Obstetric and neonatal outcomes after frozen-thawed embryos transfer with laser-assisted hatching: a retrospective cohort study.

PURPOSE: To ascertain if assisted hatching (AH) increases the risk of placenta-associated diseases and perinatal outcomes after frozen-thawed cleavage-stage embryo transfer. METHODS: We retrospectively evaluated 924 women who conceived with frozen-thawed cleavage-stage embryos transfer with (n = 390) or without (n = 534) laser-AH between 2013 and 2015. Data were obtained from the database on in vitro fertilization (IVF) patients in Shanghai First Maternity and Infant Hospital. We assessed neonatal (preterm birth, low birthweight, fetal macrosomia, stillbirth) and obstetric (miscarriage, ectopic pregnancy, post-term pregnancy, gestational diabetes (GDM), preeclampsia, intrahepatic cholestasis (ICP), placenta previa, placental abruption, premature rupture of membranes) outcomes. RESULTS: In twins, the median birthweight was lower in the AH group than that in the control group, and the prevalence of low birthweight (< 2500 g) was significantly higher in the AH group; after adjusting for maternal age, body mass index, mode of fertilization, and parity, no significant difference was found. In twins, no significant difference was detected in the prevalence of stillbirth or preterm pregnancy. In singleton births, there was no significant difference in the prevalence of low birthweight, macrosomia, preterm pregnancy or post-term pregnancy between the two groups. In singletons and twins, there were no significant differences in the prevalence of miscarriage, ectopic pregnancy, preeclampsia, GDM, ICP, or placenta abruption between the two groups. CONCLUSIONS: AH is a relatively safe method and our study provides important information for using this method in carefully selected patients.

FACS selection of valuable mutant mouse round spermatids and strain rescue via round spermatid injection.

Round spermatid injection (ROSI) into mammalian oocytes can result in the development of viable embryos and offspring. One current limitation to this technique is the identification of suitable round spermatids. In the current paper, round spermatids were selected from testicular cells with phase contrast microscopy (PCM) and fluorescence-activated cell sorting (FACS), and ROSI was performed in two strains of mice. The rates of fertilization, embryonic development and offspring achieved were the same in all strains. Significantly, round spermatids selected by PCM and FACS were effectively used to rescue the infertile Pten-null mouse. The current results indicate that FACS selection of round spermatids can not only provide high-purity and viable round spermatids for use in ROSI, but also has no harmful effects on the developmental capacity of subsequently fertilized embryos. It was concluded that round spermatids selected by FACS are useful for mouse strain rederivation and rescue of infertile males; ROSI should be considered as a powerful addition to the armamentarium of assisted reproduction techniques applicable in the mouse.

Reprogramming of round spermatids by the germinal vesicle cytoplasm in mice.

The birthrate following round spermatid injection (ROSI) remains low in current and evidence suggests that factors in the germinal vesicle (GV) cytoplasm and certain substances in the GV such as the nucleolus might be responsible for genomic reprogramming and embryonic development. However, little is known whether the reprogramming factors in GV oocyte cytoplasm and/or nucleolus in GV are beneficial to the reprogramming of round spermatids and development of ROSI embryos. Here, round spermatids were treated with GV cytolysates and injected this round spermatid alone or co-injected with GV oocyte nucleolus into mature metaphase II oocytes. Subsequent embryonic development was assessed morphologically and by Oct4 expression in blastocysts. There was no significant difference between experimental groups at the zygote to four-cell development stages. Blastocysts derived from oocytes which were injected with cytolysate treated-round spermatid alone or co-injected with nucleoli injection yielded 63.6% and 70.3% high quality embryos, respectively; comparable to blastocysts derived by intracytoplasmic sperm injection (ICSI), but higher than these oocytes which were co-injected with lysis buffer-treated round spermatids and nucleoli or injected with the lysis buffer-treated round spermatids alone. Furthermore, the proportion of live offspring resulting from oocytes which were co-injected with cytolysate treated-round spermatids and nucleoli or injected with cytolysate treated-round spermatids alone was higher than those were injected with lysis buffer treated-round spermaids, but comparable with the ICSI group. Our results demonstrate that factors from the GV cytoplasm improve round spermatid reprogramming, and while injection of the extra nucleolus does not obviously improve reprogramming its potential contribution, although which cannot be definitively excluded. Thus, some reprogramming factors are evidently present in GV oocyte cytoplasm and could significantly facilitate ROSI technology, while the nucleolus in GV seems also having a potential to improve reprogramming of round spermatids.