Oocyte cryopreservation and in vitro culture affect calcium signalling during human fertilization

• Published in: Human reproduction (Oxford) 2014-01, Vol.29 (1), p.29-40
• Main Authors: Nikiforaki, D., Vanden Meerschaut, F., Qian, C., De Croo, I., Lu, Y., Deroo, T., Van den Abbeel, E., Heindryckx, B., De Sutter, P.
• Format: Article
• Language: English
• Subjects: Adult; Calcium Signaling - physiology; Cryopreservation - methods; Female; Fertilization - physiology; Freezing; Fura-2; Humans; In Vitro Oocyte Maturation Techniques; Male; Oocytes - physiology; Sperm Injections, Intracytoplasmic; Vitrification
• ISSN: 0268-1161; 1460-2350
• DOI: 10.1093/humrep/det404

STUDY QUESTION What are the precise patterns of calcium oscillations during the fertilization of human oocytes matured either in vivo or in vitro or aged in vitro and what is the effect of cryopreservation? SUMMARY ANSWER Human oocytes matured in vivo exhibit a specific pattern of calcium oscillations, which is affected by in vitro maturation, in vitro ageing and cryopreservation. WHAT IS KNOWN ALREADY Oscillations in cytoplasmic calcium concentration are crucial for oocyte activation and further embryonic development. While several studies have described in detail the calcium oscillation pattern during fertilization in animal models, studies with human oocytes are scarce. STUDY DESIGN, SIZE, DURATION This was a laboratory-based study using human MII oocytes matured in vivo or in vitro either fresh or after cryopreservation with slow freezing or vitrification. Altogether, 205 human oocytes were included in the analysis. PARTICIPANTS/MATERIALS, SETTING, METHODS In vivo and in vitro matured human oocytes were used for this research either fresh or following vitrification/warming (V/W) and slow freezing/thawing (F/T). Human oocytes were obtained following written informed consent from patients undergoing ovarian hyperstimulation. For the calcium pattern analysis, oocytes were loaded with the ratiometric calcium indicator fluorescent dye Fura-2. Following ICSI using sperm from a single donor, intracellular calcium was measured for 16 h at 37°C under 6% CO2. The calcium oscillation parameters were calculated for all intact oocytes that showed calcium oscillations and were analyzed using the Mann–Whitney U-test. MAIN RESULTS AND THE ROLE OF CHANCE Human in vivo MII oocytes display a specific pattern of calcium oscillations following ICSI. This pattern is significantly affected by in vitro ageing, with the calcium oscillations occurring over a longer period of time and with a lower frequency, shorter duration and higher amplitude (P < 0.05). In vitro matured oocytes from the GV and MI stage exhibit a different pattern of calcium oscillations with calcium transients being of lower frequency and shorter duration compared with in vivo matured MII. In MI oocytes that reached the MII stage within 3 h the calcium oscillations additionally appear over a longer period of time (P < 0.05). In vivo MII oocytes show a different calcium oscillation pattern following V/W with calcium oscillations occurring over a longer period of time, with a higher amplitude and a lower frequ