Study on the optimal time limit of frozen embryo resuscitation transfer and the effect of a long-term frozen embryo on pregnancy outcome

Abstract Background ：Embryo freezing is now being utilized more frequently in clinical practice, not just to assist patients with fertility preservation, but also to provide patients who are not candidates for fresh embryo transfer more time to prepare. However, there is still fewer studies on the best timing to resuscitate embryos after freezing and whether long-term embryo freezing has a deleterious impact on pregnancy outcomes. Therefore, the aim of this study is to explore whether the length of time embryo freezing has an impact on pregnancy outcomes and to further explore the optimal time for resuscitation transfer after embryo freezing. Methods ：This retrospective study was conducted at Sichuan Jinxin Xinan Women and Children's Hospital, focusing on Frozen-thawed Embryo Transfer (FET). Data were collected from January 2015 to December 2021, encompassing patients who underwent the procedure. The participants were divided into three groups based on the duration of embryo freezing: ≤1 year, 1 to 6 years, and ≥6 years. The association between embryo freezing time and pregnancy outcome after FET was analyzed using various statistical analyses, including one-way ANOVA, Kruskal-Wallis test, chi-square test, and generalized estimating equation model. The studies were adjusted for confounding factors, and odds ratios (OR) and adjusted odds ratios (aOR), along with 95% confidence intervals (CI), were calculated. The primary outcomes assessed were the clinical pregnancy rate (CPR) and live birth rate (LBR). Secondary outcomes examined included the biochemical pregnancy rate, multiple pregnancy rate, ectopic pregnancy rate, early miscarriage rate, late miscarriage rate, preterm birth rate, neonatal birth weight, weeks at birth, and newborn sex. Results： We ultimately included 47,006 cycles for analysis. Patients were grouped according to embryo frozen time: Group 1 (≤ 1 year) consisted of 40,461 cycles, Group 2 (1~6 years) included 6,337 cycles, and Group 3 (≥ 6 years) comprised 208 cycles. After controlling for confounding factors, compared with the control group, the remaining groups had a decreased likelihood of achieving biochemical pregnancy rate, clinical pregnancy rate, and live birth rate (OR＜1, aOR＜1, P ＜0.05). Additionally, an increased incidence of ectopic pregnancy increased (OR >1, aOR>1), with a significant increase noted after six years of freezing time [aOR=4.141, 95% CI (1.013-16.921), P =0.05]. A frozen time of more than one year was associated with an increased risk of early miscarriage and preterm birth (OR>1, aOR>1). No statistically significant differences were found in birth weight or sex between groups. However, the birth rate of male infants was higher than that of female infants across all groups. Conclusions : This retrospective study demonstrates that better pregnancy outcomes are associated with embryo freezing time resuscitation transfer within one year, whereas freezing for more than one year may lower clinical pregnancy and live birth rates while increasing the risk of ectopic pregnancy and preterm birth. Therefore, it is recommended to minimize embryo freezing time.