Establishment Of Three-dimensionally Culture Model Of Ovine Endometrial Cells And Impact Of 17β-estradiol On The Expression Of β-defensin Gene 1

Defensins are rich in basic amino acids cationic peptides. Previous researches had shown that 17β-estradiol are able to induce the expression of SBD-1, however, previous studys are confined to two-dimensional cells culture. Cells in vivo are encapsulated in a complex three-dimensional (3D) structure called extracellular matrix (ECM), which is crucial for cells to conduct complicated biochemical and physical signaling. It is a very serious piece of research of culturing ovine endometrial cells with three-dimensionally in vitro.The main purpose of this study was to explore the economical and simple culture system for growing ovine endometrial cells in three-dimensional culture.And on this basis ovine endometrial cells are treated with 17p-estradiol at dose from 10-6to10-10mol/L, respectively, and the total RNA are extracted from cells at 2 h to24 h for examination by real-time PCR. Under the two-dimensional culture, three methods, which are conventional tissue culture, trypsin digestion and trypsin digestion+tissue culture, are compared. The results show that the about three methods all can obtain ovine endometrial epithelial cells and stromal cells in vitro and these are used for seed cells of three-dimensional culture. The growth curves shape of cells obtained are "S". Ovine endometrial epithelial cells which have polygon and grew like pave road stone. They are stained positively by cytokeratin. Stromal cells are long spindle and grow in swirl. They are stained positively by vimentin and negatively by cytokeratin.Endometrium cells with three kinds of support in vitro to structure endometrium-like structures, which are matrigel,collagen solution,collagen solution:matrigel(1:1). Results showed that collagen solution are unstable, matrigel are expensive. Collagen Solution:matrigel(1:1) are an efficient way to establish three-dimensional cultured model of ovine endometrial cells.The results of real-time PCR showed that 17β-estradiol is able to induce and increase the expression of SBD-1 gene in a time-and concentration-dependent manner.