Abstract

Abstract Study question Is reduction in oxygen tension from 5% to 2% after D3 of extended culture beneficial for blastocyst development in patients with previous failed culture? Summary answer Ultra-low oxygen tension in extended culture improves blastocyst development (overall blastulation rate OBR), quality (top blastulation rate TPR) and increases the pregnancy rate (PR). What is known already The practice ESHRE guidelines recommends human embryo culture at low oxygen concentrations (2-8%). Traditionally atmospheric conditions (20% O2) were used but better outcomes were observed at low monophasic 5% O2. Human embryo leaves the oviduct (5-7% O2) to cross the uterus-tubal junction around D3-4 and reaches the uterin ultra-low O2 tension (2%). This decreasing gradient coincides with the shift from pre-compaction oxidative phosphorylation in the oviduct, to post-compaction glycolysis in the uterus. Few studies with ultra-low 2% O2 incubation suggested that the best O2 level may be stage-dependent and its reduction after D3 may be a more physiological approach. Study design, size, duration Prospective observational study in a single private hospital during 2023. 71 patients experienced a failed cycle of extended culture (n = 29 cIVF, n = 37 ICSI and n = 5 Sibling oocytes) with a poor embryo quality and low OBR, under standard monophasic 5% O2 incubation. In the subsequent cycle for these 71 patients (n = 24 cIVF and n = 46 ICSI and n = 1 Sibling oocytes) the O2 level was reduced to 2% from D3 onwards. Participants/materials, setting, methods After a failed extended culture, considered as the control cycle, the patients were enrolled for the study. Blastocysts were rated by Gardner’s classification. In the control cycle embryos were cultured under monophasic 5% O2 until D6. In the subsequent cycle embryos were cultured under 5% O2 until D3 and in ultra-low 2% O2 from D3 to D6. Sage 1-Step™ media was used for both cycles. The statistical analyses were done by chi-square and fisher test. Main results and the role of chance 71 patients were included in the study regardless of age, indication or technique. Each patient was its own control. The mean age was not different between the 2 groups (36.5±4 vs 37.2±4) but the attempt rank was logically statistically higher in the study cycle (2.1±1.4 vs 3.4±1, p<0.001). Oocyte maturity (75% vs 76.4%) and fertilization rate (66.8 vs 70.3%) were similar between groups. The average embryo quality at D3 was poor in both groups. OBR and TBR (40.7% vs 52.4%, p=0.004 and 15.8% vs 37.5%, p<0.001) were statistically higher after culture in ultra-low O2 conditions. Extended culture failure with absence of usable blastocyst (57.7% vs 19.7%, p<0.001) was statistically lower in ultra-low incubation group and the pregnancy rate PR per cycle (3.6% vs 25%, p=0.004) was seven fold higher after ultra-low culture. Limitations, reasons for caution The small size of the sample needs to be expanded in order to confirm the encouraging findings in a larger population. The study will continue during 2024 and will compel the live birth rate and the cumulative live birth rate. Wider implications of the findings Providing embryos with the most physiologic culture system would improve IVF cost-effectiveness and a better infertility management. Embryo energetic adaptation to suboptimal conditions results in impaired viability and lower pregnancy rates. In our study extended culture of low-quality embryos under 2% O2 after D3 yielded better culture and pregnancy outcomes. Trial registration number NA

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