Development of Primary Monolayer Cell Model and Organotypic Model of Uterine Leiomyoma

Abstract

Cellular technologies are one of the most promising areas of biomedicine, which is based on the isolation of cells of various types, followed by their cultivation and use, or the use of their metabolic products, for medical purposes. Today, a significant part of biomedical research is carried out in vitro. On the other hand, organotypic culture can be used as a powerful model system and can complement cell culture and in vivo studies in different biomedical applications. Uterine leiomyoma (UL) is a very common benign tumor and often leads to many reproductive complications. Herein we describe a fast and reliable method of isolation and UL primary cells culturing along with the development of a UL organotypic model. We propose the usage of UL primary cells in experimental work at a first passage to prevent loss of driver mutations in MED12 and HMGA2 genes. New optimized conditions for the growth and maintenance of 2D and 3D models of uterine leiomyoma in vitro are suggested.

Keywords: HMGA2; MED12; cell model; organotypic model; uterine leiomyoma.

Figure 1

Representative scheme of UL cellular and organotypic model development.

Figure 2

Representative microphotographs of the cellular and organotypic UL models: (a) primary UL cells at p0 (magnification × 100; scale bar is 100 µm); (b) typical appearance of leiomyoma fragments.

Figure 3

Detection of heterozygous mutation c.131G >A in MED12 gene in the same primary UL cell line at different passages: (a) p1; (b) p2; (c) p3; (d) control–healthy myometrium.

Figure 4

Measurement of HMGA2 gene expression (fold change relative to reference β-actin gene) in two independent primary UL cell lines at different passages.

Figure 5

Histological characterization of the organotypic model. Hematoxyline-eosine stained cryosections of UL fragments after 7 (a), 14 (b), and 21 (c) days of culture (magnification × 100; scale bar is 200 µm). Arrows mark extracellular matrix.