Cytoarchitecture modifications of the human uterine endocervical mucosa in long-term three-dimensional organotypic culture

Abstract

We assayed the effects of phenol red ( pr), estrogen (Es), and progesterone (Pg) in three-dimensional organotypic cultures of human uterine endocervix. Small intact fragments deposited on sponges submerged in DMEM with 10% fetal bovine serum were cultured in three different conditions: with pr (DMEM pr+), without pr (DMEM pr−), and without pr but with the addition of physiological concentrations of Es and Pg [DMEM pr−(Es+Pg)]. Cell viability and cellular responses were assayed after 4, 10, and 21 days using immunohistochemistry for the expression and distribution of the following markers: mucins and mucopolysaccharides (PAS staining), pan-cytokeratins (AE1/AE3), CK19, p63, Ki-67, vimentin, estrogen receptor- α (ER- α), and progesterone receptor (PR). The fragments cultivated in DMEM pr+ showed a cuboidal, poorly differentiated epithelial phenotype and signs of stroma degeneration. In DMEM pr−, both tissue architecture and cellular heterogeneity were much better preserved: epithelial cells showed a more columnar shape, and stroma was very well conserved, maintaining cell density. The addition of Es and Pg further improved the histology and physiology of the fragments: in DMEM pr−(Es+Pg), epithelial cells retained a columnar shape, secreted mucins, and formed areas of squamous hyperplasia.