Name: a syngeneic murine model of endometriosis using naturally cycling mice
Uploaded: 2020-11-24T15:00:00
Description: Watch this Scientific Journal Video about A Syngeneic Murine Model of Endometriosis using Naturally Cycling Mice at JoVE.com

Vi vill tacka medlemmarna i Reizes-laboratoriet för deras kritiska granskning och insikter under utarbetandet av manuskriptet, samt imaging- och histologikärnorna vid Lerner Research Institute för deras hjälp med datainsamling och dataanalys. Detta arbete stöddes genom en intern bidragsfinansiering genom forskningsprogramkommittén vid Cleveland Clinic och genom ett externt bidrag genom Society for Reproductive Investigation och Bayer. Forskning i Reizes Laboratory finansieras också genom VeloSano Bike to Cure, Center of Research Excellence i gynekologisk cancer, och genom Laura J. Fogarty Endowed Chair for Uterine Cancer Research. Cleveland Clinic äger upphovsrättstillståndet för figur 1 och figur 2.

OBS: Användningen av djur i denna studie godkändes av Institutional Animal Care and Use Committee (IACUC) vid Cleveland Clinic Lerner Research Institute. Alla allmänt tillgängliga standarder för djurvård och användning utfördes enligt riktlinjer från National Institutes of Health. Denna procedur använder aseptiska tekniker. Petriskålen är steril. Pbs/saltlösningen som används är steril. De kirurgiska instrumenten för obduktion och vävnad dissekering steriliseras via autoklav. Vi använder 70% EtOH på i...

<ol>
	<li>Zondervan, K. T., Becker, C. M., Missmer, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endometriosis.">Endometriosis.</a> England Journal of Medicine. 382 (13), 1244-1256 (2020).</li><li>Schwartz, K., Llarena, N. C., Rehmer, J. M., Richards, E. G., Falcone, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+pharmacotherapy+in+the+treatment+of+endometriosis+across+the+lifespan.">The role of pharmacotherapy in the treatment of endometriosis across the lifespan.</a> Expert Opinion on Pharmacotherapy. 21 (8), 893-903 (2020).</li><li>Giudice, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+practice.+Endometriosis.">Clinical practice. Endometriosis.</a> New England Journal of Medicine. 362 (25), 2389-2398 (2010).</li><li>D'Hooghe, T. M., Debrock, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endometriosis,+retrograde+menstruation+and+peritoneal+inflammation+in+women+and+in+baboons.">Endometriosis, retrograde menstruation and peritoneal inflammation in women and in baboons.</a> Human Reproduction Update. 8 (1), 84-88 (2002).</li><li>Ahn, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathophysiology+and+immune+dysfunction+in+endometriosis.">Pathophysiology and immune dysfunction in endometriosis.</a> BioMed Research International. 2015, (2015).</li><li>Falcone, T., Flyckt, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clinical+management+of+endometriosis.">Clinical management of endometriosis.</a> Obstetrics and Gynecology. 131 (3), 557-571 (2018).</li><li>Vercellini, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Association+between+endometriosis+stage,+lesion+type,+patient+characteristics+and+severity+of+pelvic+pain+symptoms:+a+multivariate+analysis+of+over+1000+patients.">Association between endometriosis stage, lesion type, patient characteristics and severity of pelvic pain symptoms: a multivariate analysis of over 1000 patients.</a> Human Reproduction. 22 (1), 266-271 (2007).</li><li>Bulun, S. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endometriosis.">Endometriosis.</a> Endocrine Reviews. 40 (4), 1048-1079 (2019).</li><li>Brueggmann, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+three-dimensional+in+vitro+models+of+ovarian+endometriosis.">Novel three-dimensional in vitro models of ovarian endometriosis.</a> Journal of Ovarian Research. 7, 17 (2014).</li><li>Dodds, K. N., Beckett, E. A. H., Evans, S. F., Hutchinson, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lesion+development+is+modulated+by+the+natural+estrous+cycle+and+mouse+strain+in+a+minimally+invasive+model+of+endometriosis.">Lesion development is modulated by the natural estrous cycle and mouse strain in a minimally invasive model of endometriosis.</a> Biology of Reproduction. 97 (6), 810-821 (2017).</li><li>Martinez, J., Bisbal, V., Marin, N., Cano, A., G&#243;mez, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+monitoring+of+lesion+size+in+a+heterologous+mouse+model+of+endometriosis.">Noninvasive monitoring of lesion size in a heterologous mouse model of endometriosis.</a> Journal of Visualized Experiments: JoVE. (144), (2019).</li><li>Pelch, K. E., Sharpe-Timms, K. L., Nagel, S. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+model+of+surgically-induced+endometriosis+by+auto-transplantation+of+uterine+tissue.">Mouse model of surgically-induced endometriosis by auto-transplantation of uterine tissue.</a> Journal of Visualized Experiments: JoVE. (59), e3396 (2012).</li><li>Nishimoto-Kakiuchi, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spontaneous+endometriosis+in+cynomolgus+monkeys+as+a+clinically+relevant+experimental+model.">Spontaneous endometriosis in cynomolgus monkeys as a clinically relevant experimental model.</a> Human Reproduction. 33 (7), 1228-1236 (2018).</li><li>Nair, H. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+efficient+model+of+human+endometriosis+by+induced+unopposed+estrogenicity+in+baboons.">An efficient model of human endometriosis by induced unopposed estrogenicity in baboons.</a> Oncotarget. 7 (10), 10857-10869 (2016).</li><li>Lagan&#224;, A. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Translational+animal+models+for+endometriosis+research:+a+long+and+windy+road.">Translational animal models for endometriosis research: a long and windy road.</a> Annals of Translational Medicine. 6 (22), 431 (2018).</li><li>Bellofiore, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=First+evidence+of+a+menstruating+rodent:+the+spiny+mouse+(Acomys+cahirinus).">First evidence of a menstruating rodent: the spiny mouse (Acomys cahirinus).</a> Amercian Journal of Obstetrics and Gynecology. 216 (1), 1-11 (2017).</li><li>Bruner-Tran, K. L., Mokshagundam, S., Herington, J. L., Ding, T., Osteen, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+models+of+experimental+endometriosis:+identifying+mechanisms+of+disease+and+therapeutic+targets.">Rodent models of experimental endometriosis: identifying mechanisms of disease and therapeutic targets.</a> Current Women's Health Reviews. 14 (2), 173-188 (2018).</li><li>Bilotas, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interplay+between+endometriosis+and+pregnancy+in+a+mouse+model.">Interplay between endometriosis and pregnancy in a mouse model.</a> PloS One. 10 (4), 0124900 (2015).</li><li>Peterse, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Of+mice+and+women:+a+laparoscopic+mouse+model+for+endometriosis.">Of mice and women: a laparoscopic mouse model for endometriosis.</a> Journal of Minimally Invasive Gynecology. 25 (4), 578-579 (2018).</li><li>Richards, E. G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=KLF11+is+an+epigenetic+mediator+of+DRD2/dopaminergic+signaling+in+endometriosis.">KLF11 is an epigenetic mediator of DRD2/dopaminergic signaling in endometriosis.</a> Reproductive Sciences. 224 (8), 1129-1138 (2017).</li><li>Jones, R. L., Lang, S. A., Kendziorski, J. A., Greene, A. D., Burns, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+a+Mouse+Model+of+Experimentally+Induced+Endometriosis+to+Evaluate+and+Compare+the+Effects+of+Bisphenol+A+and+Bisphenol+AF+Exposure.">Use of a Mouse Model of Experimentally Induced Endometriosis to Evaluate and Compare the Effects of Bisphenol A and Bisphenol AF Exposure.</a> Environmental Health Perspectives. 126 (12), 127004 (2018).</li><li>Greaves, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+mouse+model+of+endometriosis+mimics+human+phenotype+and+reveals+insights+into+the+inflammatory+contribution+of+shed+endometrium.">A novel mouse model of endometriosis mimics human phenotype and reveals insights into the inflammatory contribution of shed endometrium.</a> The American Journal of Pathology. 184 (7), 1930-1939 (2014).</li><li>Nothnick, W. B., Graham, A., Holbert, J., Weiss, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=miR-451+deficiency+is+associated+with+altered+endometrial+fibrinogen+alpha+chain+expression+and+reduced+endometriotic+implant+establishment+in+an+experimental+mouse+model.">miR-451 deficiency is associated with altered endometrial fibrinogen alpha chain expression and reduced endometriotic implant establishment in an experimental mouse model.</a> PloS One. 9 (6), 100336 (2014).</li></ol>

Vår studie visar att endometrios kan induceras tillförlitligt hos möss utan att kräva användning av ovariectomy och/eller överlevnad kirurgi, och att ectopic endometrial skador kan identifieras och kvantifieras med hjälp av en standardiserad undersökning av buken och histologic analys.
Många murin studier av endometrios använder kirurgiskt inducerad endometrios där givaren endometrium sutu...

Endometrios är en gåtfull sjukdom i det kvinnliga reproduktionssystemet med betydande ekonomiska och hälsomässiga bördor på kvinnor1,2. Etiologin av endometrios är inte helt klarlagt, och flera förklaringar har föreslagits inklusive coelomic metaplasi, embryonala Müllerian vilar, rekrytering av benmärg härledda stamceller och bakåtsträvande menstruation3. Medan flera aspekter av dessa föreslagna mekanismer kan vara inblandade, och ingen enskild förklaring kan förklara alla former av sjukdomen, är den ledande modellen av endometrios patogenes retrograd menstruation. Retrograd menstruation är passagen av menstruationsutflöde genom äggledarna och in i peritonealhålan; det uppskattas att 90% av menstruerande kvinnor regelbundet genomgår retrograd menstruation4,5. Med tanke på detta vanliga fenomen av bakåtsträvande menstruation, varför endometrios utvecklas endast i en delmängd av kvinnor är oklart5. För att bättre förstå etiologin av denna sjukdom är direkta mänskliga studier inte genomförbara och djurstudier är motiverade.
Endometrios är en utmaning både att behandla och studera. Prevalensen av sjukdomen är inte känd men uppskattas till 10%1. Medan vissa avancerade typer av endometrios kan identifieras noggrant genom noninvasive imaging, uppnås en slutgiltig diagnos endast genom histopatologisk analys av kirurgiskt erhållna biopsi exemplar. lesioner som visuellt verkar vara sjuka, kan i själva verket vara fibros eller ärrbildning från andra orsaker6. Sjukdomens svårighetsgrad och omfattning korrelerar inte med symptomatologi7.
Endometriosskador består av heterogena celltyper och populationer som interagerar på komplexa sätt inom mikromiljön, vilket begränsar nyttan av cellulära modeller8,9. In vivo-modeller finns, men dessa har inneboende utmaningar ochbegränsningar 10,11,12. Primatmodeller är idealiska men är ofta integenomförbara 13,14,15. Få icke-primat däggdjur menstruerar och utvecklar endometrios spontant16. Gnagare modeller av endometrios finns men var och en har begränsningar17. Många av dessa modeller kräver överlevnadskirurgi för sutur eller implantat endometrievävnad i donatormottagaren vägg eller tarm, lägger till teknisk komplexitet, behov av anestesi och förvirrar immunfaktorer från själva operationen18,19,20. Dessutom kräver många modeller ovariectomy och östrogen tillskott; samtidigt öka lesion avkastning, detta lägger tid, kostnader och ytterligare överlevnad kirurgi. Intraperitoneal (IP) injektionsmodeller kräver inte anestesi eller överlevnad kirurgi, och dessa modeller simulerar logiskt bakåtsträvande menstruation bättre än suturing modeller21,22,23. De flesta IP-modeller, dock, är föremål för mer variation i lesion plats på grund av slumpmässig spridning av livmodercancer fragment efter injektion och därför för att mer partiskhet i lesion identifiering och mätning.
Här presenterar vi en murinmodell av endometrios som integrerar flera funktioner i befintliga endometriosmodeller i ett nytt, förenklat och effektivt system som förlitar sig på mikroskopisk kvantifiering i stället för subjektiv gradering.

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			<td>Supplies for injecting PMSG into donor mouse</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>1 mL Tuberculin syringe with 27G needle</td>
			<td>Fisher Scientific</td>
			<td>14-826-87</td>
			<td></td>
		</tr>
		<tr>
			<td>Pregnant mare serum gonadotropin</td>
			<td>Sigma-Aldrich</td>
			<td>9002-70-4</td>
			<td></td>
		</tr>
		<tr>
			<td>Supplies for necropsy of donor mouse and tissue processing</td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td>6&#8221; serrated forceps, curved tip</td>
			<td>Electron Microscopy Sciences</td>
			<td>72993-6C</td>
			<td></td>
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		<tr>
			<td>70% ethanol solution</td>
			<td>Pharmco</td>
			<td>33000HPLCCS4L</td>
			<td>70% solution dilute ethyl acetate 200 proof</td>
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			<td>Analytical balance</td>
			<td>Mettler Toledo</td>
			<td>ME54TE</td>
			<td></td>
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			<td>Carbon dioxide</td>
			<td>TriGas Supplier</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dissecting tray</td>
			<td>Fisher Scientific</td>
			<td>S14000</td>
			<td></td>
		</tr>
		<tr>
			<td>No. 10 disposable scalpel</td>
			<td>Fisher Scientific</td>
			<td>NC9999403</td>
			<td></td>
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		<tr>
			<td>Scissors, curved</td>
			<td>Electron Microscopy Sciences</td>
			<td>72941</td>
			<td></td>
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			<td>Scissors, straight</td>
			<td>Electron Microscopy Sciences</td>
			<td>72940</td>
			<td></td>
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		<tr>
			<td>Stereo microscope</td>
			<td>Leica Microsystems</td>
			<td>Leica SE 4</td>
			<td>For tissue dissection</td>
		</tr>
		<tr>
			<td>Sterile phosphate buffered saline (PBS)</td>
			<td>Institutional core facility supplies</td>
			<td></td>
			<td></td>
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		<tr>
			<td>Surgical instrument sterilization tray</td>
			<td>Electron Microscopy Sciences</td>
			<td>66112-02</td>
			<td></td>
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		<tr>
			<td>Tissue culture dishes</td>
			<td>Fisher Scientific</td>
			<td>08-772E</td>
			<td></td>
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		<tr>
			<td>Weighing dishes</td>
			<td>Fisher Scientific</td>
			<td>02-202-103</td>
			<td></td>
		</tr>
		<tr>
			<td>Supplies for injecting into recipient mouse</td>
			<td></td>
			<td></td>
			<td></td>
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			<td>1 cc syringe</td>
			<td>BD Biosciences</td>
			<td>301025</td>
			<td></td>
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			<td>18 G needle</td>
			<td>Fisher Scientific</td>
			<td>148265d</td>
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			<td>200 uL pipette tip</td>
			<td>Fisher Scientific</td>
			<td>02-707-422</td>
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			<td>Double distilled water</td>
			<td>Institutional core facility supplies</td>
			<td></td>
			<td></td>
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		<tr>
			<td>Latex bulb</td>
			<td>Fisher Scientific</td>
			<td>03-448-21</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro cover glass slip</td>
			<td>VWR</td>
			<td>48366-067</td>
			<td></td>
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			<td>Microscope slide</td>
			<td>Fisher Scientific</td>
			<td>12-544-7</td>
			<td></td>
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		<tr>
			<td>Standard light microscope</td>
			<td>Leica Microsystems</td>
			<td>DM IL</td>
			<td>For evaluating vaginal cytology smears</td>
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		<tr>
			<td>Supplies for harvesting tissue from recipient mouse</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>10% Buffered formalin</td>
			<td>Fisher Scientific</td>
			<td>SF100-4</td>
			<td></td>
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		<tr>
			<td>Biopsy foam pads</td>
			<td>Fisher Scientific</td>
			<td>22-038-222</td>
			<td></td>
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			<td>Precision Digital Calipers</td>
			<td>Electron Microscopy Sciences</td>
			<td>62065-40</td>
			<td></td>
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			<td>Processing/embedding cassettes</td>
			<td>Fisher Scientific</td>
			<td>22-272416</td>
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a syngeneic murine model of endometriosis using naturally cycling mice

Endometrios är en ledande orsak till bäckensmärta och infertilitet. Det definieras av förekomsten av endometrievävnad på extrauterin platser. Utvecklingen av nya terapier och diagnostiska verktyg för endometrios har begränsats delvis på grund av utmaningar i att studera sjukdomen. Utanför primater menstruerar få däggdjur, och ingen utvecklar spontan endometrios. Gnagare modeller är populära men kräver konstgjord induktion av endometrios, med många använder antingen immunkomprometterade möss eller kirurgiskt inducerad sjukdom. Nyligen har mer uppmärksamhet lagts på modeller som involverar intraperitoneal injektion. Vi presenterar en murinmodell av endometrios som integrerar flera funktioner i befintliga endometriosmodeller i ett nytt, förenklat system som förlitar sig på mikroskopisk kvantifiering i stället för subjektiv gradering. I denna modell utför vi hormonell stimulering av donatormöss, intraperitoneal injektion, systematisk bukundersökning och vävnadsskörd och histologic kvantifiering som kan utföras och verifieras när som helst efter obduktion. Denna modell kräver minimala resurser och utbildning. kräver inte expertis från labbtekniker vid murinöverlevnadskirurgi eller vid identifiering av grova endometriotiska lesioner, kan användas i immunkomprometterade, immunkompetenta och/eller muterade möss, och på ett tillförlitligt sätt skapar endometriotiska skador som är histologiskt förenliga med mänskliga endometriotisk sjukdom.

För ett första bevis på konceptexperiment injicerades donatorendometrium från RFP möss i wildtype mottagare möss. H&E färgning visade histopatologisk bekräftelse av klassisk arkitektur av endometrios lesion (Figur 3A). Fluorescerande mikroskopi bekräftade att den observerade lesionen i fråga härrörde från givaren (figur 3B).
Det andra experimentet utfördes med hjälp av 10 wildtype C57BL/6J givare och 10 mottagare. Ytter...

Watch this Scientific Journal Video about A Syngeneic Murine Model of Endometriosis using Naturally Cycling Mice at JoVE.com

A Syngeneic Murine Model of Endometriosis using Naturally Cycling Mice

Många gnagare modeller av endometrios begränsas av teknisk komplexitet, reproducerbarhet och/eller behov av immunkomprometterade djur eller speciella reportermöss. Vi presenterar ett förenklat system av lesion induktion med hjälp av någon experimentell mus med ett oberoende verifierbart, objektiva poängsystem och utan krav på ovariectomy eller överlevnad kirurgi.

En syngeneisk murinmodell av endometrios med naturligt cyklande möss

Endometriosis is a leading cause of pelvic pain and infertility. It is defined by the presence of endometrial tissue in extrauterine locations. The ...

Our mirroring model of endometriosis is a simplified and efficient system that integrates the best features of existing models. And perhaps most importantly, relies on microscopic quantification in lieu of subjective grading. Modeling endometriosis can be difficult because there can be many confounding factors and sources of bias.In our approach, endometriosis is induced without ovariectomy or survival surgery. And the endometrial lesions are objectively quantified. These Innovations provide a highly reliable and reproducible animal model of endometriosis that can be used to further endometriosis disease research.To facilitate ongoing estrous cycling, place urine soaked bedding from a male mouse cage into the recipient female mouse cage periodically throughout development, and again at 72 hours before endometrial tissue transplant. When the donor female mice are between 22 to 24 days old, use a one milliliter syringe equipped with a 25 to 27 gauge needle to subcutaneously inject two international units of pregnant mare serum gonadotropin in 200 microliters of sterile saline into the lower mouse abdomen. 38 to 42 hours after injection, use dissecting scissors to make a shallow transverse incision through the skin and subcutaneous tissue of the euthanized mouse at the midline and use blunt traction to open the skin on either side of the incision.Before removing the uterus, trim the adjacent connective tissue and transact each uterine horn just below its respective fallopian tubes, then transect the cervix to allow removal of the entire uterus and block. Inspect the uterus carefully, removing any additional peripheral fat or connective tissue and place the uterus in a droplet of cold PBS on a Petri dish for weighing. Transect each horn across the uterine fundus as close to the fundus as possible and place the dish under a dissecting microscope.Placing one blade of the dissecting scissors inside the lumen of the first horn, cut along the major axis of the tube keeping in mind, which side is this serosa and which side is the epithelium. Then use forceps to grasp and peel away the endometrial layer and mince the tissue into 10 to 12, one by one millimeter fragments. Alternatively, mince the tissue into one by one millimeter pieces without separating off the myometrium.Then place the tissues into 500 microliters of saline in a new Petri dish and harvest the endometrium from the second uterine horn as demonstrated. For peritoneal delivery of the tissue fragments into the recipient animal, use the blunt end of a one milliliter syringe to collect the tissue pieces. A total volume of one milliliter of tissue and saline should be collected.Attach an 18 gauge needle to the syringe and gently depress the plunger to load the fluid into the needle. A mock injection into the Petri dish can be performed to ensure that all of the tissue will pass through the needle tip. Either before or after the injection, use a bulb syringe to administer 10 microliters of saline to obtain a vaginal smear of the recipient animal for estrous cycle documentation.Holding the syringe at a 45 degree angle, intraperitoneally inject the fragments. If fragments remain after the injection, draw an additional 200 microliters of saline into the syringe to allow the rest of the pragmatists to be injected. Once you're assured of no bleeding or complications, place the mouse back in the home cage with a normal diet.Approximately 21 days after fragment injection, spray the recipient mouse abdomen was 70%ethanol, intent the skin to allow creation of a superficial subcutaneous incision to bluntly open the abdomen. Perform a complete survey of the abdominal wall and peritoneum, pancreas and mesenteric fat and para uterine connective fat and tissue for gross lesions. Harvest each region of tissue entirely and place the dissected tissue samples into an appropriate labeled cassette for formalin fixation and processing according to standard protocols.Then section the formalin blocks onto two slides per tissue area at two uniform depths. To score the endometriosis lesions, scan the tissue sections at a 40 times magnification and archive the slides. After scanning, use an appropriate digital slide reading software program to define and mark the longest distance between the edges of each endometriosis lesion.A continuous lesion is defined by glands surrounded by stroma. The line does not necessarily traverse only the endometriosis tissue, but the two end points must be connected by continuous stroma and or glands. Make a second line, 90 degrees across the first line as demonstrated.If multiple non-contiguous lesions are encountered, give each their own X and Y measurements. Calculate the final score for each slide as the summation of the areas of each lesion and use the larger of the scores from the two sections on each slide as the final score for that tissue region. Then total the scores from each region to give the final microscopic score for that animal.In this representative sample, histopathologic analysis after donor endometrium injection revealed a classic architecture of endometriosis lesion. Fluorescent microscopy of the tissue sample confirmed that the lesion originated from the donor. It is important to note that the macroscopic examination for lesions alone is not sufficient for the quantification of the endometriosis disease burden as the gross lesions observed in these representative samples were ultimately determined not to be endometriosis upon histological examination.Collection of all the tissue within the designated regions reduces bias by allowing for a systematic, objective approach to lesion quantification. Once this reliable and reproducible model has been achieved, researchers can use it to investigate a multitude of endometriosis pathways and therapies.

Research

JoVE Journal

Medicine

Mouse Model of Surgically-induced Endometriosis by Auto-transplantation of Uterine Tissue

Musmodell av kirurgiskt inducerad Endometrios av Auto-transplantation av uterus Tissue

Endometriosis is a chronic, painful disease whose etiology remains unknown. Furthermore, treatment of endometriosis can require laparoscopic removal of ...

Murine Model of Wound Healing

Murinmodell sårläkning

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Murina Cervical Heart Transplantation Modell hjälp av en modifierad Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

A Murine Model of Subarachnoid Hemorrhage

En mus modell av subaraknoidalblödning

In this video publication a standardized mouse model of subarachnoid  hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of  Willis ...

A Preclinical Murine Model of Hepatic Metastases

En Prekliniska mus modell av levermetastaser

Numerous murine models have been developed to study human cancers and advance the understanding of cancer treatment and development. Here, a preclinical, ...

A "Patient-Like" Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis

En &quot;Patient-Like&quot; Orthotopic syngena musmodell för Hepatocellulär cancer metastas

The majority of cancer-related deaths are caused by the metastasis of the cancer rather than the primary tumor itself. Yet, the underlying mechanisms of ...

Injection of Syngeneic Murine Melanoma Cells to Determine Their Metastatic Potential in the Lungs

Injektion av syngena murina melanomceller för att bestämma deras metastatisk potential i lungorna

Approximately 90% of human cancer deaths are linked to metastasis.  Despite the prevalence and relative harm of metastasis, therapeutics for treatment or ...

Murine Distal Colostomy, A Novel Model of Diversion Colitis in C57BL/6 Mice

Murina distala kolostomi, en ny modell av avledning kolit i C57BL/6 möss

Diversion colitis (DC) is a frequent clinical condition occurring in patients with bowel segments excluded from the fecal stream as a result of a ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

Isometrisk kontraktilitet mätning av mus mesenterica artär med Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Murine Cervical Aortic Transplantation Model using a Modified Non-Suture Cuff Technique

Murina cervikal aorta transplantation modell med en modifierad icke-sutur manschetten teknik

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. ...