In vitro development of bovine embryos cultured with stem cell factor or insulin-like growth factor-I following IVF with semen of two bulls having different field fertility

Abstract

The usefulness of IVF as a potential tool to evaluate the field fertility of bulls is equivocal and growth factor addition to culture media research is needed to delineate components needed for providing defined environments for embryos. The overall aim was to evaluate the in vitro development of embryos derived using a serum supplemented and serum-free production systems and semen from two bulls of different field fertility. The study was conducted to determine the combinatorial effect of stem cell factor (SCF) and/or insulin-like growth factor-I (IGF-I) in culture on subsequent embryo development in cattle. Oocytes were aspirated separately from ≥3 to <3 mm follicles to test different follicle size populations and were matured in TCM-199 supplemented with LH, FSH, estradiol and BSA (Fraction V). Matured oocytes were fertilized in BSA supplemented synthetic oviductal fluid (SOF)-IVF medium. Presumptive zygotes were cultured for 8 d (in humidified 5% CO₂ at 38.5 °C) in BSA supplemented SOF-in vitro culture (IVC) medium. SOF-IVC medium was supplemented with fetal bovine serum (4%), IGF-I (100 ng/mL), SCF (50 ng/mL) or IGF-I (100 ng/mL) + SCF (50 ng/mL). The development competence of embryos did not differ between the bulls and among the culture environments. Nevertheless, there was an effect of follicle size on cleavage rate (P < 0.05) and a greater cleavage rate resulted from oocytes aspirated from ≥3 mm follicles (71.0 ± 1.5%) compared to those collected from <3 mm follicles (64.8 ± 1.6%). The overall cleavage rate (%); blastocyst formation (%); and expanded/hatched blastocyst formation (%) were 68.2 ± 1.5 and 67.7 ± 1.7; 29.4 ± 1.4 and 28.6 ± 1.5; and 18.6 ± 1.2 and 18.5 ± 1.1, respectively, for the bull of above and below average field fertility. The results indicate that follicle size for oocyte aspiration is effective for determining IVC success and that IVF may not discriminate among bulls of different field fertility.

Introduction

The success of AI depends on many factors. The estimated relative conception rate (ERCR) is a measure of the fertility of an individual sire (Clay and McDaniel, 2001). The ERCR predicted fertility value is a 70-d non-return rate of an AI service relative to the service sires of herd mates. Many laboratory procedures have been used to test different aspects of semen quality to predict field fertility. Yet, the usefulness of these techniques remains inconclusive (Rodriguez-Martinez, 2003). The ability of sperm to fertilize oocytes following in vitro fertilization (IVF) differs among bulls (Lonergan, 1994, Shi et al., 1990) and sire used in IVF has a profound effect on in vitro embryonic development (Ward et al., 2003).

The in vitro culture environment has a great influence on the development and quality of embryos. Though in vitro embryo production using serum supplemented media is widely accepted, serum contains many undefined factors that may stimulate or inhibit embryo development (Yamashita et al., 1999). Development may vary with serum from different sources (Batt and Miller, 1988), and serum has a biphasic effect on embryo development in cattle. Serum can inhibit the first cleavage division, but it can stimulate blastocyst development (Van Langendonckt et al., 1997). Experiments have been attempted to investigate the effects of growth factors on embryonic development in cattle. Growth factors improved embryo development, but they have not yielded results comparable to fetal bovine serum (FBS). However, previous research reported that the use of growth factors such as epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I) improved the rate of blastocyst formation of embryos. Kurzawa et al. (2004) demonstrated that EGF and IGF-I, at a concentration of 100 ng/mL, had a positive effect on mouse embryos. Culture medium supplemented with IGF-I was beneficial for preimplantation embryo development because it increased cell proliferation in human, rabbit and mice blastocysts (Makarevich and Markkula, 2002). With embryo culture in cattle, EGF supplementation was less effective that IGF-I or stem cell factor (SCF, Dhali et al., 2007b). Also, it is recognized that oocyte quality and the development competence following IVF is determined by the characteristics of follicles and follicular size (Qian et al., 2001).

The overall hypotheses were that growth factor supplementation to culture was equal to an undefined medium containing FBS for embryo development and in the absence of serum supplement field fertility differences could be evaluated with in vitro embryo development. The objective was to investigate the in vitro development of embryos in cattle after IVF with the semen of bulls of above and below average field fertility. For the precise evaluation of the sire effects on embryo development, a total serum-free system was used for in vitro embryo production to exclude the possible undesirable influence of serum on embryo development.