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Whole-ovary decellularization generates an effective 3D bioscaffold for ovarian bioengineering

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A Correction to this article was published on 28 July 2023

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Abstract

Purpose

To develop a new protocol for whole-ovary decellularization for the production of a 3D bioscaffold suitable for in vitro/ex vivo studies and for the reconstruction of a bioengineered ovary.

Methods

Porcine ovaries were subjected to the decellularization process (DECELL; n = 20) that involved a freeze-thaw cycle, followed by sequential incubations in 0.5% SDS for 3 h, 1% Triton X-100 for 9 h, and 2% deoxycholate for 12 h. Untreated ovaries were used as a control (CTR; n = 6). Both groups were analyzed to evaluate cell and DNA removal as well as ECM preservation. DECELL bioscaffolds were assessed for cytotoxicity and cell homing ability.

Results

DECELL ovaries maintained shape and homogeneity without any deformation, while their color turned from red to white. Histological staining and DNA quantification confirmed a decrease of 98.11% in DNA content, compared with the native tissue (CTR). Histochemical assessments demonstrated the preservation of intact ECM microarchitecture after the decellularization process. This was also confirmed by quantitative analysis of collagen, elastin, and GAG contents. DECELL bioscaffold showed no cytotoxic effects in co-culture and, when re-seeded with homologous fibroblasts, encouraged a rapid cell adhesion and migration, with repopulating cells increasing in number and aggregating in cluster-like structures, consistent with its ability to sustain cell adherence, proliferation, and differentiation.

Conclusion

The protocol described allows for the generation of a 3D bioscaffold that may constitute a suitable model for ex vivo culture of ovarian cells and follicles, as well as a promising tool for the reconstruction of a bioengineered ovary.

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Acknowledgments

The Laboratory of Biomedical Embryology is a member of the COST Action CA16119 In vitro 3-D total cell guidance and fitness (CellFit). SEM analysis was carried out at NOLIMITS, an advanced imaging facility established by the Università degli Studi di Milano.

Funding

This work was funded by Carraresi Foundation and PSR2017.

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Authors and Affiliations

  1. Laboratory of Biomedical Embryology, Department of Health, Animal Science and Food Safety and Center for Stem Cell Research, Università degli Studi di Milano, via Celoria 10, 20133, Milan, Italy

    Georgia Pennarossa, Matteo Ghiringhelli & Tiziana A. L. Brevini

  2. Department of Agricultural and Environmental Sciences - Production, Landscape, Agroenergy and Center for Stem Cell Research, Università degli Studi di Milano, via Celoria 2, 20133, Milan, Italy

    Fulvio Gandolfi

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  1. Georgia Pennarossa
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  2. Matteo Ghiringhelli
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  4. Tiziana A. L. Brevini
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Correspondence to Tiziana A. L. Brevini.

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Pennarossa, G., Ghiringhelli, M., Gandolfi, F. et al. Whole-ovary decellularization generates an effective 3D bioscaffold for ovarian bioengineering. J Assist Reprod Genet 37, 1329–1339 (2020). https://doi.org/10.1007/s10815-020-01784-9

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