Effect of hormones and growth factors on in vitro development of sheep preantral follicles

The present investigation attempts to improve the frequency of in vitro maturation of oocytes by culturing small (150-250 mu m) and large (>250-400 mu m) preantral follicles (PFs) of sheep for 6 days in various combinations/sequences of thyroxin (T-4), FSH, LH, transforming growth factor alpha (TGF-a), epidermal growth factor (EGF) and heat-treated foetal calf serum (FCS). Bicarbonate-buffered tissue culture medium 199, supplemented with 50 mu g ml(-1) gentamicin sulphate, served as the control medium. In vitro development was initially assessed by the proportion of PFs exhibiting an increase in size, mean increase in diameter and antrum formation. Nuclear maturation to the metaphase II stage of the oocytes isolated from cultured PFs, after an additional 24-h in vitro maturation, indicated success. A total of 15% of oocytes from small PFs and 55% from large PFs, cultured in T-4+FSH, matured to metaphase II. Culture of PFs in other combinations/sequences of hormones and growth factors, including the control medium, supported a significantly lower proportion of oocytes maturing to metaphase II stage. It is concluded that 6-day in vitro culture of sheep PFs in thyroxin and FSH greatly improves the frequency of oocyte maturation to metaphase II stage. (C) 2006 Elsevier B. V. All rights reserved.