Caprine Endometrial Mesenchymal Stromal Stem Cell: Multilineage Potential, Characterization, and Growth Kinetics in Breeding and Anestrous Stages

Abstract

The endometrial layer of the uterus contains a population of cells with similar characteristics of mesenchymal stem cells (MSCs). In the present study, caprine endometrial mesenchymal stromal stem cells (En-MSCs) characters and differentiation potential to chondrogenic, osteogenic, and adipogenic cell lines as well as their growth kinetics in breeding and anestrous stages were evaluated. En-MSCs were enzymatically isolated from endometrial layer of the uterus of adult goats and were cultured and subcultured until passage 4. The growth kinetics and population doubling time (PDT) of caprine En-MSCs in breeding and anestrous stages were determined. En-MSCs in passage 4 were used for the karyotyping and differentiation into chondrocytes, osteocytes, and adipocytes. The PDT in anestrus phase was 40.6 h and in cyclic goats was 53 h. En-MSCs were fibroblast-like in all passages. The number of chromosomes was normal (2n = 60) with no chromosomal instability. Chondrogenic, osteogenic, and adipogenic differentiation of En-MSCs was confirmed by staining with Alcian blue, Alizarin red, and Oil Red O, respectively. Caprine En-MSCs demonstrated to be an alternative source of MSCs for cell therapy purposes in regenerative medicine.

Figure 1

Uteri of goats (a) in breeding stage with corpus luteum (black arrow) and (b) in anestrus stage. The locations of sampling of endometrial tissues are shown by white arrows on body and horn of the uterus.

Figure 2

Fibroblast-like morphology of caprine endometrial mesenchymal stromal stem cells in (a) anestrous and (b) breeding stages (magnification: ×200).

Figure 3

Comparison of growth curves of goat endometrial mesenchymal stromal stem cells after seeding of 2.2 × 10⁴ cells (a) in 12-well culture plates between anestrous and breeding stage and (b) between 12-well and 24-well culture plates in anestrus stage.

Figure 4

Karyotype of endometrial mesenchymal stromal stem cells in passage 4 of cyclic goats.

Figure 5

Differentiation of endometrial mesenchymal stromal stem cells (En-MSCs) into (a) chondrocytes stained with Alcian blue shows methachromatic appearance (magnification: ×40). (b) Control cells that remained colorless (magnification: ×100). (c) Alizarin red staining revealed calcification and osteogenic differentiation of En-MSCs in goat (magnification: ×100). (d) Oil Red O staining revealed lipid droplets and adipogenic differentiation of En-MSCs in goat (magnification: ×40).