Objective:Breast cancer is the most common malignancy in women and is characterized by a high recurrence rate that severely impacts patient survival.Regulatory T cells(Tregs)in the tumor microenvironment(TME)promote i...Objective:Breast cancer is the most common malignancy in women and is characterized by a high recurrence rate that severely impacts patient survival.Regulatory T cells(Tregs)in the tumor microenvironment(TME)promote immune evasion and metastasis,increasing recurrence risk.This study determined how the epigenetic regulators,DNMT3A and METTL7A,modulate Treg infiltration via the DDR1/STAT3/CXCL5 axis and influence breast cancer recurrence and prognosis.Methods:RNA sequencing(RNA-seq)was used to identify differentially expressed genes(DEGs),followed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment.Machine learning algorithms,including least absolute shrinkage and selection operator(LASSO),supported vector machine-recursive feature elimination(SVM-RFE)and ElasticNet identified DDR1 as a key gene.Validation included RT-qPCR,western blot,MSP,MeRIP-qPCR,and Co-IP to assess epigenetic regulation.Functional assays(CCK-8,Transwell,and Treg differentiation/chemotaxis)and xenograft models evaluated the role of DDR1 in tumor progression and recurrence.Results:DNMT3A upregulated DDR1 via DNA methylation,while METTL7A enhanced DDR1 mRNA stability via m6A modification.Co-regulation activated the DDR1/STAT3/CXCL5 axis,which boosted cancer cell proliferation,migration,and invasion.CXCL5 secretion increased Treg infiltration and accelerated tumor growth in vivo.DDR1 silencing reversed these effects,confirming that DDR1 has a pivotal role in breast cancer recurrence.Conclusion:DNMT3A and METTL7A were shown to cooperatively regulate DDR1 via DNA/m6A methylation,which drives Tregmediated immune suppression and recurrence.This study provided novel insights and therapeutic targets for breast cancer prognosis and treatment.展开更多
目的探讨星状神经节阻滞(SGB)通过长链非编码RNA(lncRNA)牛磺酸上调基因1(TUG1)-NOD样受体热蛋白结构域相关蛋白3(NLRP3)轴在体外脑缺血再灌注模型中对炎症反应和自噬溶酶体形成的调节作用。方法培养大鼠海马神经元细胞系H19-7,并将细...目的探讨星状神经节阻滞(SGB)通过长链非编码RNA(lncRNA)牛磺酸上调基因1(TUG1)-NOD样受体热蛋白结构域相关蛋白3(NLRP3)轴在体外脑缺血再灌注模型中对炎症反应和自噬溶酶体形成的调节作用。方法培养大鼠海马神经元细胞系H19-7,并将细胞分为8组:(i)正常对照组:正常培养的神经元细胞;(ii)氧-糖剥夺/复氧(OGD/R)组:采用氧-糖剥夺/复氧法模拟脑缺血再灌注损伤;(iii)OGD/R+SGB组:OGD/R联合麻醉药0.5%布比卡因用于体外模拟SGB;(iv)OGD/R+SGB+TUG1过表达阴性对照组:OGD/R联合布比卡因并联合TUG1过表达阴性对照质粒转染细胞;(v)OGD/R+SGB+TUG1过表达组:OGD/R联合布比卡因并联合TUG1过表达质粒转染细胞;(vi)OGD/R+SGB+TUG1过表达+MCC950组:OGD/R联合布比卡因、TUG1过表达质粒转染及NLRP3抑制剂MCC950处理细胞;(vii)OGD/R+TUG1过表达组:OGD/R联合TUG1过表达质粒转染细胞;(viii)OGD/R+MCC950组:OGD/R联合NLRP3抑制剂MCC950处理细胞。进一步通过实时定量PCR(Quantitative Real Time PCR,qRTPCR)实验检测细胞中lncRNATUG1的表达;利用Western blot法检测细胞中NLRP3、微管相关蛋白1轻链3(LC3)-I、LC3-II、自噬相关基因5(Atg5)、苄氯素1(beclin1)、自噬接头蛋白(p62)、溶酶体相关膜蛋白1(LAMP1)的表达水平;利用透射电镜(TEM)检测自噬溶酶体的数量;并用酶联免疫吸附测定(ELISA)法检测细胞培养上清中白细胞介素(IL)-1β、IL-6、IL-18、肿瘤坏死因子(TNF)-α的含量。结果与正常对照组相比,OGD/R组lncRNA TUG1、NLRP3、Atg5、beclin1、p62、LAMP1的表达水平以及LC3-II/I比值均显著上调(^(均)P<0.05),自噬溶酶体数量增加(P<0.05),IL-1β、IL-6、IL-18、TNF-α的含量显著升高(^(均)P<0.05)。与OGD/R组相比,OGD/R+SGB组的上述指标均显著下调(P<0.05)。与OGD/R+SGB+TUG1过表达阴性对照组相比,OGD/R+SGB+TUG1过表达组的lncRNA TUG1、NLRP3、Atg5、beclin1、p62、LAMP1的表达水平以及LC3-II/I比值均显著上调(^(均)P<0.05),自噬溶酶体数量增加(P<0.05),IL-1β、IL-6、IL-18、TNF-α的含量显著升高(^(均)P<0.05),然而,加入NLRP3的抑制剂MCC950后,除lncRNA TUG1外其余指标均显著下调(^(均)P<0.05)。另外,与OGD/R组比,OGD/R+TUG1过表达组的上述指标进一步上调(^(均)P<0.05)。与OGD/R组比,OGD/R+MCC950组则抑制了除lncRNA TUG1外的其余指标(^(均)P<0.05)。结论星状神经节阻滞通过调节lncRNA TUG1-NLRP3轴有效减轻体外脑缺血再灌注损伤引起的炎症反应和自噬溶酶体形成,提示其可能作为治疗缺血性脑损伤的潜在策略。展开更多
目的研究分析维持性血液透析患者血清巨噬细胞炎性蛋白-1α(macrophage inflammatory protein-1α,MIP-1α)、几丁质酶-3样蛋白1(chitinase-3-like protein 1,CHI3L1/YKL-40)、含pyrin结构域的NOD样受体3(NOD like receptor family pyri...目的研究分析维持性血液透析患者血清巨噬细胞炎性蛋白-1α(macrophage inflammatory protein-1α,MIP-1α)、几丁质酶-3样蛋白1(chitinase-3-like protein 1,CHI3L1/YKL-40)、含pyrin结构域的NOD样受体3(NOD like receptor family pyrin domain-containing protein 3,NLRP3)水平与导管相关性感染及预后的相关性。方法选取泸州市中医医院在2020年1月─2024年8月收治的维持性血液透析患者296例,根据是否发生导管感染分为感染组(n=68)和未感染组(n=228),根据预后情况分为预后良好组(n=42)和预后不良组(n=26),经ELISA检测血清MIP-1α、YKL-40、NLRP3水平,收集分析一般临床资料,多因素Logistic回归分析患者预后影响因素,受试者工作特征(receiver operating characteristic,ROC)曲线分析MIP-1α、YKL-40、NLRP3对患者预后的预测价值,Pearson及Spearman法分析MIP-1α、YKL-40、NLRP3与炎症因子及预后的相关性。结果与非感染组相比,感染组的血清MIP-1α、YKL-40、NLRP3、C反应蛋白(C-reactive protein,CRP)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、降钙素原(procalcitonin,PCT)水平显著升高(t=14.532、14.561、22.823、28.495、11.487、22.234,均P<0.001);预后不良组的透析龄、透析频率、MIP-1α、YKL-40、NLRP3、CRP、TNF-α、PCT水平显著高于预后良好组(t=3.293、3.293、8.410、7.432、7.511、15.045、3.694、4.007,P=0.002、0.001、<0.001、<0.001、<0.001、<0.001、<0.001、<0.001);ROC曲线显示MIP-1α、YKL-40、NLRP3三者联合对患者预后预测的AUC显著高于MIP-1α(Z=2.101,P=0.036)、YKL-40(Z=2.610,P=0.009)、NLRP3(Z=2.965,P=0.003)单独预测;MIP-1α、YKL-40、NLRP3与炎症因子CRP(r=0.462、0.466、0.472,均P<0.001)、TNF-α(r=0.436、0.426、0.451,均P<0.001)、PCT(r=0.450、0.444、0.455,均P<0.001)以及预后(r=0.521、0.502、0.504,P<0.001)均呈正相关。结论血清MIP-1α、YKL-40、NLRP3与维持性血液透析患者的导管相关性感染及预后具有一定相关性。展开更多
基金supported by the National Natural Science Foundation of China(Grant No.82060479)Key Research and Development Program of Ningxia Hui Autonomous Region(Grant No.2021BEG03062)Ningxia Natural Science Fund Key Project(Grant No.2024AAC02080).
文摘Objective:Breast cancer is the most common malignancy in women and is characterized by a high recurrence rate that severely impacts patient survival.Regulatory T cells(Tregs)in the tumor microenvironment(TME)promote immune evasion and metastasis,increasing recurrence risk.This study determined how the epigenetic regulators,DNMT3A and METTL7A,modulate Treg infiltration via the DDR1/STAT3/CXCL5 axis and influence breast cancer recurrence and prognosis.Methods:RNA sequencing(RNA-seq)was used to identify differentially expressed genes(DEGs),followed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment.Machine learning algorithms,including least absolute shrinkage and selection operator(LASSO),supported vector machine-recursive feature elimination(SVM-RFE)and ElasticNet identified DDR1 as a key gene.Validation included RT-qPCR,western blot,MSP,MeRIP-qPCR,and Co-IP to assess epigenetic regulation.Functional assays(CCK-8,Transwell,and Treg differentiation/chemotaxis)and xenograft models evaluated the role of DDR1 in tumor progression and recurrence.Results:DNMT3A upregulated DDR1 via DNA methylation,while METTL7A enhanced DDR1 mRNA stability via m6A modification.Co-regulation activated the DDR1/STAT3/CXCL5 axis,which boosted cancer cell proliferation,migration,and invasion.CXCL5 secretion increased Treg infiltration and accelerated tumor growth in vivo.DDR1 silencing reversed these effects,confirming that DDR1 has a pivotal role in breast cancer recurrence.Conclusion:DNMT3A and METTL7A were shown to cooperatively regulate DDR1 via DNA/m6A methylation,which drives Tregmediated immune suppression and recurrence.This study provided novel insights and therapeutic targets for breast cancer prognosis and treatment.
文摘目的探讨星状神经节阻滞(SGB)通过长链非编码RNA(lncRNA)牛磺酸上调基因1(TUG1)-NOD样受体热蛋白结构域相关蛋白3(NLRP3)轴在体外脑缺血再灌注模型中对炎症反应和自噬溶酶体形成的调节作用。方法培养大鼠海马神经元细胞系H19-7,并将细胞分为8组:(i)正常对照组:正常培养的神经元细胞;(ii)氧-糖剥夺/复氧(OGD/R)组:采用氧-糖剥夺/复氧法模拟脑缺血再灌注损伤;(iii)OGD/R+SGB组:OGD/R联合麻醉药0.5%布比卡因用于体外模拟SGB;(iv)OGD/R+SGB+TUG1过表达阴性对照组:OGD/R联合布比卡因并联合TUG1过表达阴性对照质粒转染细胞;(v)OGD/R+SGB+TUG1过表达组:OGD/R联合布比卡因并联合TUG1过表达质粒转染细胞;(vi)OGD/R+SGB+TUG1过表达+MCC950组:OGD/R联合布比卡因、TUG1过表达质粒转染及NLRP3抑制剂MCC950处理细胞;(vii)OGD/R+TUG1过表达组:OGD/R联合TUG1过表达质粒转染细胞;(viii)OGD/R+MCC950组:OGD/R联合NLRP3抑制剂MCC950处理细胞。进一步通过实时定量PCR(Quantitative Real Time PCR,qRTPCR)实验检测细胞中lncRNATUG1的表达;利用Western blot法检测细胞中NLRP3、微管相关蛋白1轻链3(LC3)-I、LC3-II、自噬相关基因5(Atg5)、苄氯素1(beclin1)、自噬接头蛋白(p62)、溶酶体相关膜蛋白1(LAMP1)的表达水平;利用透射电镜(TEM)检测自噬溶酶体的数量;并用酶联免疫吸附测定(ELISA)法检测细胞培养上清中白细胞介素(IL)-1β、IL-6、IL-18、肿瘤坏死因子(TNF)-α的含量。结果与正常对照组相比,OGD/R组lncRNA TUG1、NLRP3、Atg5、beclin1、p62、LAMP1的表达水平以及LC3-II/I比值均显著上调(^(均)P<0.05),自噬溶酶体数量增加(P<0.05),IL-1β、IL-6、IL-18、TNF-α的含量显著升高(^(均)P<0.05)。与OGD/R组相比,OGD/R+SGB组的上述指标均显著下调(P<0.05)。与OGD/R+SGB+TUG1过表达阴性对照组相比,OGD/R+SGB+TUG1过表达组的lncRNA TUG1、NLRP3、Atg5、beclin1、p62、LAMP1的表达水平以及LC3-II/I比值均显著上调(^(均)P<0.05),自噬溶酶体数量增加(P<0.05),IL-1β、IL-6、IL-18、TNF-α的含量显著升高(^(均)P<0.05),然而,加入NLRP3的抑制剂MCC950后,除lncRNA TUG1外其余指标均显著下调(^(均)P<0.05)。另外,与OGD/R组比,OGD/R+TUG1过表达组的上述指标进一步上调(^(均)P<0.05)。与OGD/R组比,OGD/R+MCC950组则抑制了除lncRNA TUG1外的其余指标(^(均)P<0.05)。结论星状神经节阻滞通过调节lncRNA TUG1-NLRP3轴有效减轻体外脑缺血再灌注损伤引起的炎症反应和自噬溶酶体形成,提示其可能作为治疗缺血性脑损伤的潜在策略。
文摘目的研究分析维持性血液透析患者血清巨噬细胞炎性蛋白-1α(macrophage inflammatory protein-1α,MIP-1α)、几丁质酶-3样蛋白1(chitinase-3-like protein 1,CHI3L1/YKL-40)、含pyrin结构域的NOD样受体3(NOD like receptor family pyrin domain-containing protein 3,NLRP3)水平与导管相关性感染及预后的相关性。方法选取泸州市中医医院在2020年1月─2024年8月收治的维持性血液透析患者296例,根据是否发生导管感染分为感染组(n=68)和未感染组(n=228),根据预后情况分为预后良好组(n=42)和预后不良组(n=26),经ELISA检测血清MIP-1α、YKL-40、NLRP3水平,收集分析一般临床资料,多因素Logistic回归分析患者预后影响因素,受试者工作特征(receiver operating characteristic,ROC)曲线分析MIP-1α、YKL-40、NLRP3对患者预后的预测价值,Pearson及Spearman法分析MIP-1α、YKL-40、NLRP3与炎症因子及预后的相关性。结果与非感染组相比,感染组的血清MIP-1α、YKL-40、NLRP3、C反应蛋白(C-reactive protein,CRP)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、降钙素原(procalcitonin,PCT)水平显著升高(t=14.532、14.561、22.823、28.495、11.487、22.234,均P<0.001);预后不良组的透析龄、透析频率、MIP-1α、YKL-40、NLRP3、CRP、TNF-α、PCT水平显著高于预后良好组(t=3.293、3.293、8.410、7.432、7.511、15.045、3.694、4.007,P=0.002、0.001、<0.001、<0.001、<0.001、<0.001、<0.001、<0.001);ROC曲线显示MIP-1α、YKL-40、NLRP3三者联合对患者预后预测的AUC显著高于MIP-1α(Z=2.101,P=0.036)、YKL-40(Z=2.610,P=0.009)、NLRP3(Z=2.965,P=0.003)单独预测;MIP-1α、YKL-40、NLRP3与炎症因子CRP(r=0.462、0.466、0.472,均P<0.001)、TNF-α(r=0.436、0.426、0.451,均P<0.001)、PCT(r=0.450、0.444、0.455,均P<0.001)以及预后(r=0.521、0.502、0.504,P<0.001)均呈正相关。结论血清MIP-1α、YKL-40、NLRP3与维持性血液透析患者的导管相关性感染及预后具有一定相关性。
