In vitro growth and steroidogenesis of dog follicles are influenced by the physical and hormonal microenvironment

The present study examined the influences of the physical and hormonal microenvironment on in vitro growth and steroidogenesis of dog follicles. Follicles were enzymatically isolated and individually encapsulated in 0.5% (w/v; n=17) or 1.5% (n=10) alginate and cultured with 0.5 IU/ml equine chorionic gonadotropin for 192 h. In a separate experiment, follicles were encapsulated in 0.5% alginate and cultured with 0 (n=22), 1 (n=23), 10 (n=20) or 100 (n=21) μg/ml FSH for 240 h. Follicle diameter and steroid production were assessed every 48 h in both studies. Follicles encapsulated in the 0.5% alginate grew faster (P<0.05) than those cultured in the 1.5% concentration. Oestradiol (E2) and progesterone (P4) increased consistently (P<0.05) over time, and follicles in the 1.5% alginate produced more (P<0.05) P4 than those in the 0.5% solution. Follicles cultured in the highest FSH concentration (100 μg/ml) increased 100% in size after 240 h compared with 50 to 70% in lower dosages. E2 concentration remained unchanged over time (P>0.05) across FSH dosages. However, P4 increased (P<0.05) as culture progressed and with increasing FSH concentration. Results demonstrate that dog follicles cultured in alginate retain structural integrity, grow in size and are hormonally active. Lower alginate and increasing FSH concentrations promote in vitro follicle growth. However, the absence of an E2 rise in follicles cultured in FSH alone suggests the need for LH supplementation to support theca cell differentiation and granulosa cell function.