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Capitalizing on transcriptome profiling to optimize and identify targets for promoting early murine folliculogenesis in vitro
journal contributionposted on 2021-07-13, 20:17 authored by Andrea Jones, Beatriz Peñalver Bernabé, Vasantha Padmanabhan, Jun Li, Ariella Shikanov
In vitro ovarian follicle culture is an active area of research towards providing fertility options for survivors of childhood cancer. Late-stage murine follicles (multilayer secondary and onwards) can be cultured successfully to maturity to obtain a meiotically competent oocyte for fertilization, but primordial and primary follicles usually die in culture because many key components of early follicle development are still unknown and difficult to mimic in vitro. To engineer a biomimetic three-dimensional culture system with high efficacy and reproducibility for the clinic, detailed mechanisms of early folliculogenesis must be uncovered. Previous studies have shown that primary murine follicles co-cultured in groups, in contrast to single follicles cultured in isolation, can reach preovulatory size and produce competent oocytes, but the factors accounting for the synergy of follicle co-culture are still unknown. To probe the underlying mechanisms of successful follicle co-culture, we conducted a time-course experiment for murine follicles encapsulated in 0.3% alginate hydrogels and compared between two conditions: groups of 5 (5X) versus groups of 10 (10X). For every 2 days during the course of 12 days, follicles were dissociated and somatic cells were isolated for microarray-based gene expression analysis (n = 380 follicles for 5X and n = 430 follicles for 10X). Gene activities in follicles co-cultured in larger groups (10X) had a distinct transcriptomic profile of key genes and pathways such as prolactin signaling and angiogenesis-related genes when compared to cells from follicles co-cultured in the smaller cohort (5X). To benchmark the results for follicles grown in culture, we compared our microarray data to data from murine follicles freshly isolated from the ovary at comparable stages of development previously published by Bernabé et al. Comparison of these datasets identified similarities and differences between folliculogenesis in the native microenvironment and the engineered in vitro system. A more detailed understanding of follicle growth in vitro will not only allow for better culture methods but also advance the field towards providing improved fertility options for survivors of childhood cancer.
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