[目的]探讨长链非编码RNA MIR4435-2HG(LncRNA MIR4435-2HG)靶向上调转化生长因子-β1(TGF-β1)对非小细胞肺癌(NSCLC)细胞迁移、增殖的作用及其初步机制。[方法]检测NSCLC癌组织及其癌旁组织、NSCLC细胞株及正常人支气管上皮细胞(HBE)...[目的]探讨长链非编码RNA MIR4435-2HG(LncRNA MIR4435-2HG)靶向上调转化生长因子-β1(TGF-β1)对非小细胞肺癌(NSCLC)细胞迁移、增殖的作用及其初步机制。[方法]检测NSCLC癌组织及其癌旁组织、NSCLC细胞株及正常人支气管上皮细胞(HBE)中MIR4435-2HG和TGF-β1表达。将pcDNA-MIR4435-2HG和pcDNA质粒转染至A549细胞,CCK-8法、Transwell法检测细胞增殖、迁移能力。Western Blot法检测细胞TGF-β1表达。免疫共沉淀验证MIR4435-2HG与TGF-β1靶向作用。[结果]NSCLC组织MIR4435-2HG(1.89±0.52 vs 0.76±0.48)和TGF-β1相对表达量(1.43±0.22 vs 0.37±0.18)均高于癌旁组织(P<0.05);与HBE细胞比较,NSCLC细胞株MIR4435-2HG相对表达量升高(P<0.05),且其中A549细胞高于H226细胞(P<0.05);与阴性对照组比较,MIR4435-2HG过表达组细胞MIR4435-2HG(4.72±0.43 vs 3.38±0.46)及TGF-β1(0.82±0.13 vs 1.07±0.24)相对表达量、细胞48 h OD值(0.43±0.03 vs 0.52±0.04)及迁移数量[(89.63±18.28)个vs(169.89±20.34)个]均增加(P<0.05),而与对照组比较,TGF-β1组细胞MIR4435-2HG表达未见显著变化(P>0.05)。[结论] NSCLC组织中MIR4435-2HG呈高表达,LncRNA MIR4435-2HG可能通过靶向上调TGF-β1促进NSCLC细胞迁移和增殖。展开更多
目的探讨LncRNA MIR4435-2HG对胃癌细胞增殖、侵袭和迁移的影响及作用机制。方法培养正常人胃黏膜上皮细胞GES-1和胃癌细胞系AGS、SGC7901、BGC823和BGC803,AGS细胞分为对照组(正常培养细胞)、si-con组(转染乱序无意义阴性序列)、si-MIR...目的探讨LncRNA MIR4435-2HG对胃癌细胞增殖、侵袭和迁移的影响及作用机制。方法培养正常人胃黏膜上皮细胞GES-1和胃癌细胞系AGS、SGC7901、BGC823和BGC803,AGS细胞分为对照组(正常培养细胞)、si-con组(转染乱序无意义阴性序列)、si-MIR4435-2HG组(转染MIR4435-2HG小干扰RNA)、miR-con组(转染模拟物对照序列)、miR-138-5p组(转染miR-138-5p模拟物)、si-HMGA1组(转染HMGA1小干扰RNA)、si-MIR4435-2HG+pcDNA组(共转染MIR4435-2HG小干扰RNA与空载体)和si-MIR4435-2HG+pcDNA-HMGA1组(共转染MIR4435-2HG小干扰RNA与HMGA1过表达载体)。qRT-PCR检测细胞中MIR4435-2HG和高迁移率族蛋白A1(HMGA1)mRNA表达,Western blot检测细胞中HMGA1蛋白表达。双荧光素酶实验验证AGS细胞中MIR4435-2HG与miR-138-5p以及miR-138-5p与HMGA1之间关系。MTT检测细胞增殖,Transwell检测细胞迁移和侵袭,Western blot检测Ki67、CyclinD1、MMP2和MMP9蛋白表达。裸鼠分为对照组(接种正常培养的AGS细胞)、sh-MIR4435-2HG组(接种转染携带MIR4435-2HG基因短发夹RNA慢病毒的AGS细胞)和sh-con组(接种转染携带无关序列片段shRNA慢病毒的AGS细胞),接种21 d后测量肿瘤重量,观察MIR4435-2HG对裸鼠移植瘤生长的影响。两组间比较采用独立样本t检验,多组间比较采用单因素方差分析,组间两两比较采用SNK-q检验。结果与GES-1细胞比较,胃癌细胞AGS、SGC7901、BGC823、BGC803中MIR4435-2HG表达水平升高,HMGA1 mRNA和蛋白表达水平升高(P均<0.05),选择AGS细胞进行后续实验。MIR4435-2HG靶向负调控miR-138-5p表达,miR-138-5p靶向负调控HMGA1表达。与si-con组比较,si-MIR4435-2HG组AGS细胞48 h OD值(1.13±0.12比0.86±0.09)、迁移数量[(179.23±18.01)个比(60.15±6.05)个]、侵袭数量[(81.26±8.16)个比(25.64±2.59)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与miR-con组比较,miR-138-5p组AGS细胞48 h OD值(1.28±0.13比0.85±0.09)、迁移数量[(178.26±17.59)个比(69.35±6.34)个]、侵袭数量[(81.02±8.06)个比(36.76±2.49)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与si-con组比较,si-HMGA1组AGS细胞48 h OD值(1.28±0.13比0.83±0.09)、迁移数量[(175.62±17.59)个比(63.26±6.34)个]、侵袭数量[(80.16±8.06)个比(24.56±2.49)个]、Ki67蛋白表达水平(0.84±0.09比0.30±0.03),CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与si-MIR4435-2HG+pcDNA组比较,si-MIR4435-2HG+pcDNA-HMGA1组AGS细胞48 h OD值(0.80±0.09比1.01±0.11)、迁移数量[(60.45±5.79)个比(119.25±12.46)个]、侵袭数量[(23.46±2.34)个比(59.46±6.29)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均升高(P均<0.05)。与sh-con组比较,sh-MIR4435-2HG组裸鼠体内移植瘤重量[(0.40±0.03)g比(0.13±0.03)g]降低(P<0.05)。结论MIR4435-2HG在胃癌细胞系中高表达,沉默MIR4435-2HG表达可能通过调控miR-138-5p/HMGA1轴抑制胃癌细胞体外增殖、迁移和侵袭,并抑制体内肿瘤生长。展开更多
AIM:To explore the effect of co-host non-coding RNA(ncRNA)MIR503HG/miR-503-5p on the angiogenesis of pterygium.METHODS:MIR503HG/miR-503-5p/fibroblast growth factor 2(FGF2)expression levels in pterygium tissues,control...AIM:To explore the effect of co-host non-coding RNA(ncRNA)MIR503HG/miR-503-5p on the angiogenesis of pterygium.METHODS:MIR503HG/miR-503-5p/fibroblast growth factor 2(FGF2)expression levels in pterygium tissues,control conjunctival tissues,and human pterygium fibroblasts(HPF)were examined by reverse transcription-polymerase chain reaction(qRT-PCR)and immunohistochemical methods.Effects of MIR503HG/miR-503-5p on low molecular weight FGF2(LWM FGF2),migration and angiogenesis of human retinal microvascular endothelial cells(HRMEC)were determined in an HPF and HRMEC co-culture model using Western blots,wound healing assay,Matrigel-based tube formation assay,and Transwell assay.RESULTS:MIR503HG/miR-503-5p/FGF2 pathway was actively increased in pterygium tissue and there was a negative correlation between the expression of the two ncRNAs.FGF2 expression level was positively correlated with MIR503HG and negatively correlated with miR-503-5p.Overexpressed MIR503HG/miR-503-5p did not affect the migration and angiogenesis of HRMECs cultured separately,but significantly affected migration and angiogenesis of HRMEC in HPF and HRMEC co-culture models.Western blotting revealed that MIR503HG/miR-503-5p overexpression significantly increased LMW FGF2 expression in HPF.CONCLUSION:MIR503HG/miR-503-5p inhibits HRMEC migration and angiogenic function by interfering with the interaction between HPF and endothelial cells via reducing LMW FGF2 in HPF.展开更多
文摘[目的]探讨长链非编码RNA MIR4435-2HG(LncRNA MIR4435-2HG)靶向上调转化生长因子-β1(TGF-β1)对非小细胞肺癌(NSCLC)细胞迁移、增殖的作用及其初步机制。[方法]检测NSCLC癌组织及其癌旁组织、NSCLC细胞株及正常人支气管上皮细胞(HBE)中MIR4435-2HG和TGF-β1表达。将pcDNA-MIR4435-2HG和pcDNA质粒转染至A549细胞,CCK-8法、Transwell法检测细胞增殖、迁移能力。Western Blot法检测细胞TGF-β1表达。免疫共沉淀验证MIR4435-2HG与TGF-β1靶向作用。[结果]NSCLC组织MIR4435-2HG(1.89±0.52 vs 0.76±0.48)和TGF-β1相对表达量(1.43±0.22 vs 0.37±0.18)均高于癌旁组织(P<0.05);与HBE细胞比较,NSCLC细胞株MIR4435-2HG相对表达量升高(P<0.05),且其中A549细胞高于H226细胞(P<0.05);与阴性对照组比较,MIR4435-2HG过表达组细胞MIR4435-2HG(4.72±0.43 vs 3.38±0.46)及TGF-β1(0.82±0.13 vs 1.07±0.24)相对表达量、细胞48 h OD值(0.43±0.03 vs 0.52±0.04)及迁移数量[(89.63±18.28)个vs(169.89±20.34)个]均增加(P<0.05),而与对照组比较,TGF-β1组细胞MIR4435-2HG表达未见显著变化(P>0.05)。[结论] NSCLC组织中MIR4435-2HG呈高表达,LncRNA MIR4435-2HG可能通过靶向上调TGF-β1促进NSCLC细胞迁移和增殖。
文摘目的探讨LncRNA MIR4435-2HG对胃癌细胞增殖、侵袭和迁移的影响及作用机制。方法培养正常人胃黏膜上皮细胞GES-1和胃癌细胞系AGS、SGC7901、BGC823和BGC803,AGS细胞分为对照组(正常培养细胞)、si-con组(转染乱序无意义阴性序列)、si-MIR4435-2HG组(转染MIR4435-2HG小干扰RNA)、miR-con组(转染模拟物对照序列)、miR-138-5p组(转染miR-138-5p模拟物)、si-HMGA1组(转染HMGA1小干扰RNA)、si-MIR4435-2HG+pcDNA组(共转染MIR4435-2HG小干扰RNA与空载体)和si-MIR4435-2HG+pcDNA-HMGA1组(共转染MIR4435-2HG小干扰RNA与HMGA1过表达载体)。qRT-PCR检测细胞中MIR4435-2HG和高迁移率族蛋白A1(HMGA1)mRNA表达,Western blot检测细胞中HMGA1蛋白表达。双荧光素酶实验验证AGS细胞中MIR4435-2HG与miR-138-5p以及miR-138-5p与HMGA1之间关系。MTT检测细胞增殖,Transwell检测细胞迁移和侵袭,Western blot检测Ki67、CyclinD1、MMP2和MMP9蛋白表达。裸鼠分为对照组(接种正常培养的AGS细胞)、sh-MIR4435-2HG组(接种转染携带MIR4435-2HG基因短发夹RNA慢病毒的AGS细胞)和sh-con组(接种转染携带无关序列片段shRNA慢病毒的AGS细胞),接种21 d后测量肿瘤重量,观察MIR4435-2HG对裸鼠移植瘤生长的影响。两组间比较采用独立样本t检验,多组间比较采用单因素方差分析,组间两两比较采用SNK-q检验。结果与GES-1细胞比较,胃癌细胞AGS、SGC7901、BGC823、BGC803中MIR4435-2HG表达水平升高,HMGA1 mRNA和蛋白表达水平升高(P均<0.05),选择AGS细胞进行后续实验。MIR4435-2HG靶向负调控miR-138-5p表达,miR-138-5p靶向负调控HMGA1表达。与si-con组比较,si-MIR4435-2HG组AGS细胞48 h OD值(1.13±0.12比0.86±0.09)、迁移数量[(179.23±18.01)个比(60.15±6.05)个]、侵袭数量[(81.26±8.16)个比(25.64±2.59)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与miR-con组比较,miR-138-5p组AGS细胞48 h OD值(1.28±0.13比0.85±0.09)、迁移数量[(178.26±17.59)个比(69.35±6.34)个]、侵袭数量[(81.02±8.06)个比(36.76±2.49)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与si-con组比较,si-HMGA1组AGS细胞48 h OD值(1.28±0.13比0.83±0.09)、迁移数量[(175.62±17.59)个比(63.26±6.34)个]、侵袭数量[(80.16±8.06)个比(24.56±2.49)个]、Ki67蛋白表达水平(0.84±0.09比0.30±0.03),CyclinD1、MMP2和MMP9蛋白表达水平均降低(P均<0.05)。与si-MIR4435-2HG+pcDNA组比较,si-MIR4435-2HG+pcDNA-HMGA1组AGS细胞48 h OD值(0.80±0.09比1.01±0.11)、迁移数量[(60.45±5.79)个比(119.25±12.46)个]、侵袭数量[(23.46±2.34)个比(59.46±6.29)个]、Ki67、CyclinD1、MMP2和MMP9蛋白表达水平均升高(P均<0.05)。与sh-con组比较,sh-MIR4435-2HG组裸鼠体内移植瘤重量[(0.40±0.03)g比(0.13±0.03)g]降低(P<0.05)。结论MIR4435-2HG在胃癌细胞系中高表达,沉默MIR4435-2HG表达可能通过调控miR-138-5p/HMGA1轴抑制胃癌细胞体外增殖、迁移和侵袭,并抑制体内肿瘤生长。
基金Supported by the National Natural Science Foundation of China(No.81770898).
文摘AIM:To explore the effect of co-host non-coding RNA(ncRNA)MIR503HG/miR-503-5p on the angiogenesis of pterygium.METHODS:MIR503HG/miR-503-5p/fibroblast growth factor 2(FGF2)expression levels in pterygium tissues,control conjunctival tissues,and human pterygium fibroblasts(HPF)were examined by reverse transcription-polymerase chain reaction(qRT-PCR)and immunohistochemical methods.Effects of MIR503HG/miR-503-5p on low molecular weight FGF2(LWM FGF2),migration and angiogenesis of human retinal microvascular endothelial cells(HRMEC)were determined in an HPF and HRMEC co-culture model using Western blots,wound healing assay,Matrigel-based tube formation assay,and Transwell assay.RESULTS:MIR503HG/miR-503-5p/FGF2 pathway was actively increased in pterygium tissue and there was a negative correlation between the expression of the two ncRNAs.FGF2 expression level was positively correlated with MIR503HG and negatively correlated with miR-503-5p.Overexpressed MIR503HG/miR-503-5p did not affect the migration and angiogenesis of HRMECs cultured separately,but significantly affected migration and angiogenesis of HRMEC in HPF and HRMEC co-culture models.Western blotting revealed that MIR503HG/miR-503-5p overexpression significantly increased LMW FGF2 expression in HPF.CONCLUSION:MIR503HG/miR-503-5p inhibits HRMEC migration and angiogenic function by interfering with the interaction between HPF and endothelial cells via reducing LMW FGF2 in HPF.
