Article(id=1324078474599740321, tenantId=1146029695717560320, journalId=1227665162245664772, issueId=1324078333767594037, articleNumber=null, orderNo=null, doi=10.20043/j.cnki.MPM.202505155, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1747065600000, receivedDateStr=2025-05-13, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1793614508533, onlineDateStr=2026-11-02, pubDate=1767974400000, pubDateStr=2026-01-10, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1793614508533, onlineIssueDateStr=2026-11-02, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1793614508533, creator=13701087609, updateTime=1793614508533, updator=13701087609, issue=Issue{id=1324078333767594037, tenantId=1146029695717560320, journalId=1227665162245664772, year='2026', volume='53', issue='1', pageStart='1', pageEnd='192', issueExtLink='null', onlineDate='null', pubDate='1767974400000', pubDateStr='2026-01-10', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1793614474956, creator='13701087609', updateTime=1793614571752, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1324078739914621270, tenantId=1146029695717560320, journalId=1227665162245664772, issueId=1324078333767594037, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1324078739914621271, tenantId=1146029695717560320, journalId=1227665162245664772, issueId=1324078333767594037, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=53, endPage=61, ext={EN=ArticleExt(id=1324078474796872610, articleId=1324078474599740321, tenantId=1146029695717560320, journalId=1227665162245664772, language=EN, title=Mechanistic study of circular RNA hsa_circ_0058493 in arsenic-induced apoptosis of lung cells, columnId=1228016570660745413, journalTitle=Modern Preventive Medicine, columnName=Environmental and Occupational Health, runingTitle=null, highlight=null, articleAbstract=
Objective

This study aims to investigate the mechanism by which circular RNA hsa_circ_0058493 mediates arsenic exposure-induced apoptosis in lung cells.

Methods

A549 and BEAS-2B cells were exposed to different concentrations of sodium arsenite (NaAsO2), and the half-maximal inhibitory concentration (IC50) of NaAsO2 was determined by the CCK-8 assay. The effects of three arsenic species (NaAsO2, MMA, DMA) on hsa_circ_0058493 expression were analyzed by real-time quantitative PCR (RT-qPCR). After silencing hsa_circ_0058493 with siRNA, cell viability, mitochondrial membrane potential, and apoptosis were evaluated using CCK-8, JC-1 staining, and Hoechst 33342/PI double staining, respectively. The JAK2/STAT3 signaling pathway and related protein expression were assessed by Western blotting.

Results

NaAsO2 significantly reduced cell viability(IC50: 35.03 μmol/L for A549, 7.809 μmol/L for BEAS-2B) and specifically induced the upregulation of hsa_circ_0058493 (P<0.05), while MMA and DMA showed no significant effects. Downregulating hsa_circ_0058493 reduced cell viability, depolarized mitochondrial membrane potential, and increased apoptosis (P<0.05). Western blotting showed that depleting hsa_circ_0058493 suppressed JAK2/STAT3 phosphorylation and downregulated c-Myc, CyclinD1, XIAP, and BCL-2 expression levels (P<0.05).After downregulating hsa_circ_0058493, cells were treated with NaAsO2, resulting in a further decrease in cell viability, and STAT3 protein expression differed between the two cell lines.

Conclusion

Downregulation of hsa_circ_0058493 induced decreased cell viability and apoptosis, which were associated with inhibition of JAK2/STAT3 pathway activity, suggesting that hsa_circ_0058493 may be involved in arsenic-induced lung cell apoptosis by regulating the JAK2/STAT3 signaling pathway.

, authors=Ting-zhi DUAN, Xue-fei YANG, Bin CHEN, Jing-chao YANG, Xin-da YANG, Qing-yun ZHAO, Yue-feng HE, authorsList=Ting-zhi DUAN, Xue-fei YANG, Bin CHEN, Jing-chao YANG, Xin-da YANG, Qing-yun ZHAO, Yue-feng HE, authorCompany=null, correspAuthors=Yue-feng HE, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1324078477896463282, articleId=1324078474599740321, tenantId=1146029695717560320, journalId=1227665162245664772, language=CN, title=环状RNA hsa_circ_0058493在砷诱导肺细胞凋亡中的机制研究, columnId=1228016571273113811, journalTitle=现代预防医学, columnName=环境与职业卫生, runingTitle=null, highlight=null, articleAbstract=
目的

本研究旨在探讨环状RNA hsa_circ_0058493在砷暴露诱导肺细胞凋亡中的作用机制。

方法

以A549和BEAS-2B细胞为研究对象,用不同浓度亚砷酸钠(NaAsO2)染毒细胞,通过CCK-8法测定NaAsO2的半抑制浓度(half-maimal inhibitory concentration,IC50)。采用RT-qPCR检测三种砷形态(NaAsO2、MMA、DMA)对hsa_circ_0058493表达的影响。利用siRNA下调hsa_circ_0058493后,采用CCK-8、JC-1染色、Hoechst 33342/PI双染法检测细胞活力、线粒体膜电位及细胞凋亡情况,Western blotting检测JAK2/STAT3通路及相关蛋白表达。

结果

NaAsO2显著降低细胞存活率(A549 IC50=35.03 μmol/L,BEAS-2B IC50=7.809 μmol/L),并且NaAsO2可诱导hsa_circ_0058493表达升高(P<0.05),而MMA和DMA处理未见显著影响。下调hsa_circ_0058493可导致细胞活力下降、线粒体膜电位去极化、细胞凋亡率升高(P<0.05)。蛋白检测结果显示,下调hsa_circ_0058493抑制JAK2/STAT3通路磷酸化水平,并下调c-Myc、CyclinD1、XIAP、BCL-2蛋白表达(P<0.05)。下调hsa_circ_0058493后用NaAsO2处理细胞,细胞存活率进一步降低,且STAT3蛋白表达在两个细胞中不同。

结论

下调hsa_circ_0058493可引起细胞活力下降和凋亡,这些效应与JAK2/STAT3通路活性抑制相关。hsa_circ_0058493可能通过调控JAK2/STAT3信号通路参与砷诱导肺细胞凋亡。

, authors=段婷芝, 杨雪飞, 陈斌, 杨净超, 杨欣达, 赵青云, 何越峰, authorsList=段婷芝, 杨雪飞, 陈斌, 杨净超, 杨欣达, 赵青云, 何越峰, authorCompany=null, correspAuthors=何越峰, authorNote=

段婷芝(2000—),女,硕士在读,研究方向:劳动与环境卫生

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何越峰,E-mail:
, copyrightStatement=本刊刊出的所有文章不代表中华预防医学会和本刊编委会的观点,除非特别声明。, copyrightOwner=中华预防医学会和四川大学华西公共卫生学院, extLink=null, articleAbsUrl=null, sourceXml=i9KOPNviiurWV+iv7PwVLQ==, magXml=VyhSZ0C7qo+iupAxBiL0OA==, pdfUrl=null, pdf=Jz//Rq3YR5q2CwA/koA/Dg==, pdfFileSize=1224553, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=A/7oKBSjU9PQ31Eil0RJQg==, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=kkH3eYHMfXBc2XEcvaWEBw==, mapNumber=null, fund=null)}, authors=[Author(id=1324078478911484856, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, orderNo=0, firstName=null, middleName=null, lastName=null, nameCn=null, orcid=null, stid=null, country=null, authorPic=null, dead=0, email=null, emailSecond=null, emailThird=null, correspondingAuthor=0, authorType=1, ext={EN=AuthorExt(id=1324078479368663994, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, authorId=1324078478911484856, language=EN, stringName=Ting-zhi DUAN, firstName=Ting-zhi, middleName=null, lastName=DUAN, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=null, address=Yunnan Provincial Key Laboratory of Public Health and Biosecurity & School of Public Health, Kunming Medical University,Kunming, Yunnan 650500, China, bio=null, bioImg=null, bioContent=null, aboutCorrespAuthor=null), CN=AuthorExt(id=1324078479708402619, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, authorId=1324078478911484856, language=CN, stringName=段婷芝, firstName=null, middleName=null, lastName=null, prefix=null, suffix=null, authorComment=null, nameInitials=null, affiliation=null, department=null, xref=null, address=云南省公共卫生与生物安全重点实验室,昆明医科大学公共卫生学院,云南 昆明 650500, bio={"content":"

段婷芝(2000—),女,硕士在读,研究方向:劳动与环境卫生

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Cancer Cell International, 2024, 24(1): 16., articleTitle=Interplay of miRNAs and lncRNAs in STAT3 signaling pathway in colorectal cancer progression, refAbstract=null), Reference(id=1324078501325844494, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, doi=null, pmid=null, pmcid=null, year=2024, volume=12, issue=1, pageStart=40, pageEnd=null, url=null, language=null, rfNumber=[20], rfOrder=20, authorNames=Zhao ZY, Yu PP, Wang Y, journalName=Cancer & Metabolism, refType=null, unstructuredReference=Zhao ZY, Yu PP, Wang Y, et al. Silencing of STEAP3 suppresses cervical cancer cell proliferation and migration via JAK/STAT3 signaling pathway[J]. Cancer & Metabolism, 2024, 12(1): 40., articleTitle=Silencing of STEAP3 suppresses cervical cancer cell proliferation and migration via JAK/STAT3 signaling pathway, refAbstract=null), Reference(id=1324078501384564751, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, doi=null, pmid=null, pmcid=null, year=2023, volume=18, issue=1, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=[21], rfOrder=21, authorNames=Tan JY, Feng RH, journalName=Open Med (Wars), refType=null, unstructuredReference=Tan JY, Feng RH. A pan-cancer analysis of STAT3 expression and genetic alterations in human tumors[J]. Open Med (Wars), 2023, 18(1): 20230792., articleTitle=A pan-cancer analysis of STAT3 expression and genetic alterations in human tumors, refAbstract=null)], funds=[Fund(id=1324078496800191478, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, awardId=82160607, language=CN, fundingSource=国家自然科学基金(82160607), fundOrder=null, country=null), Fund(id=1324078496871494647, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, awardId=202401AT070177, language=CN, fundingSource=云南省科技研究计划(202401AT070177), fundOrder=null, country=null), Fund(id=1324078496942797816, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, awardId=202405AS350016, language=CN, fundingSource=云南省创新团队(202405AS350016), fundOrder=null, country=null), Fund(id=1324078497005712377, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, awardId=CXTD202201, language=CN, fundingSource=昆明医科大学创新团队(CXTD202201), fundOrder=null, country=null), Fund(id=1324078497081209850, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, awardId=J114150330366, language=CN, fundingSource=昆明医科大学硕士研究生教育创新基金(J114150330366), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1324078478533997491, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, xref=null, ext=[AuthorCompanyExt(id=1324078478563357620, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, companyId=1324078478533997491, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=Yunnan Provincial Key Laboratory of Public Health and Biosecurity & School of Public Health, Kunming Medical University,Kunming, Yunnan 650500, China), AuthorCompanyExt(id=1324078478575940534, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, companyId=1324078478533997491, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=云南省公共卫生与生物安全重点实验室,昆明医科大学公共卫生学院,云南 昆明 650500)])], figs=[ArticleFig(id=1324078493599937508, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 1, caption=Effects of different concentrations of NaAsO2 on the viability of A549 and BEAS-2B cells, figureFileSmall=QHQ0ygNYm+vH7pubywKUAQ==, figureFileBig=A/7oKBSjU9PQ31Eil0RJQg==, tableContent=null), ArticleFig(id=1324078493688017893, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图1, caption=不同浓度NaAsO2染毒对A549和BEAS-2B细胞存活率的影响

注:A为不同浓度NaAsO2对A549细胞存活率的影响;B为不同浓度NaAsO2对BEAS-2B细胞存活率的影响;与对照组比,*表示P<0.05;**表示P<0.01。

, figureFileSmall=QHQ0ygNYm+vH7pubywKUAQ==, figureFileBig=A/7oKBSjU9PQ31Eil0RJQg==, tableContent=null), ArticleFig(id=1324078495487374310, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 2, caption=Effects of NaAsO2, MMA, and DMA on the expression of hsa_circ_0058493 in A549 and BEAS-2B cells, figureFileSmall=kmUfeW0AgwJEZyQthx5Fqw==, figureFileBig=5B7Q5q1F65weOypH44eJLw==, tableContent=null), ArticleFig(id=1324078495571260391, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图2, caption=NaAsO2、MMA、DMA对A549和BEAS-2B细胞中hsa_circ_0058493表达的影响

注:A为不同浓度NaAsO2对A549细胞中hsa_circ_0058493表达的影响;B为不同浓度NaAsO2对BEAS-2B细胞中hsa_circ_0058493表达的影响;C为MMA、DMA和NaAsO2(30 μmol/L)对A549细胞中hsa_circ_0058493表达的影响;D为MMA、DMA和NaAsO2(6 μmol/L)对BEAS-2B细胞中hsa_circ_0058493表达的影响;*表示P<0.05;**表示P<0.01。

, figureFileSmall=kmUfeW0AgwJEZyQthx5Fqw==, figureFileBig=5B7Q5q1F65weOypH44eJLw==, tableContent=null), ArticleFig(id=1324078495634174952, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 3, caption=Validation of the silencing efficiency of siRNA in A549 and BEAS-2B cells, figureFileSmall=AFoHUpZLH2boHYgADiHYgA==, figureFileBig=MDvZdqwNmnh38vUp8BENqw==, tableContent=null), ArticleFig(id=1324078495713866729, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图3, caption=A549和BEAS-2B细胞中siRNA沉默效率验证

注:A为A549细胞中siRNA沉默效率验证;B为BEAS-2B细胞中siRNA沉默效率验证;*表示P<0.05;**表示P<0.01。

, figureFileSmall=AFoHUpZLH2boHYgADiHYgA==, figureFileBig=MDvZdqwNmnh38vUp8BENqw==, tableContent=null), ArticleFig(id=1324078495890027498, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 4, caption=Effects of hsa_circ_0058493 downregulation on cell viability and mitochondrial membrane potential in A549 and BEAS-2B cells, figureFileSmall=s+/TX4RdisLp2aCqN43qQw==, figureFileBig=mXsMK03wYByf+P9f5BXNIw==, tableContent=null), ArticleFig(id=1324078495961330667, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图4, caption=下调hsa_circ_0058493对A549和BEAS-2B细胞活力及线粒体膜电位的影响

注:A为下调hsa_circ_0058493对A549细胞存活率的影响;B为下调hsa_circ_0058493对BEAS-2B细胞存活率的影响;C为下调hsa_circ_0058493对A549细胞线粒体膜电位的影响;D为下调hsa_circ_0058493对BEAS-2B细胞线粒体膜电位的影响;*表示P<0.05;**表示P<0.01。

, figureFileSmall=s+/TX4RdisLp2aCqN43qQw==, figureFileBig=mXsMK03wYByf+P9f5BXNIw==, tableContent=null), ArticleFig(id=1324078496028439532, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 5, caption=Effects of hsa_circ_0058493 downregulation on apoptosis in A549 and BEAS-2B cells, figureFileSmall=qu0LjTGBnWF4BNqZ2Kj1Hg==, figureFileBig=BA+76lnATxWNvlrbIUknLw==, tableContent=null), ArticleFig(id=1324078496095548397, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图5, caption=下调hsa_circ_0058493对A549和BEAS-2B细胞凋亡的影响

注:A为A549细胞Hoechst 33342/PI双染结果(黄色箭头示早期凋亡细胞),放大倍数为200倍;B为BEAS-2B细胞Hoechst 33342/PI双染结果;C为A549细胞凋亡率统计;D为BEAS-2B细胞凋亡率统计;*表示P<0.05;**表示P<0.01。

, figureFileSmall=qu0LjTGBnWF4BNqZ2Kj1Hg==, figureFileBig=BA+76lnATxWNvlrbIUknLw==, tableContent=null), ArticleFig(id=1324078496175240174, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 6, caption=Effects of hsa_circ_0058493 downregulation on related protein expression in A549 and BEAS-2B cells, figureFileSmall=LDnfr2+gmrHym1/tLB5hJQ==, figureFileBig=q00SEAp9C/2OsfrCNug15A==, tableContent=null), ArticleFig(id=1324078496238154735, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图6, caption=下调hsa_circ_0058493对A549和BEAS-2B细胞相关蛋白表达的影响

注:A、D为Western blot检测A549和BEAS-2B细胞中蛋白表达变化;B、E为A549细胞中相关蛋白相对表达量;C、F为BEAS-2B细胞中相关蛋白相对表达量;*表示P<0.05;**表示P<0.01。

, figureFileSmall=LDnfr2+gmrHym1/tLB5hJQ==, figureFileBig=q00SEAp9C/2OsfrCNug15A==, tableContent=null), ArticleFig(id=1324078496309457904, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Figure 7, caption=Effects of NaAsO2 treatment following downregulation of hsa_circ_0058493 on cell viability and related proteins in A549 and BEAS-2B cells, figureFileSmall=eUdJrQGi3+4+Bs7MOZ+TNg==, figureFileBig=CblPRUWqZphZ8ksQrgCoCQ==, tableContent=null), ArticleFig(id=1324078496380761073, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=图7, caption=下调hsa_circ_0058493后给予NaAsO2处理对A549和BEAS-2B细胞存活率及相关蛋白的影响

注:A为下调hsa_circ_0058493后给予NaAsO2处理对A549细胞存活率的影响;B为下调hsa_circ_0058493后给予NaAsO2处理对BEAS-2B细胞存活率的影响;C为Western blot检测A549细胞中STAT3蛋白表达变化;D为Western blot检测BEAS-2B细胞中STAT3蛋白表达变化;E为A549细胞中蛋白相对表达量;F为BEAS-2B细胞中蛋白相对表达量;*表示在同一处理组内与对照组比较存在统计学差异(P<0.05);^表示si-circ+NaAsO2组的NC、siRNA#1、siRNA#2与对应si-circ组的NC、siRNA#1、siRNA#2比较存在统计学差异(P<0.05)。

, figureFileSmall=eUdJrQGi3+4+Bs7MOZ+TNg==, figureFileBig=CblPRUWqZphZ8ksQrgCoCQ==, tableContent=null), ArticleFig(id=1324078496473035762, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Table 1, caption=

Primer sequences

, figureFileSmall=null, figureFileBig=null, tableContent=
基因正向引物序列(5'~3')反向引物序列(5'~3')
β-actinGCCGAGGACTTTGATTGCACTGGACTTGGGAGAGGACTGG
hsa_circ_0058493GCAACGGAGATCAAGAGGGATATTCTCACCAGGGTTCAGC
), ArticleFig(id=1324078496531756019, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=表1, caption=

引物序列

, figureFileSmall=null, figureFileBig=null, tableContent=
基因正向引物序列(5'~3')反向引物序列(5'~3')
β-actinGCCGAGGACTTTGATTGCACTGGACTTGGGAGAGGACTGG
hsa_circ_0058493GCAACGGAGATCAAGAGGGATATTCTCACCAGGGTTCAGC
), ArticleFig(id=1324078496611447796, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=EN, label=Table 2, caption=

siRNA interference fragment sequence

, figureFileSmall=null, figureFileBig=null, tableContent=
基因正义链(5'~3')反义链(5'~3')
Negative controlUUCUCCGAACGUGUCACGUTTACGUGACACGUUCGGAGAATT
siRNA#1UUAUUCUCACCAGGGUUCATTUGAACCCUGGUGAGAAUAATT
siRNA#2AUUCUCACCAGGGUUCAGCTTGCUGAACCCUGGUGAGAAUTT
), ArticleFig(id=1324078496682750965, tenantId=1146029695717560320, journalId=1227665162245664772, articleId=1324078474599740321, language=CN, label=表2, caption=

siRNA干扰片段序列

, figureFileSmall=null, figureFileBig=null, tableContent=
基因正义链(5'~3')反义链(5'~3')
Negative controlUUCUCCGAACGUGUCACGUTTACGUGACACGUUCGGAGAATT
siRNA#1UUAUUCUCACCAGGGUUCATTUGAACCCUGGUGAGAAUAATT
siRNA#2AUUCUCACCAGGGUUCAGCTTGCUGAACCCUGGUGAGAAUTT
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环状RNA hsa_circ_0058493在砷诱导肺细胞凋亡中的机制研究
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段婷芝 , 杨雪飞 , 陈斌 , 杨净超 , 杨欣达 , 赵青云 , 何越峰
现代预防医学 | 环境与职业卫生 2026,53(1): 53-61
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现代预防医学 |环境与职业卫生 2026 , 53 (1) : 53 -61
环状RNA hsa_circ_0058493在砷诱导肺细胞凋亡中的机制研究
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段婷芝, 杨雪飞, 陈斌, 杨净超, 杨欣达, 赵青云, 何越峰
作者信息
  • 云南省公共卫生与生物安全重点实验室,昆明医科大学公共卫生学院,云南 昆明 650500
通讯作者:
何越峰,E-mail:
作者简介:

段婷芝(2000—),女,硕士在读,研究方向:劳动与环境卫生

Mechanistic study of circular RNA hsa_circ_0058493 in arsenic-induced apoptosis of lung cells
Ting-zhi DUAN, Xue-fei YANG, Bin CHEN, Jing-chao YANG, Xin-da YANG, Qing-yun ZHAO, Yue-feng HE
Affiliations
  • Yunnan Provincial Key Laboratory of Public Health and Biosecurity & School of Public Health, Kunming Medical University,Kunming, Yunnan 650500, China
出版时间: 2026-01-10 doi: 10.20043/j.cnki.MPM.202505155
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目的

本研究旨在探讨环状RNA hsa_circ_0058493在砷暴露诱导肺细胞凋亡中的作用机制。

方法

以A549和BEAS-2B细胞为研究对象,用不同浓度亚砷酸钠(NaAsO2)染毒细胞,通过CCK-8法测定NaAsO2的半抑制浓度(half-maimal inhibitory concentration,IC50)。采用RT-qPCR检测三种砷形态(NaAsO2、MMA、DMA)对hsa_circ_0058493表达的影响。利用siRNA下调hsa_circ_0058493后,采用CCK-8、JC-1染色、Hoechst 33342/PI双染法检测细胞活力、线粒体膜电位及细胞凋亡情况,Western blotting检测JAK2/STAT3通路及相关蛋白表达。

结果

NaAsO2显著降低细胞存活率(A549 IC50=35.03 μmol/L,BEAS-2B IC50=7.809 μmol/L),并且NaAsO2可诱导hsa_circ_0058493表达升高(P<0.05),而MMA和DMA处理未见显著影响。下调hsa_circ_0058493可导致细胞活力下降、线粒体膜电位去极化、细胞凋亡率升高(P<0.05)。蛋白检测结果显示,下调hsa_circ_0058493抑制JAK2/STAT3通路磷酸化水平,并下调c-Myc、CyclinD1、XIAP、BCL-2蛋白表达(P<0.05)。下调hsa_circ_0058493后用NaAsO2处理细胞,细胞存活率进一步降低,且STAT3蛋白表达在两个细胞中不同。

结论

下调hsa_circ_0058493可引起细胞活力下降和凋亡,这些效应与JAK2/STAT3通路活性抑制相关。hsa_circ_0058493可能通过调控JAK2/STAT3信号通路参与砷诱导肺细胞凋亡。

砷  /  hsa_circ_0058493  /  细胞凋亡  /  STAT3信号通路
Objective

This study aims to investigate the mechanism by which circular RNA hsa_circ_0058493 mediates arsenic exposure-induced apoptosis in lung cells.

Methods

A549 and BEAS-2B cells were exposed to different concentrations of sodium arsenite (NaAsO2), and the half-maximal inhibitory concentration (IC50) of NaAsO2 was determined by the CCK-8 assay. The effects of three arsenic species (NaAsO2, MMA, DMA) on hsa_circ_0058493 expression were analyzed by real-time quantitative PCR (RT-qPCR). After silencing hsa_circ_0058493 with siRNA, cell viability, mitochondrial membrane potential, and apoptosis were evaluated using CCK-8, JC-1 staining, and Hoechst 33342/PI double staining, respectively. The JAK2/STAT3 signaling pathway and related protein expression were assessed by Western blotting.

Results

NaAsO2 significantly reduced cell viability(IC50: 35.03 μmol/L for A549, 7.809 μmol/L for BEAS-2B) and specifically induced the upregulation of hsa_circ_0058493 (P<0.05), while MMA and DMA showed no significant effects. Downregulating hsa_circ_0058493 reduced cell viability, depolarized mitochondrial membrane potential, and increased apoptosis (P<0.05). Western blotting showed that depleting hsa_circ_0058493 suppressed JAK2/STAT3 phosphorylation and downregulated c-Myc, CyclinD1, XIAP, and BCL-2 expression levels (P<0.05).After downregulating hsa_circ_0058493, cells were treated with NaAsO2, resulting in a further decrease in cell viability, and STAT3 protein expression differed between the two cell lines.

Conclusion

Downregulation of hsa_circ_0058493 induced decreased cell viability and apoptosis, which were associated with inhibition of JAK2/STAT3 pathway activity, suggesting that hsa_circ_0058493 may be involved in arsenic-induced lung cell apoptosis by regulating the JAK2/STAT3 signaling pathway.

Arsenic  /  hsa_circ_0058493  /  Apoptosis  /  STAT3 signaling pathway
段婷芝, 杨雪飞, 陈斌, 杨净超, 杨欣达, 赵青云, 何越峰. 环状RNA hsa_circ_0058493在砷诱导肺细胞凋亡中的机制研究. 现代预防医学, 2026 , 53 (1) : 53 -61 . DOI: 10.20043/j.cnki.MPM.202505155
Ting-zhi DUAN, Xue-fei YANG, Bin CHEN, Jing-chao YANG, Xin-da YANG, Qing-yun ZHAO, Yue-feng HE. Mechanistic study of circular RNA hsa_circ_0058493 in arsenic-induced apoptosis of lung cells[J]. Modern Preventive Medicine, 2026 , 53 (1) : 53 -61 . DOI: 10.20043/j.cnki.MPM.202505155
砷是一种广泛存在于自然环境中的有毒类金属,长期暴露可导致呼吸系统损伤甚至肺癌等严重疾病[1-2]。流行病学数据表明,即使低至中等浓度(<150 μg/L)的砷暴露也可显著增加肺癌发病与死亡风险[3],这已成为全球范围内重大的公共卫生问题。现有研究表明,砷可通过诱发氧化应激、破坏线粒体功能、引起内质网应激以及异常激活信号通路等多种机制诱导细胞凋亡[4-5]。其中,氧化应激介导的DNA损伤和p53通路激活等机制已较为明确[6],但非编码RNA[尤其是环状RNA(circRNA)]在砷诱导肺细胞凋亡过程中的调控作用与分子机制仍缺乏研究。
circRNA是一类具有共价闭合环状结构的RNA分子,其高稳定性和组织特异性表达特征使其在多种疾病的发生发展中发挥重要作用。hsa_circ_0058493是由RHBDD1基因的第4和第5外显子区域反向剪接形成,其序列长度为656 bp,定位于chr2:227729319-227732034[7]。目前已发现hsa_circ_ 0058493在多种疾病中发挥重要作用:在慢性粒细胞白血病中,hsa_circ_0058493通过分子海绵效应参与耐药表型调控,为疾病的诊疗提供潜在靶点[7];在肝细胞癌中,hsa_circ_0058493经m6A修饰及核质转运调控定位于细胞质,通过激活促癌信号通路加速肿瘤增殖与转移[8];hsa_circ_0058493在肺纤维化患者血清中高表达,提示该circRNA可能作为疾病的潜在诊断标志物[9]。目前尚未有研究探讨hsa_circ_0058493在砷暴露诱导肺细胞凋亡中的作用机制,hsa_circ_0058493在砷毒性中是否存在其他功能模式也值得深入探索。
JAK-STAT信号通路广泛参与免疫调控、细胞增殖与凋亡、肿瘤发生等过程[10]。已有研究证实,砷暴露可通过METTL3介导的SOCS3 m6A修饰、或直接激活受体酪氨酸激酶等途径,异常上调STAT3水平,进而影响细胞存活[11]。值得注意的是,近年研究发现circRNA常通过直接或间接调控JAK2/STAT3通路影响疾病进程,例如Circ_0072088通过miR-375激活JAK2/STAT3信号通路促进肝癌进展[12]。而下调CircRNA 104633可抑制JAK2/STAT3通路减轻痛风性关节炎炎症与高凝状态[13]。据此,我们推测hsa_circ_0058493可能通过调控JAK2/STAT3通路参与砷诱导的肺细胞凋亡过程。
基于上述研究背景,本研究以A549和BEAS-2B细胞为模型,通过体外实验明确NaAsO2暴露对hsa_circ_0058493表达的影响,探究其在砷诱导肺细胞凋亡中的功能及可能的分子机制,旨在填补相关研究空白、拓展砷毒理学研究维度,为砷致肺疾病的诊疗提供理论与实验依据。
人肺腺癌细胞系(A549细胞,中国科学院昆明动物研究所)、人支气管上皮细胞系(BEAS-2B细胞,中国科学院昆明动物研究所)。
恒温培养箱(美国Thermo Forma公司);酶标仪(美国Bio Tek公司);研究级荧光倒置显微镜(江西凤凰phenix光学股份有限公司);聚合酶链式反应(polymerase chain reaction,PCR)仪(美国Bio-Rad公司);实时荧光定量PCR反应仪(德国Roche公司);蛋白显影成像仪(上海嘉鹏科技有限公司);1640培养基、MEM培养基(美国Corning公司);亚砷酸钠(NaAsO2)(纯度≥99.0%,成都西亚试剂公司);一甲基胂酸(MMA)、二甲基胂酸(DMA)(纯度均≥99.0%,美国AccuStandard公司);逆转录试剂(康为世纪生物科技有限公司);RFect试剂(常州百代生物公司);siRNA片段(吉玛公司);β-actin单克隆抗体、c-Myc鼠单克隆抗体、STAT3鼠单克隆抗体、BCL-2鼠单克隆抗体、XIAP鼠单克隆抗体(Proteintech公司);p-JAK2(Y1007+Y1008)兔单克隆抗体、JAK2兔单克隆抗体、p-STAT3(Y705)兔单克隆抗体、CyclinD1兔单克隆抗体(杭州华安生物科技有限公司)。
用T25瓶培养A549和BEAS-2B细胞,恒温培养箱条件:5%CO2,37℃。A549细胞使用1640完全培养基(包含10%胎牛血清和1%青链霉素混合液),BEAS-2B细胞使用MEM完全培养基。每日观察细胞生长状态,当细胞生长密度达到80%左右时,使用胰酶消化细胞进行传代操作。
取对数生长期细胞,接种于96孔板,接种密度为3 000个/孔,培养26 h后加NaAsO2染毒处理,分别以终浓度为0、10、20、30、40、60和80 μmol/L的NaAsO2处理A549细胞,以终浓度为0、2、4、6、8和10 μmol/L的NaAsO2处理BEAS-2B细胞,设空白组,染毒48 h后加入CCK-8试剂,酶标仪测定吸光度值,计算细胞存活率,并计算两个细胞的NaAsO2半抑制浓度(half-maximal inhibitory concentration,IC50)。取对数生长期细胞,接种于6孔板,接种密度为2.0×105个/孔,培养26 h后分组染毒。以NaAsO2终浓度为10、20和30 μmol/L以及DMA(30 μmol/L)、MMA(30 μmol/L)分别对A549细胞进行染毒,以NaAsO2终浓度为2、4和6 μmol/L以及DMA(6 μmol/L)、MMA(6 μmol/L)分别对BEAS-2B细胞进行染毒,仅加培养液处理组作为对照组,染毒48 h后提取RNA。
根据1.4步骤将细胞接种于6孔板并分组染毒处理后,使用Trizol裂解液提取总RNA,使用逆转录试剂合成cDNA,反应条件为:26℃预热5 min,42℃孵育45 min,85℃孵育5 min,10℃冷却2 min。以上述制备的cDNA为模板,使用RT-qPCR法检测目的基因表达,内参基因为管家基因β-肌动蛋白(ACTB),条件为95℃预变性520 s;95℃变性10 s、60℃退火10 s、72℃延伸10 s,扩增40个循环,以2-△△ct计算目的基因的相对表达量,引物序列见表1。
取对数生长期细胞分别接种于96孔板(接种密度为2 000个/孔)或6孔板(接种密度为1.0×105个/孔),培养18 h,使用RFect转染试剂转染siRNA片段,序列见表2,分别设置对照(negative control,NC)组、siRNA#1组、siRNA#2组,转染24 h后换完全培养基,继续培养48 h进行后续实验。将细胞接种于6孔板转染后,同1.5步骤对hsa_circ_0058493表达进行检测,验证siRNA沉默效率。
接种96孔板,细胞密度:2 000个/孔,同1.6步骤转染后,检测细胞存活率。同上,将细胞接种于96孔板并转染后,以1 μl的JC-1(100X)试剂加入1 ml空白培养基的比例配制JC-1工作液,每孔加入60 μl的JC-1工作液,37℃孵育15 min,酶标仪测定荧光强度,计算红/绿荧光强度比。
接种6孔板(接种密度为1×105个/孔),同1.6步骤转染后,配置HO-PI工作液(HO与PI的比例为1∶1)对细胞进行染色,每孔加入1 ml的HO-PI工作液,常温避光孵育15 min,观察细胞凋亡情况用荧光显微镜,每个视野分别记录可见光、蓝色和红色荧光下细胞形态,拍照结束后选取3个视野计算不同组细胞凋亡率,凋亡率=(凋亡细胞数/视野内总细胞数)×100%。
将细胞接种于6孔板并转染后,加入蛋白酶抑制剂和裂解液提取细胞总蛋白,测定蛋白浓度并计算上样量。将蛋白样品与上样缓冲液混合,加热变性后进行SDS-PAGE电泳分离。电泳后将蛋白转移到PVDF膜上,使用快速封闭液封闭20 min。使用针对目标蛋白的一抗进行孵育,(p-JAK2、JAK2、p-STAT3、STAT3、c-Myc、CyclinD1、XIAP、BCL-2、β-actin)一抗4℃过夜孵育,之后加入二抗孵育2 h。使用化学发光检测试剂盒进行显影图像采集。对显影后的条带进行灰度值分析,通过灰度值计算蛋白相对表达量=目的蛋白灰度值/内参蛋白灰度值。
为探究下调hsa_circ_0058493对NaAsO2诱导肺细胞凋亡的调控作用,设置两个分组:(1)单独hsa_circ_0058493下调组(si-circ组):分别将细胞接种于96孔板或6孔板,A549/BEAS-2B细胞培养18 h后转染siRNA,24 h后换完全培养基,继续培养48 h;(2)hsa_circ_0058493下调+NaAsO2处理组(si-circ+NaAsO2组):A549/BEAS-2B细胞培养18 h后转染siRNA,24 h后更换含NaAsO2(A549:30 μmol/L;BEAS-2B:6 μmol/L)的培养基,继续培养48 h。培养结束后检测细胞存活率及目的蛋白表达。
采用SPSS软件对数据统计分析。计量资料满足正态分布,采用两独立样本t检验,多组间比较,采用单因素方差分析,并进一步两两比较。样本率的比较使用χ2检验。检验水准α=0.05。
在A549细胞中,与对照组(NaAsO2浓度=0 μmol/L)相比,NaAsO2浓度为10~80 μmol/L组,细胞存活率降低,差异具有统计学意义(10~80 μmol/L组:P<0.001),见图1A。在BEAS-2B细胞中,与对照组相比,NaAsO2浓度为2~10 μmol/L组,细胞存活率降低,差异具有统计学意义(2 μmol/L组:P=0.006,4~10 μmol/L组:P<0.001),见图1B。经计算,A549细胞IC50=35.03 μmol/L,95% CI:33.48~36.65 μmol/L。BEAS-2B细胞IC50=7.809 μmol/L,95% CI:7.349~8.341 μmol/L。后续将在此基础上选取适宜浓度进行实验。
用不同浓度NaAsO2及MMA、DMA处理A549和BEAS-2B细胞,结果显示,A549细胞中,与对照组相比,10、20和30 μmol/L NaAsO2处理组,hsa_circ_0058493表达升高,差异具有统计学意义(P=0.012、0.003、<0.001),见图2A。BEAS-2B细胞中,与对照组相比,2 μmol/L NaAsO2处理组hsa_circ_0058493表达无明显变化(P=0.204),4和6 μmol/L NaAsO2处理组hsa_circ_0058493表达升高,差异具有统计学意义(P=0.027、0.002),见图2B。MMA、DMA处理两组细胞后,hsa_circ_0058493表达与对照组相比均无明显变化(A549细胞:P=0.058、0.197;BEAS-2B细胞:P=0.676、0.909),见图2C、D。
使用siRNA下调hsa_circ_0058493,通过RT-qPCR检测hsa_circ_0058493相对表达量,结果显示,在A549和BEAS-2B细胞中,与对照组相比,siRNA#1组和siRNA#2组hsa_circ_0058493相对表达量显著降低,见图3A、B,差异具有统计学意义(A549细胞:P<0.001、<0.001;BEAS-2B细胞:P<0.001、<0.001)。
下调hsa_circ_0058493后,通过CCK-8实验对细胞活力进行检测,在A549细胞中,各组细胞活力分别为100%、62.5%、56.05%,BEAS-2B细胞中分别为100%、71.63%、53.34%。与对照组相比,siRNA#1组和siRNA#2组细胞活力降低,差异具有统计学意义(A549细胞:P<0.001、<0.001;BEAS-2B 细胞:P<0.001、<0.001),见图4A、B。通过JC-1实验检测细胞线粒体膜电位变化情况,结果表明下调hsa_circ_0058493组(siRNA#1、siRNA#2)线粒体膜电位和对照组相比降低,差异有统计学意义(A549细胞:P<0.001、<0.001;BEAS-2B 细胞:P=0.003、0.041),见图4C、D。经Hoechst 33342/PI双染后,与对照组相比,下调hsa_circ_0058493组呈现出更多亮蓝色和红色的细胞,见图5A、B,早期凋亡细胞和晚期凋亡细胞占比增加,见图5C、D,差异有统计学意义(P<0.001、<0.001;P<0.001、<0.001)。
与对照组相比,下调hsa_circ_0058493组(siRNA#1、siRNA#2),p-JAK2、JAK2、p-STAT3、STAT3蛋白相对表达量降低(图6A),差异具有统计学意义(P<0.05)(图6B、C)。此外,下调hsa_circ_0058493组(siRNA#1、siRNA#2)中c-Myc、XIAP、CyclinD1和BCL-2蛋白的相对表达量也呈现降低趋势(图6D),与对照组相比,差异具有统计学意义(P<0.05)(图6E、F)。
下调hsa_circ_0058493(siRNA#1、siRNA#2)后,A549和BEAS-2B细胞存活率与对照组相比均显著降低(P<0.05)。下调hsa_circ_0058493后给予NaAsO2处理,各组的细胞存活率较单独下调hsa_circ_0058493组进一步降低(P<0.05),见图7A、B。Western blot检测A549和BEAS-2B细胞中STAT3蛋白表达变化如图7C、D,在A549细胞中,与单独下调hsa_circ_0058493组相比,下调hsa_circ_0058493后给予NaAsO2处理时,NC、siRNA#1分组STAT3蛋白相对表达量无明显变化(P>0.05),siRNA#2分组STAT3蛋白相对表达量升高(P<0.05),见图7E。在BEAS-2B细胞中,上述处理后,NC分组STAT3蛋白相对表达量降低(P<0.05),siRNA#1和siRNA#2分组的STAT3蛋白相对表达量无明显变化(P>0.05),见图7F。
细胞凋亡是砷致肺损伤的关键过程,尽管氧化应激、DNA损伤等经典通路已被证实参与其中[14-15],但circRNA在这一过程中的作用仍有待深入探究。本研究发现hsa_circ_0058493可能通过调控STAT3通路参与砷诱导的肺细胞凋亡,这为揭示砷毒性的表观遗传调控机制提供了新的视角。我们的研究结果表明,经NaAsO2处理,A549和BEAS-2B细胞的存活率显著降低,且呈现出剂量关系,这与之前的研究结果一致,证实了砷的细胞毒性[16]。但本研究发现仅NaAsO2能够上调A549和BEAS-2B细胞中hsa_circ_0058493的表达,而MMA和DMA无显著作用,这与砷代谢的毒性差异一致,无机砷的细胞毒性显著高于有机砷代谢产物[17-18]。推测hsa_circ_0058493可能参与无机砷的早期应激响应,hsa_circ_0058493表达变化或可作为无机砷暴露的潜在生物标志物。为了研究hsa_circ_0058493在肺细胞中的功能,我们设计了特异性siRNA下调hsa_circ_0058493。
通过siRNA下调hsa_circ_0058493后,A549和BEAS-2B细胞活力显著降低,线粒体膜电位下降,细胞凋亡率增加。在wu等人[8]的研究中,也发现下调hsa_circ_0058493会增加肝癌细胞凋亡率,抑制细胞增殖。进一步机制研究显示,下调hsa_circ_0058493可降低JAK2/STAT3通路相关蛋白(p-JAK2、p-STAT3、JAK2、STAT3)及下游靶基因蛋白(c-Myc、CyclinD1、BCL-2等)的表达。而JAK/STAT3通路作为调控细胞增殖与凋亡的核心信号轴,其抑制与上述表型变化吻合[19-20],表明hsa_circ_0058493可能通过调控JAK2/STAT3通路影响细胞生理功能。
hsa_circ_0058493下调可进一步增强NaAsO2的细胞毒性,提示二者在抑制细胞存活方面存在效应叠加。值得注意的是,STAT3蛋白表达呈现细胞系特异性差异,这一现象可能与细胞背景相关。已有研究表明,不同类型肿瘤组织与正常组织间的STAT3表达存在显著差异,其中肺癌组织的STAT3表达水平高于正常肺组织[21]。据此推测,A549作为癌细胞,其STAT3基础活性较高,可能部分抵消hsa_circ_0058493下调所致的抑制效应;而BEAS-2B作为正常细胞,STAT3活性较低,更易受hsa_circ_0058493下调的主导影响。这种差异提示,hsa_circ_0058493对STAT3的调控可能并非单一直接路径,其复杂的作用网络有待进一步探究。
综上,本研究表明,下调hsa_circ_0058493可引起A549和BEAS-2B细胞活力下降、凋亡增加,这与JAK2/STAT3通路活性抑制相关,提示hsa_circ_0058493可能通过调控JAK2/STAT3通路参与砷诱导的肺细胞凋亡,为解析砷致肺损伤机制及相关干预提供了新视角。
  • 国家自然科学基金(82160607)
  • 云南省科技研究计划(202401AT070177)
  • 云南省创新团队(202405AS350016)
  • 昆明医科大学创新团队(CXTD202201)
  • 昆明医科大学硕士研究生教育创新基金(J114150330366)
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2026年第53卷第1期
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doi: 10.20043/j.cnki.MPM.202505155
  • 接收时间:2025-05-13
  • 首发时间:2026-11-02
  • 出版时间:2026-01-10
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  • 收稿日期:2025-05-13
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国家自然科学基金(82160607)
云南省科技研究计划(202401AT070177)
云南省创新团队(202405AS350016)
昆明医科大学创新团队(CXTD202201)
昆明医科大学硕士研究生教育创新基金(J114150330366)
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    云南省公共卫生与生物安全重点实验室,昆明医科大学公共卫生学院,云南 昆明 650500

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鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
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红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
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