Establishment of a new method for isolation and culture of equine endometrial epithelial and stromal cells

A new method of in vitro culture of equine endometrial epithelial and stromal cells is described. After dissection, the endometrial tissue was dissociated in collagenase type II, sieved through nylon gauze and the different cell types were separated by centrifugation and differential adhesion. Cells were seeded at a density of 1x10(4) viable cells per cm(2) on several surfaces and cultured at 37 degrees C in humidified atmosphere. For both cell types the same serum-supplemented culture medium was utilized whereby confluent monolayer evolved after 10 to 14 days of incubation. Confluent cultures were dissociated using 0.25% trypsin-EDTA (stromal cells) or alfazyme (epithelial cells) and passaged up to 19 times. Further on, a successful cryoconservation-protocol could be established. With it, we created the precondition to establish a co-culture system for equine endometrial epithelial and stromal cells with the long-term objective to study growth characteristics, effects of steroid hormones and mutual influences between the cell populations, as well as potential pathogenetic factors and etiological influences on equine endometrosis.