新建IVF实验室中挥发性有机化合物的浓度变化及其对鼠胚体外发育的影响

目的:探讨新建成的体外受精(IVF)胚胎实验室挥发性有机化合物(volatile organic compound,VOC)的浓度变化及不同浓度VOC环境对小鼠体外受精和胚胎发育的影响,同时比较目前常用的去除实验室内VOC方法的效率。方法:测定胚胎实验室装修后不同时间段的VOC浓度变化,比较不同浓度的VOC对小鼠体外受精率、2-细胞率和囊胚率的影响;比较排风扇抽风机、Coda空气净化器和单纯活性炭包去除室内VOC的效率。结果:装修后第1个月(M1)VOC浓度为384.00 ppb,显著高于对照组(199.00 ppb)(P0.05);装修后第2个月(M2)VOC浓度为279.50 ppb,与对照组相比差异具有统计学意义(P0.05);装修后第3个月(M3)VOC浓度为210.25 ppb,与对照组(206.00 ppb)相比差异无统计学意义(P0.05)。装修后1个月(M1)实验组体外受精率、2-细胞率和囊胚率分别为62.3%、71.5%、72.0%,显著低于对照组(91.8%、92.2%、93.0%),组间差异具有统计学意义(P0.05)。装修后3个月(M3)时受精率、2-细胞率和囊胚率(91.5%、90.7%、91.4%)与对照组(89.8%、90.1%、92.5%)相比差异无统计学意义(P0.05)。Coda空气净化器处理3个月后可去除室内60.15%的VOC,显著高于排风扇抽风(51.88%)和单纯活性炭吸附(28.95%)的去除效率。结论:实验室内高浓度的VOC会对小鼠体外受精胚胎造成严重的胚胎毒性,导致受精率、2-细胞率和囊胚率降低,胚胎质量下降和发育受阻;Coda空气过滤器是一种有效的去除实验室内VOC的方法。     

鼠胚实验在新建体外受精胚胎培养室质量控制中的作用

目的:检测新建体外受精实验室培养体系的可靠与否,是否符合人类胚胎体外培养的要求。方法:对昆明白小鼠进行超促排卵,手术获取昆明白小鼠雌雄配子进行体外受精和体内受精的胚胎;并进行体外培养观察两组鼠胚的受精率、2-细胞率、囊胚率及采用密度梯度离心法和上游法两种精液处理方法对小鼠体外受精胚胎发育情况的影响。结果:进行了30个周期的体内受精体外培养实验,共获卵1136枚;进行了35个周期的体外受精实验,共获卵1486枚。体内受精组获得的受精率(93.13±1.46)%和2-细胞率(94.52±1.67)%高于体外受精组的受精率(84.39±2.87)%和2-细胞率(87.80±3.21)%,两组间差异有统计学意义(P0.05)。密度梯度离心法处理精液后,其受精率(90.32±3.14)%和2-细胞率(88.54±2.28)%高于上游法的受精率(76.46±2.35)%和2-细胞率(82.67±2.62)%,两者差异有统计学意义(P0.05)。结论:小鼠卵母细胞体内受精和体外受精后均有高的成胚率,体内受精后进行体外培养优于体外受精后培养,采用密度梯度离心处理精液实施体外受精优于上游法,鼠胚实验可对新建胚胎培养室的培养体系进行评估。     

重组人白血病抑制因子对体外移植前鼠胚发育的影响

目的：观察重组人白血病抑制因子（rhLIF)对体外移植前鼠胚发育的影响。方法：将36只小鼠随机分成3组,每组12只。组I（体内对照）小鼠注射人绒毛膜促性腺激素（hCG)116-120h后处死,组Ⅱ及组Ⅲ（体外对照）小鼠注射hCG44-48h后处死,收集组Ⅱ及组Ⅲ的2细胞期鼠胚,组Ⅱ的鼠胚用人输卵管液（HTF）＋10％人血清培养,组Ⅲ的鼠胚用HTF＋10％人血清＋rhLIF(1000U/ml）培养,观察并记录各细胞期鼠胚发育的数目。结果：（1）组Ⅱ和组Ⅲ发育到4细胞期、8细胞期、桑椹胚阶段的鼠胚百分率（分别为93．4％、87．7％、75．0％和94．5％、91．2％、85．4％）相似,差异无显著性（P＞0．05）。（2）组Ⅱ发育到囊胚期、扩张囊胚期和孵出期的鼠胚百分率低于组Ⅲ（分别为48．1％、32．1％、18．4％和82．3％、59．7％、36．3％）,差异有显著性（P＜0．05）。（3）组I和组Ⅲ发育到囊胚期的鼠胚百分率（分别为86．0％和82．3％）相比,差异无显著性（P＞0．05）。结论：rhLIF对鼠胚的早期发育没有显著影响,但能促进移植前晚期鼠胚的生长、分化和孵出。     

白细胞介素对小鼠体外受精及早期胚胎发育的影响

本文就人白细胞介素（ＩＬ－１β、５、６）对小鼠的体外受精率、体外受精卵的发育率，以及从交配雌鼠体内回收的２细胞期胚（体内受精卵）的发育率带来的影响进行了探讨。ＩＬ－１β５０ｎｇ／ｍｌ、ＩＬ－５５ｎｇ－ｍｌ以上、ＩＬ－６０．５ｎｇ／ｍｌ以上时对小鼠的体外受精率有明显抑制（Ｐ＜０．０５）。体外受精卵序的发育仅ＩＬ－６实验组见到抑制，０．５ｎｇ／ｍｌ以上对４细胞期胚发育有抑制，５０ｎｇ／ｍｌ时对８细胞期胚到囊胚期的发育有抑制。对体内受精卵的发育ＩＬ－１β和ＩＬ－６实验组与对照组间均未见到差异，但ＩＬ－５对囊胚期的发育有促进。以上结果提示，子宫内膜异位症时，人腹腔液中存在的ＩＬ－１β、５、６由于其抑制受精率而造成不孕。     

子宫内膜异位症子宫内膜细胞对鼠胚体外早期发育的影响

子宫内膜异位症（EMs,内异症）与妇女不育有着密切的关系,约有25％～40％的不孕妇女患内异症,而内异症患者中,不育症约占30％～40％。为探讨子宫内膜异位症患者的子宫内膜是否对胚胎的发育产生不利的影响,本实验将两细胞期鼠胚与内异症妇女子宫内膜细胞体外共培养,观察其对鼠胚早期发育的影响。     

低氧对体外受精-胚胎移植胚胎发育潜能的影响

目的:探讨低氧环境(体积分数5%O2)对体外受精-胚胎移植(in vitro fertilization-em-bryo transfer,IVF-ET)中胚胎发育潜能及临床结局的影响。方法:将接受IVF-ET长方案治疗的265名不孕症患者随机分为研究组(n=156):患者取卵后受精及整个胚胎培养过程全部在三气培养箱(体积分数5%O2)中进行,对照组(n=109):患者取卵后受精及整个胚胎培养过程全部在常规培养箱(体积分数20%O2)内进行,所有患者均移植授精第2日或第3日胚胎。比较组间受精率、正常受精率、卵裂率、正常卵裂率、优质胚胎率、可用胚胎率、生化妊娠率、临床妊娠率和异位妊娠率。结果:组间患者年龄、不孕年限、体质量指数、基础性激素、获卵数、成熟卵数、授精至移植时间和移植胚胎数均无统计学差异(P>0.05)。研究组受精率(84.4%)、正常受精率(72.0%)、卵裂率(97.6%)、优质胚胎率(43.3%)和可用胚胎率(72.5%)均显著高于对照组(分别为80.8%、68.7%、96.1%、35.1%、59.5%)(P<0.01或P<0.05),研究组与对照组正常卵裂率(97.7%vs 98.0%)、生化妊娠率(50.0%vs 39.4%)、临床妊娠率(44.9%vs 35.8%)、异位妊娠率(8.6%vs 12.8%)均无统计学差异(P>0.05)。结论:低氧环境(5%O2)似乎能够提高胚胎的发育潜能,获得更多优质胚胎和可用胚胎。     

人子宫内膜细胞共培养体系对早期鼠胚体外发育的影响

目的:研究人子宫内膜共培养体系对早期鼠胚体外发育的影响及移植后的妊娠情况。方法:将2-细胞小鼠胚胎与人子宫内膜细胞进行体外共培养,对照组为无营养细胞的单纯培养液,每日在显微镜下观察胚胎的发育情况。将培养到囊胚期的胚胎移植回小鼠的子宫腔,观察着床情况。结果:共培养体系中68.3％的2-细胞胚胎发育至桑椹胚期,50.8％发育至囊胚期,囊胚的孵化率为36.7％,胚胎的着床率为25.0％。而对照组只有24.8％的2-细胞胚胎发育至桑椹胚期,11.4％到达囊胚期,且其中大部分为早期囊胚即停止发育。另外对照组细胞碎片出现早且多,卵裂球不均匀,胚形态差,移植后胚胎的着床率仅为3.1％。结论:人子宫内膜细胞共培养体系可以促进小鼠胚胎的体外发育,改善胚胎的质量,提高着床率。     

不育、习惯性流产妇女血清对2－细胞鼠胚体外生长的影响

本文报道２０例原发不育妇女、２５例习惯性流产妇女以及１１例正常生育妇女血清作ＢＷＷ培养基添加剂分别对昆明小白鼠的８１３个、１０８５个和３８１个２—细胞鼠胚进行体外培养的比较研究。结果显示：各组７２ｈ囊胚形成百分率分别为４８．７０±１３．５０％，３７．３２±２０．３５％和５９．０５±７．３３％。不育组、习惯性流产组囊胚形成率与正常生育组囊胚形成百分率经统计学处理分别为Ｐ＜０．０５，Ｐ＜０．０２，均有显著性差异。这些结果提示：部分不育与习惯性流产妇女血清中可能存在有抑制早期胚胎发育的因子，因此检测血清中鼠胚生长抑制因子，有助于分析免疫性妊娠失败的病因。     

不育、习惯性流产妇女血清对2－细胞鼠胚体外生长的影响

本文报道２０例原发不育妇女、２５例习惯性流产妇女以及１１例正常生育妇女血清作ＢＷＷ培养基添加剂分别对昆明小白鼠的８１３个、１０８５个和３８１个２—细胞鼠胚进行体外培养的比较研究。结果显示：各组７２ｈ囊胚形成百分率分别为４８．７０±１３．５０％，３７．３２±２０．３５％和５９．０５±７．３３％。不育组、习惯性流产组囊胚形成率与正常生育组囊胚形成百分率经统计学处理分别为Ｐ＜０．０５，Ｐ＜０．０２，均有显著性差异。这些结果提示：部分不育与习惯性流产妇女血清中可能存在有抑制早期胚胎发育的因子，因此检测血清中鼠胚生长抑制因子，有助于分析免疫性妊娠失败的病因。     

重组人促性腺激素对早期鼠胚发育的影响

目的:探讨重组人促性腺激素对小鼠早期胚胎发育的影响。方法:将35只昆明种小鼠随机分为7组,实验组注射一定剂量的重组人促卵泡激素(rhFSH 20 IU)结合不同剂量的重组人促黄体激素(rhLH)对小鼠进行超促排卵,对照组注射孕马血清(PMSG10 IU)和hCG(10 IU),受精0.5 d处死各组雌鼠,收集2-细胞期的鼠胚进行体外培养,观察并记录鼠胚发育至各细胞期的数目。结果:10 IU和15 IU rhLH 组收集的胚胎数与对照组无差异,20 IU rhLH组则高于对照组,但20 IU rhLH组鼠胚发育至4-细胞、8-细胞、桑椹胚、囊胚、扩张期囊胚和孵出期的比率却较低,与对照组比有统计学差异,其余各组收集的胚胎数和胚胎发育至各细胞期数均低于对照组。结论:合适剂量的rhLH对小鼠早期胚胎的发育有明显的促进作用,但剂量过高则对胚胎发育有抑制作用。     

The Effect of Endometrial Polyps on Outcomes of In Vitro Fertilization (IVF) Cycles

Purpose: Our purpose was to investigate the effect of endometrial polyps on pregnancy outcome in an in vitro fertilization (IVF) program. Methods: Endometrial polyps less than 2 cm in diameter were suspected by transvaginal ultrasound before oocyte recovery in 83 patients. Forty-nine women (Group I) had standard IVF–embryo transfer, while in 34 women (Group II) hysteroscopy and polypectomy were performed immediately following oocyte retrieval, the suitable embryos were all frozen, and the replacement cycle took place a few months later. Results: Of the 32 hysteroscopies, a polyp was diagnosed in 24 cases (75%) and polypoid endometrium in another 5 patients (15.6%). An endometrial polyp was confirmed by histopathological examination in 14 women (58.3%). The pregnancy rate in group I was similar to the general pregnancy rate of our unit over the same period (22.4 vs 23.4%) but the miscarriage rate was higher (27.3 vs 10.7%, P = 0.08). In Group II, the pregnancy and miscarriage rates were similar to those of the frozen embryo cycles at Bourn Hall (30.4 and 14.3 vs 22.3 and 12.1%, respectively). Conclusions: Small endometrial polyps, less than 2 cm, do not decrease the pregnancy rate, but there is a trend toward increased pregnancy loss. A policy of oocyte retrieval, polypectomy, freezing the embryos, and replacing them in the future might increase the take-home baby rate.     

Improvement of mouse embryo quality by myo-inositol supplementation of IVF media

Objective

Myo-inositol (myoIns) has a positive role in mammalian development and human reproduction. Since experiments on farming species suggest a similar role in preimplantation development, we evaluated the hypothesis that the inclusion of myoIns in human embryo culture media would produce an increase in embryo quality in IVF cycles, using the mouse embryo assay.

Methods

To determine the effect of myoIns on completion of preimplantation development in vitro, one-cell embryos of the inbred C57BL/6N mouse strain were produced by ICSI, cultured in human fertilization media in the presence of myoIns (myoIns+) or in its absence (myoIns–) and evaluated morphologically. Daily progression through cleavage stages, blastocyst production and expansion and blastomere number at 96 hours post fertilization were assessed.

Results

Compared to myoIns– embryos, myoIns+ embryos displayed a faster cleavage rate and by the end of preimplantation development, the majority of myoIns+ blastocysts was expanded and formed by a higher number of blastomeres.

Conclusion

The presence of myoIns resulted in both an increase in proliferation activity and developmental rate of in vitro cultured early mouse embryos, representing a substantial improvement of culture conditions. These data may identify myoIns as an important supplement for human embryo preimplantation culture.     

干预治疗对子宫内膜形态不良者体外受精-胚胎移植结局的影响

目的:探讨干预治疗对超声检查显示子宫内膜形态不良的不孕患者体外受精-胚胎移植(IVF-ET)结局的影响。方法:采用回顾性对照研究,选择近排卵期子宫内膜超声检查形态不良者并行子宫内膜搔刮的干预性治疗后至内膜回声正常的患者30例(34周期)为干预组,子宫内膜回声不良而未行干预性治疗的患者37例(40周期)为未干预组;另选择同期子宫内膜形态正常的不孕患者51例(53周期)为对照组。比较3组患者的临床特征、内膜厚度、卵巢反应和妊娠结局。结果:干预组30例患者中,19例干预治疗1次,2例干预治疗4次,其余9例干预治疗2~3次后至内膜形态基本恢复正常。3组患者的基本临床特征、内膜厚度、卵巢反应、受精数以及移植胚胎数等比较,差异均无统计学意义(P0.05)。干预组的胚胎种植率、周期临床妊娠率(27.1%,50.0%)均高于未干预组(13.4%,25.0%),差异有统计学意义(P0.05),但与对照组(19.4%,35.8%)相比,差异无统计学意义(P0.05)。未干预组的胚胎种植率、周期临床妊娠率低于对照组,但差异无统计学意义(P0.05)。结论:超声检查提示子宫内膜形态不良的患者行干预治疗至内膜回声正常后有利于提高子宫内膜容受性,改善IVF-ET治疗结局。     

小鼠及人胚胎分割技术的研究

植入前小鼠胚胎（ｐｒｅｉｍｐｌａｎｔａｔｉｏｎ ｅｍｂｒｙｏ）的透明带经软化后,用固定在显微操作臂上的微玻璃针行胚胎分割,２－、４－、８－细胞胚胎及桑椹胚分割成功率分别为９０．６％、８５．０％、７３．６％、７１．４％。所获半胚体外培养后,胚泡发育率分别为６２．１％、８０．１％、８３．０％、８９．２％。随胚胎细胞数目增多,分割成功率降低,而发育率则提高。将８－细胞胚组及桑椹胚组的半胚移植到假孕母鼠后,幼仔出生率分别为１４．６％、１５．１％。同法分割４－细胞人胚胎２个,获半胚４个,分割成功率为１００％。     

体外受精胚胎冷冻储存及其移植的应用

本文的目的在于说明冷冻储存胚胎，适时解冻移植可以增加体外受精胚胎移植的妊娠成功率。第一例应用－Ｔｕｒｎｅｒ’ｓ综合征患者，丈夫无精症，在激素替代疗法下移植冻融胚胎，妊娠，于１９９５年２月６日分娩健康女婴，追踪检查生长发育良好。另外２例在超排卵刺激下获多个卵子及体外受精胚胎当月移植未获妊娠。而分别于５个月及６个月后在正常周期中移植自己的冻融胚胎，临床妊娠成功，妊娠在继续中，一例为双胞胎，三例都采用在２～８分裂球阶段冷冻。细胞内冷冻保护剂为丙二醇，细胞外保护剂为蔗糖。Ｔｅｓｔａｒｔ慢冻过程中诱发冰晶步骤很重要，可以控制溶液中的冰晶形成，避免细胞损害。     

Cytogenetic Analysis of Embryos Generated from In Vitro Matured Mouse Oocytes Reveals an Increase in Micronuclei Due to Chromosome Fragmentation

Purpose : (i) To determine the prevalence of micronuclei in the cytoplasm of embryos generated from in vitro matured oocytes. (ii) Assess whether micronuclei presence are the result of chromosome fragmentation or the loss of whole chromosomes. Methods : In vitro fertilization was performed on mature oocytes generated from superovulated mice (control) and in vitro matured mouse oocytes. Fertilized oocytes were cultured to the two-cell stage and fixed to slides. Micronuclei assessment was performed after staining with Giemsa. Centromere assessment was made using immunofluorescent staining (CREST) of the centromeric kinetochores. Results : Micronuclei were observed in 2% (4/197) of control two-cell embryos and 36.2% (46/127) of two-cell embryos generated from in vitro matured oocytes (P < 0.02). Centromeres were not detected in micronuclei from either group. Conclusions : A significant increase in micronuclei was observed in embryos generated from in vitro matured oocytes. The lack of accompanying centromeres would suggest the micronuclei are the result of chromosome fragmentation.     

Impact of short-term preconceptional exposure to particulate air pollution on treatment outcome in couples undergoing in vitro fertilization and embryo transfer (IVF/ET)

Purpose  

To assess the potential effects of short-term exposure to particulate air pollution during follicular phase on clinical, laboratory, and pregnancy outcomes of women undergoing IVF/ET.