Effect of Essential and Nonessential Amino Acids in North Carolina State University (NCSU)-23 Medium on Development of Porcine In vitro Fertilized Embryos

Eu Gine Lee,Ok-Jae Koo,Ji-Hye Kim,Dilip P Bhandari,Sue Kim,Yeon Woo Jeong,Md Abul Hashem,Sung Keun Kang,Seon Mi Park,Sun Woo Park,Byeong Chun Lee,Mohammad Shamim Hossein

doi:10.5713/ajas.2007.693

Abstract

The present study was conducted to examine the effect of different levels of essential and nonessential amino acid in NCSU-23 medium on the in vitro-produced porcine embryo as it develops from the zygote to the blastocyst stage. Four experiments were performed, each with a completely randomized design involving 5 to 8 replications of treatments. In order to know the effect of nonessential amino acids in NCSU-23 medium, 0, 5, 10 and 20 μl/ml MEM were supplemented there to, (Exp. 1) and the medium was supplemented with same level of essential amino acids (Exp. 2). The combined effect of nonessential (0, 5, 10 and 20 μl/ml MEM) and essential amino acids (0, 5, 10 and 10 μl/ml MEM) in NCSU-23 medium (Exp. 3), first 72 h with non-essential amino acids (at 0, 5, 10 and 20 μl/ml MEM), and last 4 d with essential amino acids with the same level as NEAA (Exp. 4) were examined. The embryo development was monitored and the quality of blastocysts was evaluated by counting the number of total cells and determining the ratio of inner cell mass (ICM) to trophoectoderm (TE) cells. When Eagle's nonessential amino acids (MEM) added to NCSU-23 medium, it significantly increased the likelihood of development to the 2- to 4-cell stage and subsequent blastocyst development. Supplementation of different levels of essential amino acids in the NCSU-23 medium decreased cleavage rate, rate of morula and blastocyst development and the number of ICMs. In the case of the combined effect of essential and nonessential amino acids, better and significant results were found for blastocysts, hatching blastocysts and for ICM numbers which were also dose dependent. With respect to the biphasic effect of nonessential and essential amino acids, nonessential amino acids increased cleavage whereas essential amino acids increased the total cell number. Neither the nonessential nor the essential group of amino acids, on their own, affected blastocyst cell number or the differentiation of cells in the blastocyst. In conclusion, this study determined the role of nonessential and essential amino acids in the culture of the porcine embryo and showed that the embryo requires different levels of amino acids as it develops from the zygote to the blastocyst stage.

Highlights

The present study was conducted to examine the effect of different levels of essential and nonessential amino acid in NCSU-23 medium of the in vitro-produced porcine embryo as it develops from the zygote to the blastocyst
Previous studies have illustrated the stimulatory effects of the combination of glutamine and the nonessential amino acids in the development of mammalian embryos in vitro
The nonessential amino acids and glutamine were found to stimulate the development of the mouse embryo by decreasing the time of the first three cleavage divisions (Lane and Gardner, 1997)

Summary

INTRODUCTION

Amino acids serve a variety of physiological functions, including: the synthesis of proteins and nucleotides (Epstein and Smith, 1973; Alexiou and Leese, 1992; Katchadourian et al, 1994), nutrition and energy provision (Lane and Gardner 1997a; 1998; Gardner, 1998; Houghton et al, 2002), osmoregulation (Van Winkle and Campione, 1996; Dumoulin et al, 1997; Dawson et al, 1998), protection. It has been suggested that in vitro produced embryos should be exposed to amino acids as early as the oocyte stage, as this increases oocyte maternal mRNA levels and promotes preimplantation development (Watson et al, 2000). Much of our understanding of the way that amino acids affect mammalian embryo development and subsequent viability has come from studies on the hamster (Carney and Bavister, 1987; Bavister and Arlotto, 1990; Bavister and Mckiernan, 1993; Mckiernan et al, 1995), mouse (Mehta and Kiessling, 1990; Gardner and Lane, 1993; Lane and Gardner, 1994; Gardner and Lane, 1996; Lane and Gardner, 1997; Lane and Gardner, 1997a), and rat (Zhang and Armstrong, 1990). The present study was undertaken to examine the effect of essential and nonessential amino acids alone or in combination, as well as the biphasic effect in NCSU-23 medium on the development of porcine IVF embryos

MATERIALS AND METHODS

RESULTS

Findings

DISCUSSION