO处理后,XBP-1和XBP-1s基因转录水平升高,而XBP-1s随处理时间增长基因转录水平变化不明显。EPO处理后,CHOP和GRP78不断增加,72 h两种蛋白质均达到最高值,并且低剂量组与高剂量组在72 h比较差异有统计学意义。结论 EPO能抑制DU145细胞生长,诱导凋亡,其机制可能与激活Er-stress,诱发内质网的相关分子CHOP、XBP-1s、XBP-1、GRP78的表达有关。展开更多
基金This work was supported by USA National Institute of Diabetes and Digestive and Kidney Diseases(NIDDK)R01 DK093807.
文摘Endoplasmic reticulum(ER)stress occurs when ER homeostasis is perturbed with accumulation of unfolded/misfolded protein or calcium depletion.The unfolded protein response(UPR),comprising of inositol-requiring enzyme 1 a(IRE1 a),double-stranded RNA-dependent protein kinase(PKR)-like ER kinase(PERK)and activating transcription factor 6(ATF6)signaling pathways,is a protective cellular response activated by ER stress.However,UPR activation can also induce cell death upon persistent ER stress.The liver is susceptible to ER stress given its synthetic and other biological functions.Numerous studies from human liver samples and animal disease models have indicated a crucial role of ER stress and the UPR signaling pathways in the pathogenesis of liver diseases,including non-alcoholic fatty liver disease(NAFLD),alcoholic liver disease(ALD),alpha-1 antitrypsin(AAT)deficiency(AATD),cholestatic liver disease,drug-induced liver injury,ischemia/reperfusion(I/R)injury,viral hepatitis and hepatocel-lular carcinoma(HCC).Extensive investigations have demonstrated the potential underlying mechanisms of the induction of ER stress and the contribution of the UPR pathways during the development of the diseases.Moreover,ER stress and the UPR proteins and genes have become emerging therapeutic targets to treat liver diseases.
基金This work was supported by the Shanghai Shenkang Hospital Development Center’s Shenkang Promotion of Clin‑ical Skills and Clinical Innovation in Municipal Hospitals Three-Year Action Plan(No.2020‒2023)the Major Clinical Research Project(No.SHDC2020CR1048B)the Pilot Construction Project of High-Level Universities in Shanghai(No.DGF501017-06),China。
文摘Endoplasmic reticulum(ER)stress,as an emerging hallmark feature of cancer,has a considerable impact on cell proliferation,metastasis,invasion,and chemotherapy resistance.Ovarian cancer(OvCa)is one of the leading causes of cancer-related mortality across the world due to the late stage of disease at diagnosis.Studies have explored the influence of ER stress on OvCa in recent years,while the predictive role of ER stress-related genes in OvCa prognosis remains unexplored.Here,we enrolled 552 cases of ER stress-related genes involved in OvCa from The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)cohorts for the screening of prognosis-related genes.The least absolute shrinkage and selection operator(LASSO)regression was applied to establish an ER stress-related risk signature based on the TCGA cohort.A seven-gene signature revealed a favorable predictive efficacy for the TCGA,International Cancer Genome Consortium(ICGC),and another GEO cohort(P<0.001,P<0.001,and P=0.04,respectively).Moreover,functional annotation indicated that this signature was enriched in cellular response and senescence,cytokines interaction,as well as multiple immune-associated terms.The immune infiltration profiles further delineated an immunologic unresponsive status in the high-risk group.In conclusion,ER stress-related genes are vital factors predicting the prognosis of OvCa,and possess great application potential in the clinic.
基金This work was supported by grants from the National Natural Science Foundation of China(No.81360199)Science and Technology Department of Guizhou Province(Guizhou Specific Grant[2019]4008)+2 种基金Science and Technology Plan Project of Guizhou Province(Basic Science and Technology Cooperation[2020]1Z060)Science and Technology Fund Project of Guizhou Health and Health Commission(No.gzwjkj2019-1-039)Science and Technology Fund Project of Southwest Guizhou Autonomous Prefecture(2019-1-10).
文摘This study aimed to elucidate the molecular mechanisms by which berberine protects against cerebral ischemia/reperfusion(I/R)injury.The oxygen-glucose deprivation/reperfusion(OGD/R)PC12 model was established.Cell counting kit-8(CCK-8)was used to detect the toxicity of berberine and the viability of PC12 cells.Hoechst 33258 staining and flow cytometry were used to observe the nuclear morphology,and changes of apoptosis and reactive oxygen species(ROS),respectively.Western blotting and immunofluorescence assay were employed to detect autophagy-related proteins[microtubule-associated protein 1A/1B-light chain 3(LC3),P62/SQSTM-1,Beclin-1]and endoplasmic reticulum(ER)stress-related markers[glucose-regulated protein 78(GRP78),C/EBP homologous protein(CHOP),Bcl-2-associated X(Bax)and cleaved caspase-3].The GFP-RFP-LC3 adenovirus was used to assay the change of autophagic flux.Our results showed that berberine could increase the viability of PC12 cells,decrease the concentrations of ROS after OGD/R treatment,and suppress OGD/R-induced ER stress and autophagy.Moreover,the results revealed the involvement of the mammalian target of rapamycin(mTOR)pathway in the induction of autophagy,and berberine could activate the phosphorylation of mTOR and thus mitigate autophagy.In conclusion,our study suggested that berberine may protect against OGD/R-induced apoptosis by regulating ER stress and autophagy,and it holds promises in the treatment of cerebral I/R injury.
基金This work was supported by USA National Institutes of Health(NIH)grants AI125350,AI085087 and AI139234 to A.Siddiqui.
文摘Autophagy is a self-eating process,in which the damaged or excessed cell organelles and misfolded protein aggregates are removed from the cellular microenvironment.Autophagy is generally thought of as a pro-survival mechanism which is not only important for balancing energy supply at times of nutrient deprivation but also in the removal of various stress stimuli to ensure homeostasis.In addition to the target materials of“self”origin,autophagy can also eliminate intracellular pathogens and acts as a defense mechanism to curb infections.In addition,autophagy is linked to the host cell's innate immune response.However,viruses have evolved various strategies to manipulate and overtake host cell machinery to establish productive replication and maintain infectious process.In fact,replication of many viruses has been found to be autophagy-dependent and suppression of autophagy can potentially affect the viral replication.Thus,autophagy can either serve as an anti-viral defense mechanism or a pro-viral process that supports viral replication.Hepatitis B virus(HBV)and hepatitis C virus(HCV)are known to co-opt cellular autophagy process as a pro-viral tool.Both viruses also induce mitophagy,which contributes to the establishment of chronic hepatitis.This review focuses on the roles of autophagy and mitophagy in the chronic liver disease pathogenesis associated with HBV and HCV infections.
基金This workwas partially supported by the USA National Institutes of Health(NIH)grants:R01 AA020518,R01 DK102142,U01 AA024733 and P20GM103549&P30GM118247.
文摘The endoplasmic reticulum(ER)is an intracellular organelle consisting of a continuous network of membranes.In the liver,the ER is highly active in protein modification,lipid metabolism,and xenobiotic detoxification.Maintaining these complicated processes requires elaborate control of the ER lumen environment as well as the ER volume.Increasing evidence suggests that autophagy plays a critical role in regulating the homeostasis of hepatic ER contents and levels of cytochrome P450(CYP)enzymes via selective ER-phagy.This review will provide an overview of ER-phagy,summarizing the possible roles of recently identified ER-phagy receptor proteins in regulating the homeostasis of hepatic ER and CYP enzymes as well as outlining the various implications of ER-phagy in ER-related liver diseases.
基金Chinese Medicine Research Program of Hunan Province(No.201999)。
文摘Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", commonly known as ‘Diabetes mellitus’ in clinics.Previous studies have shown that SGY-P could reduce insulin resistance and repair damaged pancreas in db/db mice, but the underlying mechanisms were unclear. Here, we investigated whether treatment with SGY-P could protect pancreatic β-cells from apoptosis and uncovered the underlying mechanisms. db/db mice were used to observe the hypoglycemic and islet protective effect in vivo. Apoptosis was induced in mouse insulinoma 6(MIN6) cells by palmitate, following which the cells were treated with SGY-P for elucidating the anti-apoptotic mechanism in vitro. Cell viability and nuclear morphology were detected by CCK-8 assay and Hoechst 33258 staining. The expression levels of apoptosis-, endoplasmic reticulum(ER) stress-, and autophagy-related proteins were measured by western blot. The results showed that SGY-P reduced fasting blood glucose, pancreatic pathological changes, and islet β-cell apoptosis in db/db mice. Palmitate-induced apoptosis in MIN6 cells was decreased by SGY-P treatment. Hence, SGY-P therapy exhibited a protective effect on pancreatic β-cells by decreasing the expression of cleaved caspase-3, cleaved PARP and Bax, and increasing Bcl-2 by suppressing ER stress(Bip/XBP1/IRE1α/CHOP/Caspase-12) and autophagy(LC3/p62/Atg5) pathways.2/Atg5 pathways.
基金supported by the Guangzhou Science and Technology Plan Project(Nos.201803040020,201903010065202102021099)+1 种基金the Guangdong Natural Science Foundation(No.2020A151501168)the Research Funds of the State Key Laboratory of Ophthalmology(No.PT1001022),China。
文摘Clinically,a large proportion of glaucoma patients undergo repeated intraocular pressure(IOP)spike(Spike IOP)attacks during their sleep,which may facilitate retinopathy.In this study,we established a mouse model of repeated transient Spike IOP to investigate the direct damage to the retina following Spike IOP attacks,and elucidated the underlying molecular mechanism.We analyzed the changes in the number of retinal ganglion cells(RGCs)via immunofluorescence.Thereafter,we detected retinal cell apoptosis via terminal deoxynucleotidyl transferase deoxyuridine triphosphate(d UTP)nick-end labeling(TUNEL)staining,and performed RNA sequencing(RNA-seq)to reveal the underlying molecular mechanism.Finally,we validated the expression of key molecules in the endoplasmic reticulum(ER)stress pathway using quantitative real-time polymerase chain reaction(q RT-PCR)and western blot analysis.Results revealed a time-dependent RGC loss in Spike IOP,evidenced by a reduction in the number of Brn3 a-positive RGCs in experimental eyes following a 7-d continuous treatment with Spike IOP.In addition,TUNEL staining indicated that apoptosis of retinal cells started in the outer nuclear layer(ONL),and then spread to the ganglion cell layer(GCL)with time.RNA-seq analysis revealed that ER stress might be involved in Spike IOP-induced retinal injury.This result was corroborated by western blot,which revealed upregulation of ER stress-related proteins including binding immunoglobulin protein/glucose-regulated protein 78(Bi P/GRP78),phosphorylated inositolrequiring enzyme 1(p-IRE1),unspliced X-box-binding protein 1(XBP1-u),spliced X-box-binding protein 1(XBP1-s),phosphorylated c-Jun N-terminal kinase(p-JNK),C/EBP-homologous protein(CHOP),and B-cell lymphoma 2(Bcl-2)-associated X protein(Bax).These findings indicate that repeated IOP transients are detrimental to the retina,while ER stress plays an important role in retinal cell apoptosis in this situation.Notably,repeated Spike IOP among glaucoma patients is a crucial factor for progressive retino
基金supported by the China Mega-project for Infectious Disease [2018ZX10102001,2018ZX10711001,2018ZX10734401,and 2018ZX10734404]the SKLID Development Grant [2011SKLID104]
文摘Objective Autophagy is a highly conserved intracellular degradation pathway. Many picornaviruses induce autophagy to benefit viral replication, but an understanding of how autophagy occurs remains incomplete. In this study, we explored whether coxsackievirus B3(CVB3) infection induced autophagy through endoplasmic reticulum(ER) stress. Methods In CVB3-infected HeLa cells, the specific molecules of ER stress and autophagy were detected using Western blotting, reverse transcription polymerase chain reaction(RT-PCR), and confocal microscopy. Then PKR-like ER protein kinase(PERK) inhibitor, inositol-requiring protein-1(IRE1) inhibitor, or activating transcription factor-6(ATF6) inhibitor worked on CVB3-infected cells, their effect on autophagy was assessed by Western blotting for detecting microtubule-associated protein light chain 3(LC3). Results CVB3 infection induced ER stress, and ER stress sensors PERK/eIF2α, IRE1/XBP1, and ATF6 were activated. CVB3 infection increased the accumulation of green fluorescent protein(GFP)-LC3 punctuation and induced the conversion from LC3-Ⅰ to phosphatidylethanolamine-conjugated LC3-1(LC3-Ⅱ). CVB3 infection still decreased the expression of mammalian target of rapamycin(mTOR) and p-mTOR. Inhibition of PERK, IRE1, or ATF6 significantly decreased the ratio of LC3-Ⅱ to LC3-Ⅰ in CVB3-infected HeLa cells. Conclusion CVB3 infection induced autophagy through ER stress in HeL a cells, and PERK, IRE1, and ATF6 a pathways participated in the regulation of autophagy. Our data suggested that ER stress may inhibit mTOR signaling pathway to induce autophagy during CVB3 infection.
基金financially supported by the National Natural Science Foundation of China (No. 21707114)Scientific Research Foundation of Hunan Provincial Education Department (No. 17A205)
文摘Recently we found that multi-walled carbon nanotube (MWCNT) exposure alters the mRNA levels of endoplasmic reticulum (ER) stress/autophagic genes, but the impact of biological molecules on this response is unclear. Herein, we compared the different actions of carboxylated MWCNTs (c-MWCNTs) pre-incubated with bovine serum albumin (BSA) or BSA-complexed free fatty acid (denoted as FFA) on macrophages derived from THP-1 monocytes (denoted as THP-1 macrophages). C-MWCNTs exhibited increased diameter and hydrodynamic size as well as decreased absolute zeta potential value after pre-incubation with BSA or FFA, which suggested a coating effect. Cytotoxicity or oxidative stress were not significantly induced after exposure to BSA-or FFA-coated c-MWCNTs. BSA-pre-incubated c-MWCNTs significantly enhanced the expression of the ER stress gene, DDIT3 and the autophagic genes, ATG5, BECN1, and PLIN2, but the mRNA levels of these genes was significantly decreased by FFA-pre-incubated c-MWCNTs. FFA-pre-incubated c-MWCNTs induced significantly higher lipid accumulation and interleukin-6 (IL-6) protein level compared with BSA-pre-incubated c-MWCNTs, which suggested that FFA-pre-incubated c-MWCNTs may more effectively induce the formation of macrophage foam cells. Collectively, our data indicated that pre-incubation with FFA may influence c-MWCNT-induced ER stress/autophagic gene expression and foam cell formation in THP-1 macrophages.
文摘目的探讨蛋白酶体抑制剂(EPO)诱导前列腺癌DU145细胞凋亡机制是否与内质网应激(Er-stress)有关。方法用不同浓度的EPO处理DU145细胞,MTS检测细胞生长情况;流式细胞仪检测细胞凋亡率;实时定量聚合酶链反应(Real-time PCR)检测Er-stress相关分子同源蛋白质(CHOP)、葡萄糖调节蛋白78(GRP78)、X盒结合蛋白1(XBP-1)、剪接型X盒结合蛋白1信使核糖核酸(XBP-1s m RNA);Western blot检测CHOP和GRP78的表达。结果 EPO抑制DU145细胞生长,并且呈现浓度梯度依赖性,处理组细胞凋亡率高于未处理组,高剂量组凋亡率高于低剂量组。EPO处理后,CHOP、剪接型X盒结合蛋白1(XBP-1s)、GRP78 m RNA明显升高,72 h最明显。低剂量组与高剂量组48 h CHOP和XBP-1s的表达比较差异均有统计学意义,两组48和72 h GRP78比较差异均有统计学意义。EPO处理后,XBP-1和XBP-1s基因转录水平升高,而XBP-1s随处理时间增长基因转录水平变化不明显。EPO处理后,CHOP和GRP78不断增加,72 h两种蛋白质均达到最高值,并且低剂量组与高剂量组在72 h比较差异有统计学意义。结论 EPO能抑制DU145细胞生长,诱导凋亡,其机制可能与激活Er-stress,诱发内质网的相关分子CHOP、XBP-1s、XBP-1、GRP78的表达有关。
