ABSTRACT
The influence of inner cell mass (ICM) and trophectoderm (TE) score on pregnancy outcomes in frozen-thawed blastocyst transfer cycles was analyzed. A retrospective analysis of 741 cycles of frozen-thawed blastosysts transfer was performed. All cycles were divided into four groups based on the number and morphological score of blastocysts: S-ICM B/TE B group (n=91), the single blastocyst transfer of ICM B and TE B; D-ICM B/TE B group (n=579), double blastocysts transfer of ICM B/TE B; D-ICM B/TE C group (n=35), double blastocysts transfer of ICM B/TE C; and D-ICM C/TE B group (n=36), double blastocysts transfer of TE B/ICM C. The pregnancy outcomes were compared among the four groups. As compared with D-ICM B/TE C group, the clinical pregnancy rate, implantation rate and multiple pregnancy rate were increased in D-ICM B/TE B group (74.96% vs. 57.14%, 57.43% vs. 37.14%, and 48.62% vs. 25%, respectively, P<0.05 for all). Clinical pregnancy rate and implantation rate in D-ICM B/TE B group were also higher than in D-ICM C/TE B group (74.96% vs. 50%, and 57.43% vs. 33.33%, both P<0.05). Multivariable Logistic regression analysis indicated that ICM score was a better predictive parameter for clinical pregnancy (OR=3.05, CI 1.70-5.46, P<0.001), while the trophectoderm score was a better one for early abortion (OR=0.074, CI 0.03-0.19, P<0.001). Clinical pregnancy rate and multiple pregnancy rate in S-ICM B/TE B group were significantly lower than those in D-ICM B/TE B group (46.15% vs. 74.96%, and 2.38% vs. 48.62%, both P<0.05), but there was no significant difference in the implantation rate between the two groups. It was suggested that the higher score of ICM and TE may be indicative of the better pregnancy outcomes. The ICM score is a better predictor of clinical pregnancy than TE, while TE score is a better one in predicting early abortion. Single ICM B/TE B blastocyst transfer in frozen-thawed cycles can also get satisfactory pregnancy outcomes.
Subject(s)
Adult , Female , Humans , Pregnancy , Analysis of Variance , Blastocyst , Cell Biology , Blastocyst Inner Cell Mass , Cell Biology , Cryopreservation , Methods , Embryo Implantation , Embryo Transfer , Methods , Fertilization in Vitro , Methods , Pregnancy Outcome , Pregnancy Rate , Retrospective StudiesABSTRACT
Embryonic stem cells [ESCs] are pluripotent cells derived from the inner cell mass [ICM] of blastocysts. A feeder layer and cytokines are necessary for culture of these embryonic cells in most species. The aim of this study was application of rat mesenchymal stem cells [MSCs] as a feeder layer for the isolation and culture of mouse embryonic stem cells. Mesenchymal stem cells were isolated from rat bone marrow and cultured in DMEM [Dulbecco's modified Eagle's medium] medium supplemented with 10% FBS [fetal bovine serum]. To verify the isolated cells, they were affected by osteocyte differentiation inducer to become bone mass. After twenty-one days, the differentiation was evaluated by Alizarin red staining. Blastocysts were obtained from Balb/c pregnant mice and cultured on this MSCs feeder layer. Two days later; after hatching of blastocysts, the cells were trypsinized and the inner cell mass dissociated to the small cell clumps. These clumps were cultured on 12-well plates covered by the same MSCs without applying any cytokines or growth inducer. Two to three days after the passage, colonies appeared which were similar to embryonic stem cell colonies in morphology. These colonies were passaged two more times using the mentioned procedure and their identities were examined by morphological observation and alkalin phosphatase staining. In this study we could easily cultured MSCs using DMEM media. The mesenchimic origin of cultured cells, which showed fibroblastic morphology, was proved by differentiation to bone masses using osteocyte inducer and detection with Alizarin red. By applying DMEM media and MSCs cells, as feeder layer, we could culture ESC without any need to cytokines or growth factors. After passage to the inner cell mass colonies were formed. These colonies were formed in two more other passages. The colonies were verified with alkaline phosphatase assay. Results of this study showed mesenchymal stem cells isolated from rat bone marrow can differentiate to osteoblast line and can be used as feeder layer for isolation, culture and forming embryonic stem cells colonies. This method, by using MSCs as feeder layer and bypassing the need of cytokine and growth factors, seems to be a simple efficient method for culture and isolation of embryonic stem cells
Subject(s)
Humans , Animals, Laboratory , Embryonic Stem Cells , Pluripotent Stem Cells , Blastocyst Inner Cell Mass , Cytokines , Mice , Rats , Culture TechniquesABSTRACT
Objective: to evaluate the effects of pre- and/or postnatal exposure to particulate air pollution on fertilization, embryo development, and cell lineage segregation in preimplantation blastocysts during late-life reproductive period, using the in vitro fertilization (IVF) mouse model. Materials and methods: five-month-old mice underwent superovulation and were pre- and/or postnatally exposed to filtered air (FA-FA), filtered-ambient air (FA-AA), or ambient air (AA-AA) in exposure chambers, 24 hours per day, during six months. Results: ovarian response was not affected by the exposure protocol. A multivariate effect for pre- and/or postnatal exposure to ambient air on blastocyst differential staining, but not on IVF and embryo development, was found. Cell counts in inner cell mass (ICM) and ICM/trophectoderm ratios in blastocysts produced in FA-FA protocol were significantly higher than in blastocysts produced in FA-AA and AA-AA protocols. Conclusions: our study suggests that the exposure to particulate air pollution of a large urban center does not affect ovarian function but may negatively affect the female reproductive health in the late-life period by disrupting the lineage specification at the blastocyst stage.
Objetivo: avaliar os efeitos da exposição pré e/ou pós-natal ao ar ambiente no final da vida reprodutiva sobre a fertilização, desenvolvimento embrionário e segregação das linhagens celulares em blastocistos pré-implantacionais, utilizando o modelo de fertilização in vitro (FIV) de camundongo. Material e métodos: fêmeas de camundongo com idade de cinco meses tiveram a ovulação estimulada e, no período pré e/ou pós-natal, foram expostas ao arfiltrado (AF-AF), ar filtrado-ar ambiente (AF-AA) ou ar ambiente-ar ambiente (AA-AA) em câmaras de exposição, 24 horas por dia durante seis meses. Resultados: a resposta ovariana não foi afetada pelo protocolo de exposição. Um efeito multivariável para a exposição pré e/ou pós-natal ao material particulado fino ambiente sobre coloração diferencial dos blastocistos, mas não sobre a FIV e o desenvolvimento embrionário, foi observado. A contagem celular na massa celular interna (MCI) e a razão MCI/trofoectoderma dos blastocistos produzidos no protocolo AF-AF foram significativamente maioresdo que aquelas em blastocistos produzidos nos protocolos FA-AA e AA-AA. Conclusões: nosso estudo sugere que a exposição à poluição ambiental particulada de um grande centro urbano não altera a função ovariana, mas pode afetar negativamente a saúde reprodutiva feminina no período final do menacme, em razão da alteração da especificação das linhagens celulares do embrião no estágio de blastocisto.