Gonadotrophin stimulation in IVF alters the immune cell profile in follicular fluid and the cytokine concentrations in follicular fluid and serum

Abstract

Study question: Are the immune cell profiles and the cytokine concentrations in follicular fluid (FF) and serum at the preovulatory stage different in conventional exogenous gonadotrophin stimulated IVF (c-IVF) compared with natural cycle IVF (NC-IVF)?

Summary answer: The cell counts of CD45+ leucocytes and T cell subpopulations and the cytokine concentrations in FF and serum are different in c-IVF compared to NC-IVF.

What is known already: FF-derived cells are heterogeneous. Immune cells are involved in intra-ovarian processes and cytokines are required for normal follicular development. Gonadotrophins stimulate the regulatory intrafollicular system and influence the local distribution of immune cells and the intrafollicular release of cytokines. Administration of exogenous gonadotrophins may have a significant effect on this local regulatory system, which then in turn could influence oocyte quality.

Study design, size, duration: The study included 105 patients, 69 undergoing c-IVF and 36 undergoing NC-IVF. c-IVF was performed by exogenous ovarian stimulation with hMG and GnRH antagonists.

Participants/materials, setting, methods: FF samples were collected from the first dominant follicle in c-IVF without pooling and from single leading preovulatory follicles in NC-IVF. Three different approaches were used to analyze FF samples: (i) microscopic investigation of CD45+ leucocytes, (ii) fluorescence-activated cell sorting to determine CD19+ B cells and CD3+ T cells including T cell subpopulations (CD4+, CD8+), and (iii) evaluation of tumour necrosis factor-alpha (TNF-α), interferon-gamma (INF-γ), interleukins (IL)-2, -6, -8, -10 and vascular endothelial growth factor (VEGF) levels in matched FF and serum samples using the Bio-Plex® platform.

Main results and the role of chance: FF obtained from c-IVF contained proportionally more CD45+ leucocytes (P = 0.0384), but fewer CD8+ cytotoxic T cells than FF from NC-IVF. CD3+ T lymphocytes were the most common type of lymphocytes, and the number thereof was comparable in the two study groups. In c-IVF, serum VEGF levels were higher (P = 0.007) than in NC-IVF while FF contained marginally decreased concentrations of IL-8 in c-IVF in comparison to NC-IVF. The cytokine concentration gradient between FF and serum in c-IVF was 10-fold for IL-8 and 8-fold for VEGF and thereby markedly lower than in NC-IVF, where the differences were 32-fold and 30-fold, respectively. Strong positive correlations were determined between FF- IL-10 and FF- VEGF in c-IVF (r = 0.85, P < 0.0001) and in NC-IVF (r = 0.81, P < 0.0001).

Large scale data: N/A.

Limitations, reasons for caution: The ovulation of NC-IVF follicles was induced by the exogenous administration of hCG, which means that the environment did not fully correspond to the physiological situation.

Wider implications of the findings: The differences in the immune profile and the cytokine concentrations in c-IVF and NC-IVF follicles support the hypothesis that conventional ovarian stimulation affects indirectly and heterogeneously the intrafollicular milieu, and thereby possibly affects the oocyte quality and the IVF outcome. However, further studies are needed to confirm our findings and to refine stimulation protocols in the context of optimizing the intrafollicular environment during oocyte maturation.

Study funding/competing interest(s): The study was supported by a research grant from IBSA Institut Biochimique SA and MSD Merck Sharp & Dohme GmbH. The authors are clinically involved in low dose mono-follicular stimulation and IVF-therapies, using gonadotrophins from all gonadotrophins distributors on the Swiss market, including Institut Biochimique SA and MSD Merck Sharp & Dohme GmbH.

Keywords: cytokines; in vitro fertilization; leucocytes; natural cycle; ovarian stimulation.