Spatiotemporal remodelling of the composition and architecture of the human ovarian cortical extracellular matrix during in vitro culture

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Abstract

Study question: How does in vitro culture alter the human ovarian cortical extracellular matrix (ECM) network structure?

Summary answer: The ECM composition and architecture vary in the different layers of the ovarian cortex and are remodelled during in vitro culture.

What is known already: The ovarian ECM is the scaffold within which follicles and stromal cells are organised. Its composition and structural properties constantly evolve to accommodate follicle development and expansion. Tissue preparation for culture of primordial follicles within the native ECM involves mechanical loosening; this induces undefined modifications in the ECM network and alters cell-cell contact, leading to spontaneous follicle activation.

Study design, size, duration: Fresh ovarian cortical biopsies were obtained from 6 women aged 28–38 years (mean ± SD: 32.7 ± 4.1 years) at elective caesarean section. Biopsies were cut into fragments of ~4×1×1 mm and cultured for 0, 2, 4 or 6 days.

Participants/materials, setting, methods: Primordial follicle activation, stromal cell density, and ECM-related protein (collagen, elastin, fibronectin, laminin) positive area in the entire cortex were quantified at each time point using histological and immunohistological analysis. Collagen and elastin content, collagen fibre characteristics and follicle distribution within the tissue were further quantified within each layer of the human ovarian cortex, namely the outer cortex, the mid-cortex, and the cortex-medulla junction regions.

Main results and the role of chance: Primordial follicle activation occurred concomitantly with a loosening of the ovarian cortex during culture, characterised by an early decrease in stromal cell density from 3.6 ± 0.2 x106 to 2.8 ± 0.1 x106 cells/mm3 at D2 (p=0.033) and a dynamic remodelling of the ECM. Notably, collagen content gradually fell from 55.5 ± 1.7% positive area at day 0 (D0) to 42.3 ± 1.1% at D6 (p=0.001), while elastin increased from 1.1 ± 0.2% at D0 to 1.9 ± 0.1% at D6 (p=0.001). Fibronectin and laminin content remained stable. Moreover, collagen and elastin distribution were uneven throughout the cortex and during culture. Analysis at the sub-region level showed that collagen deposition was maximal in the outer cortex and the lowest in the mid-cortex (69.4 ± 1.2% vs 53.8 ± 0.8% positive area, p

Limitations, reasons for caution: Ovarian cortical biopsies were obtained from women undergoing caesarean sections. As such, the data obtained may not accurately reflect the ECM distribution and structure of non-pregnant women.

Wider implications of the findings: Clarifying the composition and architecture signature of the ovarian cortical ECM provides a foundation for further exploration of ovarian microenvironments. It is also critical for understanding of ECM-follicle interactions regulating follicle quiescence and awakening, leading to improvements in both in vitro activation (IVA) and in vitro growth (IVG) techniques.
Study funding/competing interest(s): MRC grant MR/R003246/1 and Wellcome Trust Collaborative Award in Science: 215625/Z/19/Z.

Original languageEnglish
Article numberdead008
Number of pages15
JournalHuman Reproduction
DOIs
Publication statusPublished - 31 Jan 2023

Keywords

  • fertility preservation
  • extracellular matrix
  • ovary
  • mechanobiology
  • tissue stiffness
  • primordial follicle activation

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