Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods...Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods:Human nasopharyngeal carcinoma CNE multicellular spheroids(MCS) were constructed with three dimensional cell culture methods.Western blot was employed to analyze the activity of JNK signaling pathway in MCS after X-ray irradiation,and the expression of caspase-3 protein before and after using SP600125(a special inhibitor of JNK).X-ray induced cell apoptosis in MCS before and after treated with SP600125 were detected by TUNEL.Results:The level of JNK phosphorylation in MCS was a dynamic course after radiation,and there was a phosphorylation peaks at 2 h later,the apoptotic rate of MCS(P < 0.05) and the expression of caspase-3 protein(P < 0.05) were significantly increased after treated with SP600125.Conclusion:The transient activation of JNK played a important role in sensitivity to radiotherapy of CNE MCS via mediating survival signals,blocking this pathway accelerate cell apoptosis,which may be related to the increased expression of caspase-3.展开更多
Prolonged activation of adenosine A1 receptor likely leads to damage of dopaminergic neurons and subsequent development of neurodegenerative diseases.However,the pathogenesis underlying long-term adenosine A1 receptor...Prolonged activation of adenosine A1 receptor likely leads to damage of dopaminergic neurons and subsequent development of neurodegenerative diseases.However,the pathogenesis underlying long-term adenosine A1 receptor activation-induced neurodegeneration remains unclear.In this study,rats were intraperitoneally injected with 5 mg/kg of the adenosine A1 receptor agonist N6-cyclopentyladenosine(CPA)for five weeks.The mobility of rats was evaluated by forced swimming test,while their cognitive capabilities were evaluated by Y-maze test.Expression of sortilin,α-synuclein,p-JUN,and c-JUN proteins in the substantia nigra were detected by western blot analysis.In addition,immunofluorescence staining of sortilin andα-synuclein was performed to detect expression in the substantia nigra.The results showed that,compared with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine(5 mg/kg)+CPA co-treated rats,motor and memory abilities were reduced,surface expression of sortin andα-synuclein in dopaminergic neurons was reduced,and total sortilin and totalα-synuclein were increased in CPA-treated rats.MN9D cells were incubated with 500 nM CPA alone or in combination with 10μM SP600125(JNK inhibitor)for 48 hours.Quantitative real-time polymerase chain reaction analysis of sortilin andα-synuclein mRNA levels in MN9D cells revealed upregulated sortilin expression in MN9D cells cultured with CPA alone,but the combination of CPA and SP600125 could inhibit this expression.Predictions made using Jasper,PROMO,and Alibaba online databases identified a highly conserved sequence in the sortilin promoter that was predicted to bind JUN in both humans and rodents.A luciferase reporter assay of sortilin promoter plasmid-transfected HEK293T cells confirmed this prediction.After sortilin expression was inhibited by sh-SORT1,expression of p-JUN and c-JUN was detected by western blot analysis.Long-term adenosine A1 receptor activation levels upregulatedα-synuclein expression at the post-transcriptional level by affecting sortilin expression.The online tool Raptor-X-Binding and Discovery Studio 4.5 prediction software predicted that sortilin can bind toα-synuclein.Co-immunoprecipitation revealed an interaction between sortilin andα-synuclein in MN9D cells.Our findings indicate that suppression of prolonged adenosine A1 receptor activation potently inhibited sortilin expression andα-synuclein accumulation,and dramatically improved host cognition and kineticism.This study was approved by the University Committee of Animal Care and Supply at the University of Saskatchewan(approval No.AUP#20070090)in March 2007 and the Animals Ethics Committee of University of South China(approval No.LL0387-USC)in June 2017.展开更多
Claudin 14 has been shown to promote nerve repair and regeneration in the early stages of Wallerian degeneration (0-4 days) in rats with sciatic nerve injury, but the mechanism underlying this process remains poorly...Claudin 14 has been shown to promote nerve repair and regeneration in the early stages of Wallerian degeneration (0-4 days) in rats with sciatic nerve injury, but the mechanism underlying this process remains poorly understood. This study reported the effects of claudin 14 on nerve degeneration and regeneration during early Wallerian degeneration. Claudin 14 expression was up-regulated in sciatic nerve 4 days after Wallerian degeneration. The altered expression of claudin 14 in Schwann cells resulted in expression changes of cytokines in vitro. Expression of claudin 14 affected c-Jun, but not Akt anal ERK1/2 patl^ways, l^urther studies reve^ed that enhanced expression of claudin 14 could promote Schwann cell proliferation and migration. Silencing of claudin 14 expression resulted in Schwann cell apoptosis and reduction in Schwann cell proliferation. Our data revealed the role of claudin 14 in early Wallerian degeneration, which may provide new insights into the molecular mechanisms of Wallerian degeneration.展开更多
The mitogen-activated protein kinase(MAPK) signaling pathway plays an important role in the regulation of cell growth, proliferation, differentiation, transformation and death. Mitogen-activated protein kinase phosp...The mitogen-activated protein kinase(MAPK) signaling pathway plays an important role in the regulation of cell growth, proliferation, differentiation, transformation and death. Mitogen-activated protein kinase phosphatase 1(MKP1) has an inhibitory effect on the p38 MAPK and JNK pathways, but it is unknown whether it plays a role in Aβ-induced oxidative stress and neuronal inflammation. In this study, PC12 cells were infected with MKP1 sh RNA, MKP1 lentivirus or control lentivirus for 12 hours, and then treated with 0.1, 1, 10 or 100 μM amyloid beta 42(Aβ42). The cell survival rate was measured using the cell counting kit-8 assay. MKP1, tumor necrosis factor-alpha(TNF-α) and interleukin-1β(IL-1β) m RNA expression levels were analyzed using quantitative real time-polymerase chain reaction. MKP1 and phospho-c-Jun N-terminal kinase(JNK) expression levels were assessed using western blot assay. Reactive oxygen species(ROS) levels were detected using 2′,7′-dichlorofluorescein diacetate. Mitochondrial membrane potential was measured using flow cytometry. Superoxide dismutase activity and malondialdehyde levels were evaluated using the colorimetric method. Lactate dehydrogenase activity was measured using a microplate reader. Caspase-3 expression levels were assessed by enzyme-linked immunosorbent assay. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase d UTP nick end labeling method. MKP1 overexpression inhibited Aβ-induced JNK phosphorylation and the increase in ROS levels. It also suppressed the Aβ-induced increase in TNF-α and IL-1β levels as well as apoptosis in PC12 cells. In contrast, MKP1 knockdown by RNA interference aggravated Aβ-induced oxidative stress, inflammation and cell damage in PC12 cells. Furthermore, the JNK-specific inhibitor SP600125 abolished this effect of MKP1 knockdown on Aβ-induced neurotoxicity. Collectively, these results show that MKP1 mitigates Aβ-induced apoptosis, oxidative stress and neuroinflammation by inhibiting the JNK signaling pathway, thereby playing a neuroprotective role.展开更多
HepatitisBvirus(HBV)infection causes acuteand chronic liver diseases,but is not directly cytopathic.Liver injury results fromrepeated attempts of the cellular immune response system to control the viral infection.Here...HepatitisBvirus(HBV)infection causes acuteand chronic liver diseases,but is not directly cytopathic.Liver injury results fromrepeated attempts of the cellular immune response system to control the viral infection.Here,we investigate the roles of cellular factors and signaling pathways involved in the regulation of HBV replication to reveal the mechanism underlying HBV infection and pathogenesis.Weshowthat collagen triple helix repeat containing 1(CTHRC1)expression is elevated in HBV-infected patients andin HBV-transfected cells through epigenetic modification and transcriptional regulation.CTHRC1 facilitates HBV replication in cultured cells and BALB/c mice by activating the PKCa/ERK/JNK/c-Jun cascade to repress the IFN/JAK/STAT pathway.HBV-activated CTHRC1 downregulates the activityof typeI interferon(IFN),theproductionof IFN-stimulatedgenes(ISGs),andthephosphorylationofsignal transducerandactivator of transcription 1/2(STAT1/2),whereas it upregulates the phosphorylation and ubiquitination of type I IFN receptors(IFNARa/b).Thus,our results showthat HBV uses a novelmechanismto hijack cellular factors and signal cascades in order to evade host antiviral immunity and maintain persistent infection.We also demonstrate that CTHRC1 has a novel role in viral infection.展开更多
文摘Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods:Human nasopharyngeal carcinoma CNE multicellular spheroids(MCS) were constructed with three dimensional cell culture methods.Western blot was employed to analyze the activity of JNK signaling pathway in MCS after X-ray irradiation,and the expression of caspase-3 protein before and after using SP600125(a special inhibitor of JNK).X-ray induced cell apoptosis in MCS before and after treated with SP600125 were detected by TUNEL.Results:The level of JNK phosphorylation in MCS was a dynamic course after radiation,and there was a phosphorylation peaks at 2 h later,the apoptotic rate of MCS(P < 0.05) and the expression of caspase-3 protein(P < 0.05) were significantly increased after treated with SP600125.Conclusion:The transient activation of JNK played a important role in sensitivity to radiotherapy of CNE MCS via mediating survival signals,blocking this pathway accelerate cell apoptosis,which may be related to the increased expression of caspase-3.
基金supported by the National Natural Sciences Foundation of China,No.81770460(to YCL)the Postdoctoral Research Fellowship of the Saskatchewan Health Research Foundation,No.SHRF,4144(to YCL)+2 种基金the third level of the Chuanshan Talent project of the University of South China,No.2017CST20(to YCL)the Aid Program,No.2017KJ268 and the Key Lab for Clinical Anatomy&Reproductive Medicine,No.2017KJ182 from the Science and Technology Bureau of Hengyang City,China(to YCL and XC)the Postgraduate Student Research Innovation Projects of Hunan Province,China,No.CX2018B62(to ABG)
文摘Prolonged activation of adenosine A1 receptor likely leads to damage of dopaminergic neurons and subsequent development of neurodegenerative diseases.However,the pathogenesis underlying long-term adenosine A1 receptor activation-induced neurodegeneration remains unclear.In this study,rats were intraperitoneally injected with 5 mg/kg of the adenosine A1 receptor agonist N6-cyclopentyladenosine(CPA)for five weeks.The mobility of rats was evaluated by forced swimming test,while their cognitive capabilities were evaluated by Y-maze test.Expression of sortilin,α-synuclein,p-JUN,and c-JUN proteins in the substantia nigra were detected by western blot analysis.In addition,immunofluorescence staining of sortilin andα-synuclein was performed to detect expression in the substantia nigra.The results showed that,compared with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine(5 mg/kg)+CPA co-treated rats,motor and memory abilities were reduced,surface expression of sortin andα-synuclein in dopaminergic neurons was reduced,and total sortilin and totalα-synuclein were increased in CPA-treated rats.MN9D cells were incubated with 500 nM CPA alone or in combination with 10μM SP600125(JNK inhibitor)for 48 hours.Quantitative real-time polymerase chain reaction analysis of sortilin andα-synuclein mRNA levels in MN9D cells revealed upregulated sortilin expression in MN9D cells cultured with CPA alone,but the combination of CPA and SP600125 could inhibit this expression.Predictions made using Jasper,PROMO,and Alibaba online databases identified a highly conserved sequence in the sortilin promoter that was predicted to bind JUN in both humans and rodents.A luciferase reporter assay of sortilin promoter plasmid-transfected HEK293T cells confirmed this prediction.After sortilin expression was inhibited by sh-SORT1,expression of p-JUN and c-JUN was detected by western blot analysis.Long-term adenosine A1 receptor activation levels upregulatedα-synuclein expression at the post-transcriptional level by affecting sortilin expression.The online tool Raptor-X-Binding and Discovery Studio 4.5 prediction software predicted that sortilin can bind toα-synuclein.Co-immunoprecipitation revealed an interaction between sortilin andα-synuclein in MN9D cells.Our findings indicate that suppression of prolonged adenosine A1 receptor activation potently inhibited sortilin expression andα-synuclein accumulation,and dramatically improved host cognition and kineticism.This study was approved by the University Committee of Animal Care and Supply at the University of Saskatchewan(approval No.AUP#20070090)in March 2007 and the Animals Ethics Committee of University of South China(approval No.LL0387-USC)in June 2017.
基金supported by grants from the National Natural Science Foundation of China,Grant No.81370982,31170946Key Program,Grant No.81130080+1 种基金the Scientific Research Foundation for Returned Scholars,Ministry of Education of Chinathe Priority Academic Program Development of Jiangsu Higher Education Institutions
文摘Claudin 14 has been shown to promote nerve repair and regeneration in the early stages of Wallerian degeneration (0-4 days) in rats with sciatic nerve injury, but the mechanism underlying this process remains poorly understood. This study reported the effects of claudin 14 on nerve degeneration and regeneration during early Wallerian degeneration. Claudin 14 expression was up-regulated in sciatic nerve 4 days after Wallerian degeneration. The altered expression of claudin 14 in Schwann cells resulted in expression changes of cytokines in vitro. Expression of claudin 14 affected c-Jun, but not Akt anal ERK1/2 patl^ways, l^urther studies reve^ed that enhanced expression of claudin 14 could promote Schwann cell proliferation and migration. Silencing of claudin 14 expression resulted in Schwann cell apoptosis and reduction in Schwann cell proliferation. Our data revealed the role of claudin 14 in early Wallerian degeneration, which may provide new insights into the molecular mechanisms of Wallerian degeneration.
文摘The mitogen-activated protein kinase(MAPK) signaling pathway plays an important role in the regulation of cell growth, proliferation, differentiation, transformation and death. Mitogen-activated protein kinase phosphatase 1(MKP1) has an inhibitory effect on the p38 MAPK and JNK pathways, but it is unknown whether it plays a role in Aβ-induced oxidative stress and neuronal inflammation. In this study, PC12 cells were infected with MKP1 sh RNA, MKP1 lentivirus or control lentivirus for 12 hours, and then treated with 0.1, 1, 10 or 100 μM amyloid beta 42(Aβ42). The cell survival rate was measured using the cell counting kit-8 assay. MKP1, tumor necrosis factor-alpha(TNF-α) and interleukin-1β(IL-1β) m RNA expression levels were analyzed using quantitative real time-polymerase chain reaction. MKP1 and phospho-c-Jun N-terminal kinase(JNK) expression levels were assessed using western blot assay. Reactive oxygen species(ROS) levels were detected using 2′,7′-dichlorofluorescein diacetate. Mitochondrial membrane potential was measured using flow cytometry. Superoxide dismutase activity and malondialdehyde levels were evaluated using the colorimetric method. Lactate dehydrogenase activity was measured using a microplate reader. Caspase-3 expression levels were assessed by enzyme-linked immunosorbent assay. Apoptosis was evaluated using the terminal deoxynucleotidyl transferase d UTP nick end labeling method. MKP1 overexpression inhibited Aβ-induced JNK phosphorylation and the increase in ROS levels. It also suppressed the Aβ-induced increase in TNF-α and IL-1β levels as well as apoptosis in PC12 cells. In contrast, MKP1 knockdown by RNA interference aggravated Aβ-induced oxidative stress, inflammation and cell damage in PC12 cells. Furthermore, the JNK-specific inhibitor SP600125 abolished this effect of MKP1 knockdown on Aβ-induced neurotoxicity. Collectively, these results show that MKP1 mitigates Aβ-induced apoptosis, oxidative stress and neuroinflammation by inhibiting the JNK signaling pathway, thereby playing a neuroprotective role.
基金supported by research grants from theMajor State Basic ResearchDevelopment Program(973 Program)(grant number 2012CB518900)the National Natural Science Foundation of China(grant numbers 31230005,31270206,31200134,and 81171525)+1 种基金the National Mega Project on Major Infectious Disease Prevention(grant numbers 2012ZX10002006-003 and 2012ZX10004-207)the Chinese Foundation for Hepatitis Prevention and Control(grant number CFHPC20132153).
文摘HepatitisBvirus(HBV)infection causes acuteand chronic liver diseases,but is not directly cytopathic.Liver injury results fromrepeated attempts of the cellular immune response system to control the viral infection.Here,we investigate the roles of cellular factors and signaling pathways involved in the regulation of HBV replication to reveal the mechanism underlying HBV infection and pathogenesis.Weshowthat collagen triple helix repeat containing 1(CTHRC1)expression is elevated in HBV-infected patients andin HBV-transfected cells through epigenetic modification and transcriptional regulation.CTHRC1 facilitates HBV replication in cultured cells and BALB/c mice by activating the PKCa/ERK/JNK/c-Jun cascade to repress the IFN/JAK/STAT pathway.HBV-activated CTHRC1 downregulates the activityof typeI interferon(IFN),theproductionof IFN-stimulatedgenes(ISGs),andthephosphorylationofsignal transducerandactivator of transcription 1/2(STAT1/2),whereas it upregulates the phosphorylation and ubiquitination of type I IFN receptors(IFNARa/b).Thus,our results showthat HBV uses a novelmechanismto hijack cellular factors and signal cascades in order to evade host antiviral immunity and maintain persistent infection.We also demonstrate that CTHRC1 has a novel role in viral infection.