Name: methods for studying uterine contributions to pregnancy establishment in an ovariectomized mouse model
Uploaded: 2023-04-07T12:50:00
Description: Watch this Scientific Journal Video about Methods for Studying Uterine Contributions to Pregnancy Establishment in an Ovariectomized Mouse Model at JoVE.com

这项工作是通过维多利亚州政府运营基础设施支持和澳大利亚政府国家卫生和医学研究委员会（NHMRC）IRIISS实现的。这项工作得到了莫纳什大学医学，护理和健康科学平台访问补助金的支持，以访问A.L.W.（WINSHIP-PAG18-0343）以访问莫纳什生殖服务平台。A.L.W. 得到了澳大利亚研究理事会 （ARC） 的 DECRA 资助 DE21010037 的支持。J.N.H.和L.R.A.由澳大利亚政府研究培训计划奖学金支持。L.R.A.由莫纳什研究生卓越奖学金支持。K.J.H.由ARC未来奖学金FT190100265支持。

所有动物都被饲养在温度控制，高屏障设施（莫纳什大学动物研究实验室）中，免费提供食物和水，并进行12小时的明暗循环。所有程序均根据莫纳什动物研究平台伦理委员会（#21908，17971）的批准执行，并按照国家卫生和医学研究委员会动物护理和使用业务守则执行。
1. 手术准备
<ol><li>在121°C的硬/干货循环中高压灭菌手术所需的所有手术器械，纱布和纸巾，?...

<ol>
	<li>Szamatowicz, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assisted+reproductive+technology+in+reproductive+medicine+-+Possibilities+and+limitations.">Assisted reproductive technology in reproductive medicine - Possibilities and limitations.</a> Ginekologia Polska. 87 (12), 820-823 (2016).</li><li>Evans, J., 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=Fertile+ground:+Human+endometrial+programming+and+lessons+in+health+and+disease.">Fertile ground: Human endometrial programming and lessons in health and disease.</a> Nature Reviews. Endocrinology. 12 (11), 654-667 (2016).</li><li>Norwitz, E. R., Schust, D. J., Fisher, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Implantation+and+the+survival+of+early+pregnancy.">Implantation and the survival of early pregnancy.</a> The New England Journal of Medicine. 345 (19), 1400-1408 (2001).</li><li>Zinaman, M. J., Clegg, E. D., Brown, C. C., O'Connor, J., Selevan, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimates+of+human+fertility+and+pregnancy+loss.">Estimates of human fertility and pregnancy loss.</a> Fertility &amp; Sterility. 65 (3), 503-509 (1996).</li><li>Kupka, M. 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=Assisted+reproductive+technology+in+Europe,+2010:+Results+generated+from+European+registers+by+ESHRE&#8224;.">Assisted reproductive technology in Europe, 2010: Results generated from European registers by ESHRE&#8224;.</a> Human Reproduction. 29 (10), 2099-2113 (2014).</li><li>Gleicher, N., Kushnir, V. A., Barad, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Worldwide+decline+of+IVF+birth+rates+and+its+probable+causes.">Worldwide decline of IVF birth rates and its probable causes.</a> Human Reproduction Open. 2019 (3), (2019).</li><li>Diaz-Gimeno, 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=A+genomic+diagnostic+tool+for+human+endometrial+receptivity+based+on+the+transcriptomic+signature.">A genomic diagnostic tool for human endometrial receptivity based on the transcriptomic signature.</a> Fertility &amp; Sterility. 95 (1), 50-60 (2011).</li><li>Amin, J., 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=Personalized+embryo+transfer+outcomes+in+recurrent+implantation+failure+patients+following+endometrial+receptivity+array+with+pre-implantation+genetic+testing.">Personalized embryo transfer outcomes in recurrent implantation failure patients following endometrial receptivity array with pre-implantation genetic testing.</a> Cureus. 14 (6), e26248 (2022).</li><li>Patel, J. A., Patel, A. J., Banker, J. M., Shah, S. I., Banker, 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=Personalized+embryo+transfer+helps+in+improving+in+vitro+fertilization/ICSI+outcomes+in+patients+with+recurrent+implantation+failure.">Personalized embryo transfer helps in improving in vitro fertilization/ICSI outcomes in patients with recurrent implantation failure.</a> Journal of Human Reproductive Sciences. 12 (1), 59-66 (2019).</li><li>Khan, K. 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=Biological+differences+between+functionalis+and+basalis+endometria+in+women+with+and+without+adenomyosis.">Biological differences between functionalis and basalis endometria in women with and without adenomyosis.</a> European Journal of Obstetrics, Gynecology, and Reproductive Biology. 203, 49-55 (2016).</li><li>Richards, J. S., Ren, Y. A., Candelaria, N., Adams, J. E., Rajkovic, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ovarian+follicular+theca+cell+recruitment,+differentiation,+and+impact+on+fertility:+2017+update.">Ovarian follicular theca cell recruitment, differentiation, and impact on fertility: 2017 update.</a> Endocrine Reviews. 39 (1), 1-20 (2018).</li><li>Corciulo, C., 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=Pulsed+administration+for+physiological+estrogen+replacement+in+mice.">Pulsed administration for physiological estrogen replacement in mice.</a> F1000Research. 10, 809 (2021).</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>Griffiths, M. J., Alesi, L. R., Winship, A. L., Hutt, K. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+an+embryo+transfer+model+to+study+uterine+contributions+to+pregnancy+in+vivo+in+mice.">Development of an embryo transfer model to study uterine contributions to pregnancy in vivo in mice.</a> Reproduction &amp; Fertility. 3 (1), 10-18 (2022).</li><li>Cousins, F. L., 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=Evidence+from+a+mouse+model+that+epithelial+cell+migration+and+mesenchymal-epithelial+transition+contribute+to+rapid+restoration+of+uterine+tissue+integrity+during+menstruation.">Evidence from a mouse model that epithelial cell migration and mesenchymal-epithelial transition contribute to rapid restoration of uterine tissue integrity during menstruation.</a> PLoS One. 9 (1), e86378 (2014).</li><li>Cousins, F. L., 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=Androgens+regulate+scarless+repair+of+the+endometrial+&amp;#34;wound&amp;#34;+in+a+mouse+model+of+menstruation.">Androgens regulate scarless repair of the endometrial &amp;#34;wound&amp;#34; in a mouse model of menstruation.</a> FASEB Journal. 30 (8), 2802-2811 (2016).</li><li>Fullerton, P. T., Monsivais, D., Kommagani, R., Matzuk, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Follistatin+is+critical+for+mouse+uterine+receptivity+and+decidualization.">Follistatin is critical for mouse uterine receptivity and decidualization.</a> Proceedings of the National Academy of Sciences of the United States of America. 114 (24), E4772-E4781 (2017).</li><li>Rowland, R. R., Reyes, E., Chukwuocha, R., Tokuda, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Corticosteroid+and+immune+responses+of+mice+following+mini-osmotic+pump+implantation.">Corticosteroid and immune responses of mice following mini-osmotic pump implantation.</a> Immunopharmacology. 20 (3), 187-190 (1990).</li><li>Barton, B. 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=Roles+of+steroid+hormones+in+oviductal+function.">Roles of steroid hormones in oviductal function.</a> Reproduction. 159 (3), R125-R137 (2020).</li><li>Lee, J. 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=Autophagy+regulates+embryonic+survival+during+delayed+implantation.">Autophagy regulates embryonic survival during delayed implantation.</a> Endocrinology. 152 (5), 2067-2075 (2011).</li><li>Hamatani, T., 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=Global+gene+expression+analysis+identifies+molecular+pathways+distinguishing+blastocyst+dormancy+and+activation.">Global gene expression analysis identifies molecular pathways distinguishing blastocyst dormancy and activation.</a> Proceedings of the National Academy of Sciences of the United States of America. 101 (28), 10326-10331 (2004).</li><li>Cui, L., 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=Transcervical+embryo+transfer+in+mice.">Transcervical embryo transfer in mice.</a> Journal of the American Association for Laboratory Animal Science. 53 (3), 228-231 (2014).</li></ol>

本文提供了有关如何进行 OVX 的分步说明，并为专注于了解子宫对怀孕和生育能力的贡献的研究提供外源性激素。在这些方法的两个实验应用中提供了两个详细的方案，包括进行胚胎移植和人工诱导蜕膜化。
虽然最初执行OVX可能具有挑战性 - 特别是对于刚接触啮齿动物模型的研究人员 - 但一旦经过适当的训练和实践，这是一个相对简单的过程。手术中的关键步骤包括在麻醉下...

近几十年来，随着辅助生殖技术的使用越来越多，许多受孕障碍已被克服，尽管存在生育问题，但仍允许许多夫妇组建家庭1。卵母细胞或精子缺陷通常可以通过 体外 受精或卵胞浆内单精子注射来绕过;然而，与子宫和子宫内膜容受性有关的问题仍然是生殖潜力难以捉摸的“黑匣子”2。
当高质量的胚胎成功地与感受性子宫内膜（子宫内膜）相互作用时，就建立了怀孕。在任何给定的月经周期中成功怀孕的机会都很低，约为 30%3，4。在成功的妊娠中，只有50%-60%的人在妊娠20周后提前，植入失败是75%未达到20周的妊娠的原因3。尽管这些数字可以追溯到1990年代后期，但该领域尚未克服子宫内膜接受能力不足造成的限制。这导致近年来试管婴儿成功率停滞不前 - 有时甚至下降 - 5，6。
患有不明原因不孕症的女性通常有一个移位的接受窗口，或者由于未知原因无法实现接受。最近，开发了子宫内膜容受性阵列，该阵列评估了数百个基因的表达，目的是根据个体的接受性窗口调整胚胎移植的时间7，8，9。然而，该领域仍然缺乏对植入过程完成后出现的妊娠并发症的发病机制的理解。
女性生殖系统是高度动态的，并受到严格的荷尔蒙控制。下丘脑-垂体-性腺（HPG）轴控制黄体生成素和卵泡刺激素的释放，它们调节卵巢周期的各个方面，包括卵泡成熟以及雌激素和孕激素活性。反过来，子宫月经周期由雌激素和黄体酮10，11调节。因此，研究子宫生物学机制因卵巢的影响而变得复杂。例如，在研究癌症疗法如何影响子宫时，很难区分观察到的任何子宫表型（例如妊娠丢失或月经不合时宜）是对子宫的直接侮辱还是卵巢损伤的间接影响。
为了全面了解生育能力，必须表征子宫对怀孕的贡献。重要的是，这种理解必须超越卵巢控制下的子宫功能。这不能在人类身上研究;因此，经常使用动物模型。因此，卵巢切除术（OVX）通常用于使研究人员能够通过外源性提供激素来调节啮齿动物发情周期（类似于月经周期）。此外，OVX允许独立于卵巢影响12研究子宫反应。然而，如果在OVX后没有立即提供激素，就会出现更年期表型，这需要研究人员仔细考虑。
OVX经常用于啮齿动物模型13，14，15，16，17，并且在经过充分训练后相对容易执行。方法取决于是单独切除卵巢还是切除卵巢和输卵管，以及动物的年龄（成年，循环动物的卵巢较大，表面有可见的黄体，这意味着它们的卵巢更容易可视化）。同样，存在许多激素补充方法，包括皮下注射14，缓释微丸15，渗透压微型泵18和卵巢移植。
在本文中，提供了有关如何进行卵巢切除术和准备三种激素补充剂的详细说明，包括皮下注射、缓释丸和渗透迷你泵。为受益于OVX的实验终点提供了两种详细的方案，然后是外源性激素补充（胚胎移植和人工蜕膜化）。本文讨论了每种方法的优缺点，目的是指导研究人员如何进行研究以隔离对子宫的影响，特别是在怀孕和生育研究领域。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>ALZET 1002 mini osmotic pumps</td>
			<td>BioScientifica</td>
			<td>1002</td>
			<td>Delivers 0.25 &#181;L/h for 14 days. Use for section 7 (Experimental procedure - Embryo transfer).</td>
		</tr>
		<tr>
			<td>ALZET 1003D mini osmotic pumps</td>
			<td>BioScientifica</td>
			<td>1003D</td>
			<td>Delivers 1 &#181;L/h for 14 days. Use for section 8 (Experimental procedure - Artificial decidualization).</td>
		</tr>
		<tr>
			<td>ALZET Reflex 7 mm clips</td>
			<td>BioScientifica</td>
			<td>0009971</td>
			<td>Either Reflex clips or Michel clips can be used for wound closure, depending on preference</td>
		</tr>
		<tr>
			<td>ALZET Reflex clip applicator</td>
			<td>BioScientifica</td>
			<td>0009974</td>
			<td>Either Reflex clips or Michel clips can be used for wound closure, depending on preference</td>
		</tr>
		<tr>
			<td>ALZET Reflex clip remover</td>
			<td>BioScientifica</td>
			<td>0009976</td>
			<td>Either Reflex clips or Michel clips can be used for wound closure, depending on preference</td>
		</tr>
		<tr>
			<td>Bupivicaine injection</td>
			<td>Pfizer</td>
			<td>NA</td>
			<td>Stock 0.5%. Use at 0.05% in saline</td>
		</tr>
		<tr>
			<td>Estradiol</td>
			<td>Sigma</td>
			<td>E8875</td>
			<td></td>
		</tr>
		<tr>
			<td>Meloxicam</td>
			<td>Ilium</td>
			<td>NA</td>
			<td>Active constituent 0.5 mg/mL. Use 3.5 mL per 200 mL cage bottle, or as your institutions vet prescribes.</td>
		</tr>
		<tr>
			<td>Michel clips</td>
			<td>Daniels</td>
			<td>NS-000242</td>
			<td></td>
		</tr>
		<tr>
			<td>Multi purpose sealant</td>
			<td>Dow Corning</td>
			<td>732</td>
			<td></td>
		</tr>
		<tr>
			<td>Non-surgical embryo transfer (NSET) device</td>
			<td>ParaTechs</td>
			<td>60010</td>
			<td>Contains 6 mm speculum. Single use only.</td>
		</tr>
		<tr>
			<td>Progesterone</td>
			<td>Sigma</td>
			<td>P0130</td>
			<td>Soluble in ethanol. Use for&#160; section 3 (Hormone preparation - subcutaneous injection) and&#160; section 4 (Hormone preparation - slow-release pellets)</td>
		</tr>
		<tr>
			<td>Progesterone</td>
			<td>Sigma</td>
			<td>P7556</td>
			<td>Soluble in water. Use for section 5 (Hormone preparation - osmotic mini pumps)</td>
		</tr>
		<tr>
			<td>Refresh eye ointment</td>
			<td>Allergan</td>
			<td>NA</td>
			<td>42.5% w/v liquid paraffin, 57.3% w/v soft white paraffin</td>
		</tr>
		<tr>
			<td>Rimadyl Carprofen</td>
			<td>Zoetis</td>
			<td>NA</td>
			<td>Stock 50 mg/mL. Use at 5 mg/kg</td>
		</tr>
		<tr>
			<td>Rubber tubing</td>
			<td>Dow Corning</td>
			<td>508-008</td>
			<td>Washed in 100% ethanol and cut into 1 cm pieces. Inside diameter 1.57 mm &#177;&#160; 0.23 mm; outside diamater 3.18 mm &#177; 0.23 mm; wall 0.81 mm.</td>
		</tr>
		<tr>
			<td>Sesame oil</td>
			<td>Sigma</td>
			<td>S3547</td>
			<td></td>
		</tr>
		<tr>
			<td>Sofsilk Silk sutures size 3-0</td>
			<td>Covidien</td>
			<td>GS-832</td>
			<td></td>
		</tr>
	</tbody>
</table>

methods for studying uterine contributions to pregnancy establishment in an ovariectomized mouse model

为了确定怀孕，一个可行的囊胚必须成功地与接受性子宫内膜（子宫内膜）相互作用，以促进植入和胎盘形成，并使持续妊娠成为可能。胚胎缺陷引起的妊娠成功的局限性是众所周知的，并且随着 体外 受精（IVF）和辅助生殖技术的兴起，近几十年来已在很大程度上克服了这些限制。然而，到目前为止，该领域尚未克服子宫内膜接受不足造成的限制，从而导致试管婴儿成功率停滞不前。卵巢和子宫内膜功能紧密交织在一起，因为卵巢产生的激素负责子宫内膜的月经周期性。因此，当使用啮齿动物怀孕模型时，很难确定观察到的结果是由于卵巢或子宫缺陷。为了克服这个问题，开发了一种具有胚胎移植或人工蜕膜化的卵巢切除小鼠模型，以允许研究子宫对怀孕的特异性贡献。本文将提供有关如何进行卵巢切除术的说明，并提供有关提供外源性激素的各种技术的见解，以支持健康供体胚胎移植后成功的人工蜕膜化或怀孕。这些技术包括皮下注射、缓释微丸和渗透微型泵。将讨论每种方法的主要优点和缺点，使研究人员能够为其特定的研究问题选择最佳的研究设计。

本协议论文描述了一个特征良好的人工蜕膜化模型（图1A）。在这里，年轻的成年雌性小鼠（8周龄）接受了第1节和第2节所述的手术卵巢切除术。然后将小鼠休息2周，以确保内源性卵巢激素消散，然后再用外源性激素支持，如第3-7节和第9节所述。通过阴道内注射芝麻油诱导人工蜕膜化，然后将动物休息直至组织收集，如第9节所述。在这项研究中，在C57BL6 / J...

Watch this Scientific Journal Video about Methods for Studying Uterine Contributions to Pregnancy Establishment in an Ovariectomized Mouse Model at JoVE.com

Methods for Studying Uterine Contributions to Pregnancy Establishment in an Ovariectomized Mouse Model

怀孕的建立是一个动态的过程，涉及复杂的胚胎和子宫串扰。母体子宫环境对这些过程的确切贡献仍然是一个活跃的研究领域。在这里，提供了详细的协议来帮助设计 体内 动物模型来解决这些研究问题。

研究卵巢切除小鼠模型中子宫对妊娠建立的贡献的方法

For pregnancy to be established, a viable blastocyst must successfully interact with a receptive uterine lining (endometrium) to facilitate implantation ...

因此，很难区分子宫和卵巢荷尔蒙对怀孕的影响，因为两者密切相关。因此，该协议允许我们直接检查子宫贡献。虽然其中一些技术（如卵巢切除术）具有很强的侵入性，但它们在练习和训练后相对简单，这使得它们适合刚接触小鼠模型的研究人员。因此，这种方法可以用来理解HPG轴的更广泛调节，因为卵巢和子宫在该轴的调节中都起着重要作用。如果您以前从未执行过这些技术，请咨询您研究所的兽医团队以获得培训和支持。首先，在脊柱驼背下方的凹痕处剃出一小块区域后，轻轻地将其轻拍在眼睛上，将眼睛润滑剂大量涂抹在麻醉的小鼠上。将0.1毫升卡洛芬皮下注射到颈部的肚子中。通过捏住动物的后脚趾并检查反射来测试动物是否被麻醉。接下来，将Betadine应用于手术部位。然后用手术布覆盖它。使用鼠齿镊子，向上拉开驼背处的皮肤，并做一个五毫米的纵向切口。然后，使用钝钳将皮肤从下面的肌肉层向下解剖到一侧朝向肾脏。通过肌肉壁识别肾脏、卵巢和卵巢脂肪垫。用镊子抓住并抬起肌肉层。然后，用剪刀做一个0.5到1厘米长的切口，用一对钝钳将卵巢垫拉过切口。接下来，使用弯曲的持针器，夹在子宫角远端的卵巢和输卵管下方，并用剪刀切除卵巢。30 秒后，取下夹子并用无菌纱布轻拍。要关闭肌肉壁切口，请使用镊子抬起切口的顶部。然后使用真丝缝线将切口与外科医生的结绑在一起。使用一毫升注射器局部涂抹几滴布比卡因，没有针头连接，并用无菌纱布轻拍多余的布比卡因。在另一侧进行类似的卵巢切除术后，通过将皮肤的两侧压在一起并应用几个七毫米的手术夹来关闭皮肤切口，为伤口肿胀留出空间。将所有必需的设备放在层流或II类生物安全罩内的箔纸上，并进行紫外线消毒20分钟。用100%脱硫醇清洗硅橡胶管，使其在引擎盖内风干。干燥后，使用手术刀将管子切成一厘米长的段。接下来，在没有柱塞的情况下将大约200微升密封剂挤入注射器中。更换柱塞并推出少量密封剂。将密封剂涂在管子的一端，用戴手套的手指将其平滑。并让密封剂在引擎盖中干燥过夜或在紫外线下干燥 30 分钟。接下来，将黄体酮粉倒入无菌培养皿中。在镊子的帮助下将粉末舀成颗粒。将颗粒的密封端沿着罩表面轻敲以冷凝粉末。然后用密封剂密封开口端。将装有颗粒的培养皿包裹在箔纸中以保护它们免受光照。在皮下插入前约72小时，通过在木炭剥离的FCS中孵育来激活颗粒。展示这一程序的是哈德逊医学研究所的早期职业研究科学家Fiona Cousins博士。在用皮下雌二醇注射给动物激素灌注后，在人工蜕膜化前两天用皮下孕酮颗粒灌注动物。手术前，麻醉小鼠并通过检查脚趾捏反射来测试麻醉深度。然后将鼠标置于俯卧位置，抬起尾巴，慢慢将六毫米的窥器插入其阴道。将鼠标的下半身放在非惯用手的前两个手指之间。然后，用食指轻轻向上推尾巴，以便更好地进入阴道。将20微升芝麻油转移到非手术胚胎移植尖端中，并将尖端通过阴道和子宫颈插入子宫角。慢慢排出油。完成后，从阴道中取出窥器。在这项研究中，80%的小鼠成功进行了蜕膜化，而20%的小鼠没有。因此，在将非甾体抗炎药装置通过子宫颈插入小鼠的子宫角时，保持所有手术设备和工作区域无菌并让患者保持无菌非常重要。诱导人工蜕膜化后，可以去除黄体酮腭以模拟月经并研究子宫内膜分解。使用这种方法，我们可以研究子宫生物学和独立于卵巢影响的调节。这在生育和妇女健康领域确实令人兴奋。

Research

JoVE Journal

Developmental Biology

Assessment of Maternal Vascular Remodeling During Pregnancy in the Mouse Uterus

产妇血管重构的评估在怀孕期间在小鼠子宫

The placenta mediates the exchange of factors such as gases and nutrients between mother and fetus and has specific demands for supply of blood from the ...

科研

发育生物学

Development of an Ethanol-induced Fibrotic Liver Model in Zebrafish to Study Progenitor Cell-mediated Hepatocyte Regeneration

乙醇引起的肝纤维化模型的斑马鱼发展研究祖细胞介导的​​肝细胞的再生

Sustained liver fibrosis with continuation of extracellular matrix (ECM) protein build-up results in the loss of cellular competency of the liver, leading ...

A Novel Use of Three-dimensional High-frequency Ultrasonography for Early Pregnancy Characterization in the Mouse

三维高频超声对小鼠早期妊娠特征的新应用

High-frequency ultrasonography (HFUS) is a common method to non-invasively monitor the real-time development of the human fetus in utero. The mouse is ...

Establishment of Larval Zebrafish as an Animal Model to Investigate Trypanosoma cruzi Motility In Vivo

Establishment of Larval Zebrafish as an Animal Model to Investigate Trypanosoma cruzi Motility In Vivo

斑马鱼幼虫动物模型的建立研究锥虫运动性体内

Chagas disease is a parasitic infection caused by Trypanosoma cruzi, whose motility is not only important for localization, but also for cellular binding ...

An In Vitro Model of a Parallel-Plate Perfusion System to Study Bacterial Adherence to Graft Tissues

An In Vitro Model of a Parallel-Plate Perfusion System to Study Bacterial Adherence to Graft Tissues

平行板灌注系统的体外模型研究了细菌对移植组织的粘附性

Various valved conduits and stent-mounted valves are used for right ventricular outflow tract (RVOT) valve replacement in patients with congenital heart ...

An In Vivo Method to Study Mouse Blood-Testis Barrier Integrity

An In Vivo Method to Study Mouse Blood-Testis Barrier Integrity

小鼠血睾丸屏障完整性的一种活体研究方法

Spermatogenesis is the development of spermatogonia into mature spermatozoa in the seminiferous tubules of the testis. This process is supported by ...

Methods to Test Endocrine Disruption in Drosophila melanogaster

Methods to Test Endocrine Disruption in Drosophila melanogaster

检测果蝇黑色素的内分泌干扰的方法

In recent years there has been growing evidence that all organisms and the environment are exposed to hormone-like chemicals, known as endocrine disruptor ...

Assessment of Oxidative Damage in the Primary Mouse Ocular Surface Cells/Stem Cells in Response to Ultraviolet-C (UV-C) Damage

Assessment of Oxidative Damage in the Primary Mouse Ocular Surface Cells/Stem Cells in Response to Ultraviolet-C UV-C Damage

评估原小鼠眼部表面细胞/干细胞对紫外线-C （UV-C） 损伤的反应

The ocular surface is subjected to regular wear and tear due to various environmental factors. Exposure to UV-C radiation constitutes an occupational ...

An Enhanced Green Fluorescence Protein-based Assay for Studying Neurite Outgrowth in Primary Neurons

增强的绿色荧光蛋白测定，用于研究原神经元的神经质生长

Neurite outgrowth is a fundamental event in the formation of the neural circuits during nervous system development. Severe neurite damage and synaptic ...

Analysis of Congenital Heart Defects in Mouse Embryos Using Qualitative and Quantitative Histological Methods

使用定性和定量组织方法对小鼠胚胎先天性心脏缺陷的分析

Congenital heart defects (CHD) are the most common type of birth defect in humans, affecting up to 1% of all live births. However, the underlying causes ...