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Endometrium Tissue Culture for In Vitro Embryo Implantation

机译：子宫内膜组织培养用于体外胚胎植入

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Introduction: Recent progress of in vitro fertilization and infertility treatment technology has much improved the infertility caused by ovary or ovulation. However, the functional recoverable treatment for infertility caused by uterus or after embryo transplantation is still unclear. In vivo embryo transplantation after blastocysts is necessary for the mammal embryo development, so that mammal embryo is not easy to be observed for continual change of development unlike reptile or fish embryos. In this study, in vitro embryo development model to investigate embryogenesis mechanism and to improve the infertility treatment has been developed by co-culture of embryo and endometrium in mouse model. Mouse blastocyst was cultured from ova in vitro and also endometrium tissue was cultured for implantation. Then, embryo adhesion onto the endometrium was observed histologically. Materials and Methods: Mouse blastocysts were prepared by IVF technique. Endometrium tissue was isolated and cultured in DMEM containing 10% FBS, progesterone, and estradiol. Change of endometrium to secretory phase by the hormones was confirmed histologically by HE staining. Additionally, the tissue was observed by Live/dead staining to confirm the cultured tissue viability. The cultured embryo and endometrium tissue were co-cultured in microtube with DMEM with 10% FBS and hormones for in vitro implantation. After co-culture for 2 days, the endometrium were observed by HE staining. Some blastocysts were cultured on collagen gel instead of endometrium in F0 medium. The collagen gel was diluted in DMEM and F-12 mixed medium with 30% FBS. Also, zona-freed blastocysts were culture in IVC1 and IVC2 medium on tissue culture dish, referred to Bedzhov's research. Result and discussion: Embryo adhesion and extension of trophoblasts were observed after 72 hour of culture on collagen gel. The culture of zona-freed blastocysts on tissue culture dish generated egg cylinders after 48 hour, but not after egg cylinder. In the endometrium culture, Live/dead staining confirmed the endometrium after 3 days of culture still survive. Also, HE staining indicated a change of endometrium by the hormones to secretory phase. Adhesion of embryo onto endometrium was observed in the co-culture of embryo and endometrium tissue, however embryo implantation could not be confirmed. Conclusion: Adhesion of embryo onto endometrium was observed by co-culture of embryo and endometrium tissue. However, the experiments could not confirm embryo implantation in vitro. Further progress may be necessary for clear observation of the embryo implantation on cultured endometrium tissue.
机译：简介：体外受精和不育治疗技术的最新进展大大改善了由卵巢或排卵引起的不育。然而，对于子宫或胚胎移植后的不育症的功能可恢复治疗仍不清楚。胚泡后的体内胚胎移植对于哺乳动物的胚胎发育是必需的，因此与爬行动物或鱼胚胎不同，不容易观察到哺乳动物胚胎的持续发育变化。在这项研究中，通过在小鼠模型中胚胎和子宫内膜的共培养，建立了用于研究胚胎发生机理和改善不育症治疗的体外胚胎发育模型。从卵中体外培养小鼠胚泡，并且还培养子宫内膜组织以进行植入。然后，通过组织学观察胚胎粘附到子宫内膜。材料与方法：采用IVF技术制备小鼠胚泡。分离子宫内膜组织，并在含有10％FBS，孕酮和雌二醇的DMEM中培养。通过HE染色在组织学上证实了激素引起的子宫内膜向分泌期的变化。另外，通过活/死染色观察组织以确认培养的组织的生存力。将培养的胚胎和子宫内膜组织与含10％FBS和激素的DMEM在微管中共培养​​，用于体外植入。共培养2天后，通过HE染色观察子宫内膜。在F0培养基中，将某些胚泡培养在胶原蛋白凝胶上而不是子宫内膜上。将胶原蛋白凝胶在DMEM和具有30％FBS的F-12混合培养基中稀释。另外，将游离带状胚泡在组织培养皿上的IVC1和IVC2培养基中培养，请参见Bedzhov的研究。结果与讨论：在胶原凝胶上培养72小时后，观察到了滋养细胞的胚胎粘附和伸长。在组织培养皿上培养无透明带的胚泡培养物在48小时后产生了蛋筒，但在蛋筒后没有。在子宫内膜培养中，活/死染色证实培养3天后子宫内膜仍能存活。同样，HE染色表明子宫内膜由激素改变为分泌期。在胚胎和子宫内膜组织的共培养中观察到了胚胎对子宫内膜的粘附，但是无法确认胚胎植入。结论：通过胚胎与子宫内膜组织的共培养观察到了胚胎对子宫内膜的粘附。但是，实验无法确定体外胚胎植入。为了清楚地观察在培养的子宫内膜组织上的胚胎植入，可能需要进一步的进展。

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