Azurocidin is associated with dienogest-resistance in ovarian endometriotic cysts

Azurocidin is associated with dienogest-resistance in ovarian endometriotic cysts | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Azurocidin is associated with dienogest-resistance in ovarian endometriotic cysts Momoe Watanabe, Yoichi Kobayashi, Manami Ishida, Atsushi Tajima, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3991408/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Endometriosis, a common estrogen-dependent inflammatory disease, and ovarian endometriotic cysts (ECs) cause dysmenorrhea and infertility. Currently available hormonal therapies for EC treatment, are sometimes ineffective. The signaling of progesterone receptors can downregulate estrogen receptors and therefore are involved in endometriosis. Dienogest (DNG) is an oral progestin that is highly selective for progesterone receptors (PRs). In our previous proteomics study, we identified that azurocidin could be associated with DNG resistance. In this study, we conducted using the human immortalized endometriotic epithelial Emosis-CC/TERT1 cell line, and examined the effect of azurocidin on ECs and its clinical significance. We examined whether azurocidin suppresses the action of DNG on IL-6 and IL-8 in ECs. We also examined whether azurocidin affects PR or ER in ECs. This study was conducted using the human immortalized endometriotic epithelial Emosis-CC/TERT1 cell line. DNG inhibited the production of IL-6 and IL-8, but this effect was suppressed in the presence of azurocidin. Inflammatory cytokines IL-6 and IL-8 enhanced azurocidin production. Furthermore, azurocidin increased estrogen receptor (ER) expression; the proliferation of EMosis-CC/TERT1 cells increased significantly when incubated with 17β-estradiol and azurocidin. The results suggest that azurocidin inhibits the action of DNG by increasing estrogen sensitivity by promoting ER mRNA expression and promoting endometriosis in endometrial epithelial cells. Thus, azurocidin may be associated with dienogest-resistance in ovarian ECs. Endometriosis dienogest progesterone receptors estrogen receptors azurocidin Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Full Text Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3991408","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":276015677,"identity":"c8928f90-7541-4e1f-8531-f8ed0a09ab78","order_by":0,"name":"Momoe Watanabe","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA7klEQVRIiWNgGAWjYPACGwMGBjYQQwImwkZISxrpWg4bEKEICnTbuxMffGw7byzfwJYm8XGPRR6DRALjhx8MfHm4tJidObvZcGbbbTODA2zHJGc8kygGamGW7GFgK8ap5UbuNmnebbeB/mdvk+Y5IJG4/0YCgzTQmYkN+LT83XbORr4BqqUBaMtvgloYtx0wYwA6DKaFDb8tIL/0/ks2NjjMlmw5A6SF52GbZY8BHr8c79344McZO8P57W2GNz4cqEtsYE8+fONHxTGcIYYAzAws0HhkBDrJ4FgCYS1ATR+QODVEaRkFo2AUjIIRAQCHU1OYSniyiQAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0009-0001-8203-7565","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":true,"prefix":"","firstName":"Momoe","middleName":"","lastName":"Watanabe","suffix":""},{"id":276015678,"identity":"eb6eddf3-ba5b-4707-b1d8-494d81028c9a","order_by":1,"name":"Yoichi Kobayashi","email":"","orcid":"","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":false,"prefix":"","firstName":"Yoichi","middleName":"","lastName":"Kobayashi","suffix":""},{"id":276015679,"identity":"fb68bcee-3909-4cb3-8346-504fbc8eefef","order_by":2,"name":"Manami Ishida","email":"","orcid":"","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":false,"prefix":"","firstName":"Manami","middleName":"","lastName":"Ishida","suffix":""},{"id":276015680,"identity":"781d1b4e-33ea-4951-84f1-7fd8dc468e9f","order_by":3,"name":"Atsushi Tajima","email":"","orcid":"","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":false,"prefix":"","firstName":"Atsushi","middleName":"","lastName":"Tajima","suffix":""},{"id":276015681,"identity":"e5986b19-9aec-48f0-8c01-ddcec235afe6","order_by":4,"name":"Shinji Tanigaki","email":"","orcid":"","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":false,"prefix":"","firstName":"Shinji","middleName":"","lastName":"Tanigaki","suffix":""},{"id":276015682,"identity":"a9a928ea-71f5-4f05-a378-0745fe8c5805","order_by":5,"name":"Tohru Morisada","email":"","orcid":"","institution":"Kyorin University Faculty of Medicine: Kyorin Daigaku Igakubu Daigakuin Igaku Kenkyuka","correspondingAuthor":false,"prefix":"","firstName":"Tohru","middleName":"","lastName":"Morisada","suffix":""}],"badges":[],"createdAt":"2024-02-26 16:09:24","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-3991408/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-3991408/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":52059441,"identity":"965a04ef-b3d7-4d0d-873e-87eafece076b","added_by":"auto","created_at":"2024-03-06 04:52:29","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":241233,"visible":true,"origin":"","legend":"\u003cp\u003eExpression of azurocidin in EMosis CC/TERT1 cells\u003c/p\u003e\n\u003cp\u003eEMosis CC/TERT1 cells were incubated with normal rabbit IgG (a) or anti-azurocidin antibody ( Scale bar 200 μ m\u003c/p\u003e","description":"","filename":"fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/656ae34664f7d0c7153cf8e0.png"},{"id":52060453,"identity":"8a03cdfa-d216-40ef-8e74-1cf2290fa714","added_by":"auto","created_at":"2024-03-06 05:08:29","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":99834,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of azurocidin on mRNA levels of IL 6 and IL 8 in EMosis\u003c/p\u003e\n\u003cp\u003eCC/TERT1 cells, evaluated using RT PCR.\u003c/p\u003e\n\u003cp\u003eThe expression\u003c/p\u003e\n\u003cp\u003eof each gene was normalized to that of GAPDH. Each value\u003c/p\u003e\n\u003cp\u003eis expressed as the mean ± SD of three independent experiments (N 3).\u003c/p\u003e\n\u003cp\u003eDNG treatment decreased the mRNA levels of IL 6 and IL 8 The mRNA\u003c/p\u003e\n\u003cp\u003eleve ls of IL 6 and IL 8 increased in cells incubated with azurocidin and DNG\u003c/p\u003e\n\u003cp\u003ecompared to those in cells incubated with DNG alone a). IL 6 mRNA\u003c/p\u003e\n\u003cp\u003eexpression increased when cells were incubated with 200 ng/m L azurocidin\u003c/p\u003e\n\u003cp\u003efor 48 h (b). * P \u0026lt;0.05.\u003c/p\u003e","description":"","filename":"fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/38a54aa48d8d66f38a30c6de.png"},{"id":52059437,"identity":"ce6cede0-0ba2-4490-ad3e-3813a30ceb3d","added_by":"auto","created_at":"2024-03-06 04:52:29","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":59376,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of IL 6 and IL 8 on azurocidin mRNA levels in EMosis\u003c/p\u003e\n\u003cp\u003eCC/TERT1 cells\u003c/p\u003e\n\u003cp\u003eThe\u003c/p\u003e\n\u003cp\u003emRNA levels of azurocidin in EMosis CC/TERT1 cells were evaluate\u003c/p\u003e\n\u003cp\u003eusing\u003c/p\u003e\n\u003cp\u003eusing RTRT--PCR. The expressionPCR. The expression of each gene of each gene waswas normalized normalized to thatto that of of GAPDH. Each value GAPDH. Each value representsrepresents the mean ± SD of four independent the mean ± SD of four independent experiments (N experiments (N == 4).4). Azurocidin mRNAAzurocidin mRNA levelslevels increased when the cells were increased when the cells were incubated with incubated with 1010 ng/mng/mLL and 100 ng/mand 100 ng/mLL of ILof IL--6 for 246 for 24 hh and with and with 1010 ng/mng/mLL and 100 ng/mand 100 ng/mLL of ILof IL--8 for 488 for 48 h. *P \u0026lt; 0.05 h. *P \u0026lt; 0.05.\u003c/p\u003e","description":"","filename":"fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/5a5a788bfeef6f6cf03ad8de.png"},{"id":52059439,"identity":"d6a2b75b-12e8-4da2-b74f-debbc28a0474","added_by":"auto","created_at":"2024-03-06 04:52:29","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":115248,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of azurocidin on PR and ER in EMosis CC/TERT1 cells\u003c/p\u003e\n\u003cp\u003eThe\u003c/p\u003e\n\u003cp\u003emRNA expression levels of PR and ER in EMosis CC/TERT1 cells were\u003c/p\u003e\n\u003cp\u003eevaluated using RT PCR. The expression levels of each gene were\u003c/p\u003e\n\u003cp\u003enormalized to those of GAPDH. Each value represents the mean ± SD of\u003c/p\u003e\n\u003cp\u003ethree independent experiments (N 3). (a) PR mRNA expression was not\u003c/p\u003e\n\u003cp\u003esignificantly affected when cells were incubated with azurocidin. (b) ER\u003c/p\u003e\n\u003cp\u003emRNA expression increased when cells were incubated with 200 ng/m L\u003c/p\u003e\n\u003cp\u003eazurocidin. *P \u0026lt; (c) ER expression was analyzed using SDS PAGE\u003c/p\u003e\n\u003cp\u003efollowed by immunoblotting using an anti ER α antibody. The figure shows\u003c/p\u003e\n\u003cp\u003ea representative blot from three independent experiments that y ielded similar\u003c/p\u003e\n\u003cp\u003eresults. The mean scanning densitometry values (expressed as arbitrary\u003c/p\u003e\n\u003cp\u003escanning units) of the bands are shown in the graph. Each value represents\u003c/p\u003e\n\u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n\u003cp\u003ethe mean ± SD of three independent experiments.\u003c/p\u003e\n\u003cp\u003ethe mean ± SD of three independent experiments. ERERαα levellevelss increased when increased when cells were incubated with 200ng/mcells were incubated with 200ng/mL L azurocidin. * P \u0026lt; 0.05.azurocidin. * P \u0026lt; 0.05.\u003c/p\u003e","description":"","filename":"fig4.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/a643e17dc6f59177e8f65a88.png"},{"id":52059624,"identity":"1465e6f5-1658-4672-88f5-4465cc8594e3","added_by":"auto","created_at":"2024-03-06 05:00:29","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":73066,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of E2 and azurocidin on EMosis CC/TERT1 cell number.\u003c/p\u003e\n\u003cp\u003eEMosis-CC/TERT1 cells were treated with E2 (10 7 M) or azurocidin (200\u003c/p\u003e\n\u003cp\u003eng/m L ) or E2 and a zurocidin for 120 h. Incubation of E M osis CC/TERT1\u003c/p\u003e\n\u003cp\u003ecells with E2 or azurocidin did not significantly affect their proliferation.\u003c/p\u003e\n\u003cp\u003eHowever , the incubation of EMosis CC/TERT1 cells both E2 and azurocidin\u003c/p\u003e\n\u003cp\u003esignificantly promoted their proliferation. Each value represents the mean ±SD of three independent experiments (N=3). * P \u0026lt; 0.05\u003c/p\u003e","description":"","filename":"fig5.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/b40b212be31790c252ddaf0d.png"},{"id":52059442,"identity":"3617e7de-f033-4c58-a3b0-3d08832c4959","added_by":"auto","created_at":"2024-03-06 04:52:29","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":175419,"visible":true,"origin":"","legend":"\u003cp\u003eSummary of this study\u003c/p\u003e","description":"","filename":"fig6.png","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1/c056373b6e31617aef17de41.png"},{"id":52942284,"identity":"6f7ddfcf-25db-4399-83fc-cdfc083e4dc0","added_by":"auto","created_at":"2024-03-19 01:32:26","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":634631,"visible":true,"origin":"","legend":"","description":"","filename":"Text.pdf","url":"https://assets-eu.researchsquare.com/files/rs-3991408/v1_covered_4820646f-3132-479f-92e1-e7d9ccc4e2ec.pdf"}],"financialInterests":"","formattedTitle":"Azurocidin is associated with dienogest-resistance in ovarian endometriotic cysts","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Endometriosis, dienogest, progesterone receptors, estrogen receptors, azurocidin","lastPublishedDoi":"10.21203/rs.3.rs-3991408/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-3991408/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"Endometriosis, a common estrogen-dependent inflammatory disease, and ovarian endometriotic cysts (ECs) cause dysmenorrhea and infertility. Currently available hormonal therapies for EC treatment, are sometimes ineffective. The signaling of progesterone receptors can downregulate estrogen receptors and therefore are involved in endometriosis. Dienogest (DNG) is an oral progestin that is highly selective for progesterone receptors (PRs). In our previous proteomics study, we identified that azurocidin could be associated with DNG resistance. In this study, we conducted using the human immortalized endometriotic epithelial Emosis-CC/TERT1 cell line, and examined the effect of azurocidin on ECs and its clinical significance.\nWe examined whether azurocidin suppresses the action of DNG on IL-6 and IL-8 in ECs. We also examined whether azurocidin affects PR or ER in ECs. This study was conducted using the human immortalized endometriotic epithelial Emosis-CC/TERT1 cell line. DNG inhibited the production of IL-6 and IL-8, but this effect was suppressed in the presence of azurocidin. Inflammatory cytokines IL-6 and IL-8 enhanced azurocidin production. Furthermore, azurocidin increased estrogen receptor (ER) expression; the proliferation of EMosis-CC/TERT1 cells increased significantly when incubated with 17β-estradiol and azurocidin. The results suggest that azurocidin inhibits the action of DNG by increasing estrogen sensitivity by promoting ER mRNA expression and promoting endometriosis in endometrial epithelial cells. Thus, azurocidin may be associated with dienogest-resistance in ovarian ECs.","manuscriptTitle":"Azurocidin is associated with dienogest-resistance in ovarian endometriotic cysts","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-03-06 04:52:25","doi":"10.21203/rs.3.rs-3991408/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"3f3dec6d-7683-4f0b-b456-b1255817f010","owner":[],"postedDate":"March 6th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-03-19T01:24:13+00:00","versionOfRecord":[],"versionCreatedAt":"2024-03-06 04:52:25","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-3991408","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3991408","identity":"rs-3991408","version":["v1"]},"buildId":"B-jG_2CBjPDmsCi4Wdhf-","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}