Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted ...Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted therapy.In our study,the role of long non-coding RNA(lncRNA)AFAP1-AS1 in gemcitabine resistance and related mechanisms were explored in cervical cancer cells.Methods:Gemcitabine-resistant cervical cancer cell lines HT-3-Gem and SW756-Gem were constructed using the gemcitabine concentration gradient method.The overall survival rates and recurrence-free survival rates were evaluated by Kaplan-Meier analysis.The interaction was verified through a Dual-luciferase reporter gene assay and a Biotinylated RNA pull-down assay.Cell proliferation ability was assessed through methyl-thiazolyl-tetrazolium(MTT),soft agar,and colony formation experiments.Cell cycle and apoptosis were detected byflow cytometry.Results:Up-regulation of AFAP1-AS1 in cervical cancer predicted a poor prognosis.Besides,patients in the gemcitabine-resistance group had higher levels of AFAP1-AS1 than the gemcitabine-sensitive group.AFAP1-AS1 promoted tumor growth and induced gemcitabine tolerance of cervical cancer cells.In addition,AFAP1-AS1 mediated epidermal growth factor receptor(EGFR)expression by serving as a molecular sponge for microRNA-7a-5p(miR-7-5p).This present study also proved that the knockdown of EGFR or overexpression of miR-7a-5p abolished the accelerative role of AFAP1-AS1 overexpression in cancer progression and gemcitabine tolerance.Conclusions:In general,the AFAP1-AS1/miR-7-5p/EGFR axis was tightly related to the progression and gemcitabine tolerance of cervical cancer,providing potential targets for the management of cervical cancer.展开更多
目的探究乳腺癌源性LncRNA OIP5-AS1经肿瘤相关巨噬细胞(TAM)M2型极化调控乳腺癌细胞迁移、侵袭及上皮—间质转化的机制。方法MDA-MB-231细胞分设Control组(空白对照)、NC组(转染NC si RNA)和si-OIP5组(转染Lnc RNAOIP5-AS1 si RNA)。RT...目的探究乳腺癌源性LncRNA OIP5-AS1经肿瘤相关巨噬细胞(TAM)M2型极化调控乳腺癌细胞迁移、侵袭及上皮—间质转化的机制。方法MDA-MB-231细胞分设Control组(空白对照)、NC组(转染NC si RNA)和si-OIP5组(转染Lnc RNAOIP5-AS1 si RNA)。RT-qPCR检测Lnc RNA OIP5-AS1、IL-4和IL-13 m RNA表达;ELISA检测培养基上清IL-4和IL-13蛋白表达。Control组培养上清按不同体积比加入RPMI1640培养基中诱导M0巨噬细胞向TAM极化,RT-qPCR检测TAM中CD206 m RNA的表达;以NC组/si-OIP5组培养基上清诱导M0型巨噬细胞,Western blot检测CD206表达;继而构建巨噬细胞MDA-MB-231共培养模型,评估MDAMB-231细胞迁移侵袭能力并检测Vimentin、N-cadherin、Ecadherin表达变化。结果相较于Control及NC组,si-OIP5组Lnc RNAOIP5-AS1表达显著下调(P<0.001),IL-13m RNA及蛋白水平均显著降低(P<0.05),而IL-4表达在各组间无明显差异。条件培养基呈体积依赖性诱导M0巨噬细胞CD206表达,40%体积比时效应最强(P<0.001)。相较NC组,si-OIP5组CD206蛋白水平显著降低(P<0.01)。共培养体系中,si-OIP5组MDA-MB-231细胞迁移侵袭能力显著减弱(P<0.001),E-cadherin表达上调,N-cadherin及Vimentin表达下调(均P<0.01)。结论乳腺癌源性Lnc RNA OIP5-AS1可经IL-13介导诱导TAM M2型极化,进而促进乳腺癌细胞的迁移、侵袭及EMT。展开更多
Objectives The discovery of novel molecular targets to enhance the osteogenesis of human bone marrow-derived mesenchymal stem cells(H-BMSCs)represents a promising strategy for preventing and treating osteoporosis.Thus...Objectives The discovery of novel molecular targets to enhance the osteogenesis of human bone marrow-derived mesenchymal stem cells(H-BMSCs)represents a promising strategy for preventing and treating osteoporosis.Thus,the primary objective of this study is to elucidate the mechanisms by which long non-coding RNA FOXD2-AS1(lncRNA FOXD2-AS1)regulates early osteogenic differentiation in H-BMSCs,thereby identifying potential therapeutic targets.Methods Lentivirus-mediated vectors were constructed to either overexpress or silence FOXD2-AS1 in H-BMSCs.The effects of FOXD2-AS1 on osteogenesis were subsequently assessed by analyzing osteogenic marker expression and alkaline phosphatase(ALP)staining.To clarify the role of the Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)pathway in this process,AG490 inhibitor(a JAK2/STAT3 pathway inhibitor)and knockdown of STAT3 were used to investigate the mechanisms of FOXD2-AS1.Results FOXD2-AS1 overexpression increased ALP activity and osteogenic marker expression,while its knockdown had the opposite effects.From a mechanistic perspective,FOXD2-AS1 overexpression promoted JAK2 and STAT3 phosphorylation,whereas its suppression attenuated their activation.Also,the osteogenic increase induced by FOXD2-AS1 overexpression was reversed by AG490 treatment or STAT3 silencing,indicating that the pathway plays a role in this process.Conclusion FOXD2-AS1 was identified as a novel genetic switch driving osteogenic commitment via JAK2/STAT3 activation,revealing a new regulatory mechanism and a potential therapeutic target for osteoporosis.展开更多
文摘Background:Drug resistance is the main factor contributing to cancer recurrence and poor prognosis.Exploration of drug resistance-related mechanisms and effective therapeutic targets are the aim of molecular targeted therapy.In our study,the role of long non-coding RNA(lncRNA)AFAP1-AS1 in gemcitabine resistance and related mechanisms were explored in cervical cancer cells.Methods:Gemcitabine-resistant cervical cancer cell lines HT-3-Gem and SW756-Gem were constructed using the gemcitabine concentration gradient method.The overall survival rates and recurrence-free survival rates were evaluated by Kaplan-Meier analysis.The interaction was verified through a Dual-luciferase reporter gene assay and a Biotinylated RNA pull-down assay.Cell proliferation ability was assessed through methyl-thiazolyl-tetrazolium(MTT),soft agar,and colony formation experiments.Cell cycle and apoptosis were detected byflow cytometry.Results:Up-regulation of AFAP1-AS1 in cervical cancer predicted a poor prognosis.Besides,patients in the gemcitabine-resistance group had higher levels of AFAP1-AS1 than the gemcitabine-sensitive group.AFAP1-AS1 promoted tumor growth and induced gemcitabine tolerance of cervical cancer cells.In addition,AFAP1-AS1 mediated epidermal growth factor receptor(EGFR)expression by serving as a molecular sponge for microRNA-7a-5p(miR-7-5p).This present study also proved that the knockdown of EGFR or overexpression of miR-7a-5p abolished the accelerative role of AFAP1-AS1 overexpression in cancer progression and gemcitabine tolerance.Conclusions:In general,the AFAP1-AS1/miR-7-5p/EGFR axis was tightly related to the progression and gemcitabine tolerance of cervical cancer,providing potential targets for the management of cervical cancer.
文摘目的探究乳腺癌源性LncRNA OIP5-AS1经肿瘤相关巨噬细胞(TAM)M2型极化调控乳腺癌细胞迁移、侵袭及上皮—间质转化的机制。方法MDA-MB-231细胞分设Control组(空白对照)、NC组(转染NC si RNA)和si-OIP5组(转染Lnc RNAOIP5-AS1 si RNA)。RT-qPCR检测Lnc RNA OIP5-AS1、IL-4和IL-13 m RNA表达;ELISA检测培养基上清IL-4和IL-13蛋白表达。Control组培养上清按不同体积比加入RPMI1640培养基中诱导M0巨噬细胞向TAM极化,RT-qPCR检测TAM中CD206 m RNA的表达;以NC组/si-OIP5组培养基上清诱导M0型巨噬细胞,Western blot检测CD206表达;继而构建巨噬细胞MDA-MB-231共培养模型,评估MDAMB-231细胞迁移侵袭能力并检测Vimentin、N-cadherin、Ecadherin表达变化。结果相较于Control及NC组,si-OIP5组Lnc RNAOIP5-AS1表达显著下调(P<0.001),IL-13m RNA及蛋白水平均显著降低(P<0.05),而IL-4表达在各组间无明显差异。条件培养基呈体积依赖性诱导M0巨噬细胞CD206表达,40%体积比时效应最强(P<0.001)。相较NC组,si-OIP5组CD206蛋白水平显著降低(P<0.01)。共培养体系中,si-OIP5组MDA-MB-231细胞迁移侵袭能力显著减弱(P<0.001),E-cadherin表达上调,N-cadherin及Vimentin表达下调(均P<0.01)。结论乳腺癌源性Lnc RNA OIP5-AS1可经IL-13介导诱导TAM M2型极化,进而促进乳腺癌细胞的迁移、侵袭及EMT。
基金supported by the Natural Science Foundation of Hubei Province of China(Grant No.2023AFB671)the National Natural Science Foundation of China(Grant Nos.82360177 and 82560182)+1 种基金the Key Project of Jiangxi Provincial Natural Science Foundation(Grant No.20224ACB206011)“Xuncheng Talents”Project in Jiujiang City,Jiangxi Province(Grant No.JJXC2023071).
文摘Objectives The discovery of novel molecular targets to enhance the osteogenesis of human bone marrow-derived mesenchymal stem cells(H-BMSCs)represents a promising strategy for preventing and treating osteoporosis.Thus,the primary objective of this study is to elucidate the mechanisms by which long non-coding RNA FOXD2-AS1(lncRNA FOXD2-AS1)regulates early osteogenic differentiation in H-BMSCs,thereby identifying potential therapeutic targets.Methods Lentivirus-mediated vectors were constructed to either overexpress or silence FOXD2-AS1 in H-BMSCs.The effects of FOXD2-AS1 on osteogenesis were subsequently assessed by analyzing osteogenic marker expression and alkaline phosphatase(ALP)staining.To clarify the role of the Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)pathway in this process,AG490 inhibitor(a JAK2/STAT3 pathway inhibitor)and knockdown of STAT3 were used to investigate the mechanisms of FOXD2-AS1.Results FOXD2-AS1 overexpression increased ALP activity and osteogenic marker expression,while its knockdown had the opposite effects.From a mechanistic perspective,FOXD2-AS1 overexpression promoted JAK2 and STAT3 phosphorylation,whereas its suppression attenuated their activation.Also,the osteogenic increase induced by FOXD2-AS1 overexpression was reversed by AG490 treatment or STAT3 silencing,indicating that the pathway plays a role in this process.Conclusion FOXD2-AS1 was identified as a novel genetic switch driving osteogenic commitment via JAK2/STAT3 activation,revealing a new regulatory mechanism and a potential therapeutic target for osteoporosis.
