Objective:To investigate the potential anti-tumor mechanisms of naphthoquinone compound shikonin(SKN)extracted from the root of Chinese herbal medicine plant lithospermum(Lithospermum erythro-rhizon Sieb.&Zucc.).Methods:We first observed that SKN treatment led to swelling and bubbles in HeLa cells that were similar to the phenotype of cell pyroptosis.Subsequently,the HeLa cells experienced a pyroptotic process with SKN,and this was then assessed using lactate dehydrogenase(LDH)release and propidium iodide(PI)/Hoechst double staining experiments.Pyroptosis is defined as gasdermin-mediated programmed necroptosis.To identify the potential pyroptosis machinery,two strategies were utilized that included a genome-wide clustered regularly interspaced short palindromic repeats(CRISPR)-associated protein 9 screening experiment and a pyroptosis reconstitution assay executed by each of the five known gas-dermins(GSDMA-E).Moreover,endogenous cleavage was also detected in a panel of tumor cell lines.Results:Compared with the control,both the LDH release and PI/Hoechst double-staining experiments suggested that SKN induced perforation and enhancement of the permeability of the cell membranes that resulted in pyroptosis in HeLa cells(P=.028 and P=.032,respectively).In addition,the reconsti-tution assays in human embryonic kidney 293T(HEK-293T)cells and endogenous cleavage assays in HeLa cells indicated that the pyroptosis was controlled by GSDME.In addition,we also found SKN could trigger pyroptosis in a panel of tumor cell lines in which the cellular morphologies were proportional to the GSDME expression levels.Additionally,the cleavage of GSDME was also detected,and this was indicative of a similar GSDME-mediated mechanism.Conclusion:Our study not only explained the molecular mechanism of cytotoxicity of SKN to various tumor cells,but also provided additional information for the potential clinical application of natural naphthoquinone compounds against cancer.

Dongxiao Cui;Sanjiao Wang;Jiajian Guo;Mingrui Yang;Yunqian Li;Yue Zhang;Wenfu Ma

School of Life Sciences,Beijing University of Chinese Medicine,Beijing,102488,China

中医科学杂志（英文）

[1]Dongxiao Cui;Sanjiao Wang;Jiajian Guo;Mingrui Yang;Yunqian Li;Yue Zhang;Wenfu Ma-.Shikonin from Chinese herbal medicine induces GSDME-controlled pyroptosis in tumor cells)[J].中医科学杂志（英文）,2022(04):432-442

Shikonin,naphthoquinone,lithospermum,rhizon,pyroptotic,dermins,GSDMA,reconsti

from,Chinese,herbal,medicine,induces,GSDME,controlled,pyroptosis,tumor,cells,Objective,To,investigate,potential,anti,mechanisms,shikonin,SKN,extracted,root,plant,Lithospermum,erythro,Sieb,Zucc,Methods,We,first,observed,that,treatment,swelling,bubbles,HeLa,were,similar,phenotype,Subsequently,experienced,process,this,was,then,assessed,using,lactate,dehydrogenase,LDH,release,propidium,iodide,Hoechst,double,staining,experiments,Pyroptosis,defined,gasdermin,mediated,programmed,necroptosis,identify,machinery,two,strategies,utilized,included,genome,wide,clustered,regularly,interspaced,short,palindromic,repeats,CRISPR,associated,protein,screening,reconstitution,executed,by,each,five,known,Moreover,endogenous,cleavage,also,detected,panel,lines,Results,Compared,both,suggested,induced,perforation,enhancement,permeability,membranes,resulted,respectively,In,assays,human,embryonic,kidney,293T,HEK,indicated,found,could,trigger,which,cellular,morphologies,proportional,expression,levels,Additionally,indicative,Conclusion,Our,study,only,explained,molecular,cytotoxicity,various,but,provided,additional,information,clinical,application,natural,compounds,against,cancer

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