目的探讨环状RNA(CircRNA)-CDR1as靶向miR-7-5p/DNA聚合酶ε亚基4(POLE4)对非小细胞肺癌(NSCLC)细胞恶性生物学行为的影响。方法收集2023年1月-2024年6月于儋州市人民医院接受手术的30例NSCLC患者的肿瘤组织和癌旁组织,检测CircRNA-CDR...目的探讨环状RNA(CircRNA)-CDR1as靶向miR-7-5p/DNA聚合酶ε亚基4(POLE4)对非小细胞肺癌(NSCLC)细胞恶性生物学行为的影响。方法收集2023年1月-2024年6月于儋州市人民医院接受手术的30例NSCLC患者的肿瘤组织和癌旁组织,检测CircRNA-CDR1as、miR-7-5p、POLE4在肿瘤组织、癌旁组织及NSCLC细胞(A549、PC9、NCI-H1299、NCI-H1650)、正常肺上皮细胞(BEAS-2B)中的表达水平;将A549细胞分为对照组、sh-CircRNA-CDR1as组、阴性对照(sh-NC)组、sh-CircRNA-CDR1as+miR-7-5p抑制剂(in-miR-7-5p)组、sh-CircRNACDR1as+miR抑制剂(in-miR-NC)组,细胞计数试剂盒-8(CCK-8)检测细胞增殖能力,Transwell实验检测细胞迁移、侵袭能力,流式细胞术检测细胞凋亡,Western blot检测基质金属蛋白酶-9(MMP-9)、程序性死亡配体-1(PD-L1)、抗BCL2相关x蛋白(BAX)、POLE4、增殖细胞核抗原(PCNA)蛋白表达;实时荧光定量PCR检测CircRNA-CDR1as、miR-7-5p、POLE4基因表达;并验证miR-7-5p与CircRNA-CDR1as或POLE4靶向关系。结果肿瘤组织中CircRNACDR1as、POLE4表达水平较癌旁组织显著升高,miR-7-5p表达水平较癌旁组织显著降低,差异均有统计学意义(t=16.877、21.546、16.383,P均<0.05);A549、PC9、NCI-H1299、NCI-H1650细胞中CircRNA-CDR1as、POLE4表达水平较BEAS-2B细胞显著升高,miR-7-5p表达水平较BEAS-2B细胞显著降低,差异均有统计学意义(F=32.878、33.836、64.688,P均<0.05)。与对照组、sh-NC组比较,sh-CircRNA-CDR1as组细胞凋亡、BAX表达、miR-7-5p表达显著增加,细胞24和48 h OD450值、PCNA表达、侵袭、迁移及MMP-9、PD-L1表达、CircRNA-CDR1as、POLE4蛋白及mRNA表达显著降低,差异均有统计学意义(P均<0.05);与sh-CircRNA-CDR1as+in-miR-NC组比较,sh-CircRNA-CDR1as+in-miR-7-5p组细胞凋亡、BAX表达、miR-7-5p表达显著降低,细胞24和48 h OD450值、PCNA表达、侵袭、迁移及MMP-9、PD-L1表达、POLE4蛋白及mRNA表达显著增加,差异均有统计学意义(P均<0.05);各组细胞凋亡、BAX、miR-7-5p表达、细胞24和48 h OD450值、PCNA表达、侵袭、迁移、MMP-9、PD-L1、CircRNA-CDR1as、POLE4蛋白及mRNA表达比较差异均有统计学意义(F=149.200、95.529、33.807、29.678、28.905、71.011、43.873、50.978、59.597、46.174、103.372、54.760、45.096,P均<0.05)。CircRNA-CDR1as-WT、POLE4-WT分别与miR-7-5p或miR-NC共转染,两组间荧光素酶活性差异均有统计学意义(P均<0.05)。结论干扰CircRNA-CDR1as靶向miR-7-5p/POLE4可对NSCLC细胞恶性生物学行为发挥抑制作用。展开更多
Ischemia–reperfusion injury is a common pathophysiological mechanism in retinal degeneration.PANoptosis is a newly defined integral form of regulated cell death that combines the key features of pyroptosis,apoptosis,...Ischemia–reperfusion injury is a common pathophysiological mechanism in retinal degeneration.PANoptosis is a newly defined integral form of regulated cell death that combines the key features of pyroptosis,apoptosis,and necroptosis.Oligomerization of mitochondrial voltage-dependent anion channel 1 is an important pathological event in regulating cell death in retinal ischemia–reperfusion injury.However,its role in PANoptosis remains largely unknown.In this study,we demonstrated that voltage-dependent anion channel 1 oligomerization-mediated mitochondrial dysfunction was associated with PANoptosis in retinal ischemia–reperfusion injury.Inhibition of voltage-dependent anion channel 1 oligomerization suppressed mitochondrial dysfunction and PANoptosis in retinal cells subjected to ischemia–reperfusion injury.Mechanistically,mitochondria-derived reactive oxygen species played a central role in the voltagedependent anion channel 1-mediated regulation of PANoptosis by promoting PANoptosome assembly.Moreover,inhibiting voltage-dependent anion channel 1 oligomerization protected against PANoptosis in the retinas of rats subjected to ischemia–reperfusion injury.Overall,our findings reveal the critical role of voltage-dependent anion channel 1 oligomerization in regulating PANoptosis in retinal ischemia–reperfusion injury,highlighting voltage-dependent anion channel 1 as a promising therapeutic target.展开更多
文摘目的探讨环状RNA(CircRNA)-CDR1as靶向miR-7-5p/DNA聚合酶ε亚基4(POLE4)对非小细胞肺癌(NSCLC)细胞恶性生物学行为的影响。方法收集2023年1月-2024年6月于儋州市人民医院接受手术的30例NSCLC患者的肿瘤组织和癌旁组织,检测CircRNA-CDR1as、miR-7-5p、POLE4在肿瘤组织、癌旁组织及NSCLC细胞(A549、PC9、NCI-H1299、NCI-H1650)、正常肺上皮细胞(BEAS-2B)中的表达水平;将A549细胞分为对照组、sh-CircRNA-CDR1as组、阴性对照(sh-NC)组、sh-CircRNA-CDR1as+miR-7-5p抑制剂(in-miR-7-5p)组、sh-CircRNACDR1as+miR抑制剂(in-miR-NC)组,细胞计数试剂盒-8(CCK-8)检测细胞增殖能力,Transwell实验检测细胞迁移、侵袭能力,流式细胞术检测细胞凋亡,Western blot检测基质金属蛋白酶-9(MMP-9)、程序性死亡配体-1(PD-L1)、抗BCL2相关x蛋白(BAX)、POLE4、增殖细胞核抗原(PCNA)蛋白表达;实时荧光定量PCR检测CircRNA-CDR1as、miR-7-5p、POLE4基因表达;并验证miR-7-5p与CircRNA-CDR1as或POLE4靶向关系。结果肿瘤组织中CircRNACDR1as、POLE4表达水平较癌旁组织显著升高,miR-7-5p表达水平较癌旁组织显著降低,差异均有统计学意义(t=16.877、21.546、16.383,P均<0.05);A549、PC9、NCI-H1299、NCI-H1650细胞中CircRNA-CDR1as、POLE4表达水平较BEAS-2B细胞显著升高,miR-7-5p表达水平较BEAS-2B细胞显著降低,差异均有统计学意义(F=32.878、33.836、64.688,P均<0.05)。与对照组、sh-NC组比较,sh-CircRNA-CDR1as组细胞凋亡、BAX表达、miR-7-5p表达显著增加,细胞24和48 h OD450值、PCNA表达、侵袭、迁移及MMP-9、PD-L1表达、CircRNA-CDR1as、POLE4蛋白及mRNA表达显著降低,差异均有统计学意义(P均<0.05);与sh-CircRNA-CDR1as+in-miR-NC组比较,sh-CircRNA-CDR1as+in-miR-7-5p组细胞凋亡、BAX表达、miR-7-5p表达显著降低,细胞24和48 h OD450值、PCNA表达、侵袭、迁移及MMP-9、PD-L1表达、POLE4蛋白及mRNA表达显著增加,差异均有统计学意义(P均<0.05);各组细胞凋亡、BAX、miR-7-5p表达、细胞24和48 h OD450值、PCNA表达、侵袭、迁移、MMP-9、PD-L1、CircRNA-CDR1as、POLE4蛋白及mRNA表达比较差异均有统计学意义(F=149.200、95.529、33.807、29.678、28.905、71.011、43.873、50.978、59.597、46.174、103.372、54.760、45.096,P均<0.05)。CircRNA-CDR1as-WT、POLE4-WT分别与miR-7-5p或miR-NC共转染,两组间荧光素酶活性差异均有统计学意义(P均<0.05)。结论干扰CircRNA-CDR1as靶向miR-7-5p/POLE4可对NSCLC细胞恶性生物学行为发挥抑制作用。
基金supported by the National Natural Science Foundation of China,Nos.82172196(to KX),82372507(to KX)the Natural Science Foundation of Hunan Province,China,No.2023JJ40804(to QZ)the Key Laboratory of Emergency and Trauma(Hainan Medical University)of the Ministry of Education,China,No.KLET-202210(to QZ)。
文摘Ischemia–reperfusion injury is a common pathophysiological mechanism in retinal degeneration.PANoptosis is a newly defined integral form of regulated cell death that combines the key features of pyroptosis,apoptosis,and necroptosis.Oligomerization of mitochondrial voltage-dependent anion channel 1 is an important pathological event in regulating cell death in retinal ischemia–reperfusion injury.However,its role in PANoptosis remains largely unknown.In this study,we demonstrated that voltage-dependent anion channel 1 oligomerization-mediated mitochondrial dysfunction was associated with PANoptosis in retinal ischemia–reperfusion injury.Inhibition of voltage-dependent anion channel 1 oligomerization suppressed mitochondrial dysfunction and PANoptosis in retinal cells subjected to ischemia–reperfusion injury.Mechanistically,mitochondria-derived reactive oxygen species played a central role in the voltagedependent anion channel 1-mediated regulation of PANoptosis by promoting PANoptosome assembly.Moreover,inhibiting voltage-dependent anion channel 1 oligomerization protected against PANoptosis in the retinas of rats subjected to ischemia–reperfusion injury.Overall,our findings reveal the critical role of voltage-dependent anion channel 1 oligomerization in regulating PANoptosis in retinal ischemia–reperfusion injury,highlighting voltage-dependent anion channel 1 as a promising therapeutic target.
