In vitro maturation (IVM) of murine and human germinal vesicle (GV) stage oocytes using variations of a human fallopian tube epithelial feeder cell system

Abstract

OBJECTIVE: To assess the maturation rate of murine and human GV oocytes using variations of a human tubal feeder cell system. DESIGN: Murine and human GV oocytes were randomized to human tubal fluid (HTF) media alone or co-culture with primary (CP58) or immortalized (CP13IM) human tubal cells. Rates of maturation were compared between groups. MATERIALS AND METHODS: Tubal epithelial cells were isolated from salpingectomy specimens by digestion with DNase/Collagenase. One batch was immortalized (CP13IM). Mouse GV oocytes were isolated from 8 week-old mice 40 h after PMSG stimulation. Human GV oocytes (n=21) were obtained from the MGH IVF lab. Murine and human GV oocytes were cultured in HTF media alone or on feeder layers of CP58 and CP13IM cells. Maturation rates were assessed by standard morphological criteria and compared between groups. RESULTS: Maturation rates of GV oocytes under the various culture conditions are shown in table 1. Tabled 1Maturation stage (% of total) of GV oocytes with different culture conditionsMurine (n=210, 24 h)Human (n=21)Culture systemGVGVBDMIIMII (24 h)MII (48 h)HTF27.5021.2551.2512.537.5CP588.7513.7577.5062.575.0CP13 IM12.5017.5070.0060.060.0 Open table in a new tab For murine GV oocytes (n=210 in 3 independent experiments), the maturation rate to the MII stage was approximately 50% higher with CP58 cells and nearly 40% higher with CP13IM cells compared to media alone. For human GV oocytes, the rate of maturation to the MII stage was approximately 5-fold higher with both the CP58 and CP13IM feeder systems at 24 h and approximately 2-fold higher at 48 h. CONCLUSIONS: This study shows that co-culture with either primary or immortalized tubal cells improved IVM rates for murine and human GV oocytes, suggesting a promising new technique with future applications in clinical reproductive medicine or in basic science research, such as in the generation of new stem cell lines.