Objective: With this study we wanted to understand, why stimulation with recombinant FSH/LH could be advantageous for patients over 35 years during an in-vitro-fertilisation treatment over recombinant FSH stimulation alone. Design Prospective case-control study: Setting Recruitment of patients at the VivaNeo Fertility Centre Wiesbaden, Germany. Measurements of the parameters at University Medical Centre of the Johannes Gutenberg University Mainz, Gynaecological Endocrinology and Reproductive Medicine, Germany. April 2012 until April 2013 Patients: After obtaining a declaration of consent 78 patients between 35 and 40 years, without endocrine disorders as low LH, aberrant menstrual cycle or low ovarian reserve, planed for intracytoplasmic sperm injection (ICSI) treatment with antagonist protocol were selected. They were divided in two stimulation groups of which 39 were stimulated with recombinant follicle-stimulating hormone plus recombinant luteinizing hormone (FSH/LH, Pergoveris®) and the other with recombinant FSH (Gonal®). Interventions: During oocyte pick-up granulosa-cells were obtained from the cumulus-cell complex and from follicular fluid. They were seeded separately and cultured for 48 hours. At average 16, 24 and 40 hours media change took place and secretion of progesterone, inhibin-A, vascular endothelial growth factor (VEGF), endothelin-1 were measured several times. After 48 hours culture was terminated and cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) were detected out of the cell-lysate. Main Outcome Measure: A granulosa cell culture could successfully be established. Secretion rate of progesterone, VEGF and cGMP in luteinized human granulosa cells is increased in after stimulation with recombinant FSH/LH in comparison to FSH alone. Results: Vitality and concentration of luteinized human granulosa cells were stable during 48 hours of culture. Secretion rates of progesterone, VEGF and endothelin-1 showed higher tendency after stimulation with recombinant FSH/LH. For inhibin-A an opposite secretion rate was observed. For cAMP no differences were found, but cGMP content favoured the combined stimulation. Conclusion: Beside the fact we did not find clear significance, we think that stimulation with recombinant FSH/LH can improve different functions of human granulosa cells during in-vitro culture. Perhaps this can lead to an explanation, why this stimulation could be beneficial for the outcome of in-vitro-fertilisation.
