Comparison of age-related differences in expression of antioxidant enzyme mRNA and activity in various tissues of pigs

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Abstract

Antioxidant enzymes (AOEs), glutathione peroxidase (GPx), superoxide dismutase(SOD) and catalase (CAT) play an important role in protecting tissues from reactive oxygen species (ROS) reactions. The objective of this study was to determine the developmental regulation of AOEs mRNA levels and activity in tissues of different growing phases pigs (Sus scrofa). Nine different tissues were collected from thirty Duroc × Landrace × Yorkshire male pigs with six animals in each age (1, 42, 84, 126 and 168 days) to assay for GPx, CAT and CuZnSOD mRNA expression and activities. Results showed that GPx, CAT, and CuZnSOD mRNA levels in liver increased (P < 0.05) at the first stage, and thereafter their levels began to decline (P < 0.05), and the maximal mRNA levels of these AOEs were seen at the age of 42, 84, and 126 days, respectively. In Muscle, GPx and CAT mRNA level increased from 1 to 84 days and 1 to 126 days, respectively, and thereafter their levels began to decline, whereas CuZnSOD mRNA level steadily increased (P < 0.05) following birth. Activity expression of AOEs in selected tissues was increased as pigs became older (P < 0.05) with the exception of CuZnSOD activity in muscle, but changes in AOEs mRNA levels between ages did not fully account for all changes in activity. GPx and CuZnSOD mRNA were most abundantly expressed in muscle, while CAT mRNA were most abundant in brain. AOEs may exert cell and tissue-specific roles in metabolic regulation beyond their mere antioxidant potential. In conclusion, expression of AOEs mRNA and activity exhibit different developmental profiles in various tissues of pigs, and the regulation of AOEs is not tightly coordinated in either tissue.

Introduction

Reactive oxygen species play different positive roles in vivo, being involved in energy production, phagocytosis, cell growth and intercellular signaling regulation. Reactive oxygen species may be also highly damaging, as they can attack biological macromolecules, namely, lipids, protein and DNA, induce oxidation and cause membrane damage, enzyme inactivation and DNA damage (Valko et al., 2004). When the level of ROS exceeds the antioxidant capacity of the cell, the intracellular redox homeostasis is altered and oxidative stress ensues (Halliwell, 1999). Biological antioxidants are natural molecules which can prevent the uncontrolled formation of free radicals and activated oxygen species, or inhibit their reactions with biological structures (Hayes and McLellan, 1999, Chaudiere and Ferrari-Iliou, 1999). Intracellular antioxidants include low molecular scavengers of oxidizing species, and enzymes which degrade superoxide and hydroperoxides. Intracellular AOEs are considered to play a major role to protect against ROS-mediated cell and tissue injury. The main AOEs include dismutases such as superoxide dismutases (SOD) and catalases (CAT), which do not consume cofactors, and peroxidases such as selenium-dependent glutathione peroxidases (GPx) in animals. On the other hand, newborn mammals experience a transition of leaving a hypoxic uterine for a relative hyperoxic environment (Rickett and Kelly, 1990). The drastic change of the environmental oxygen concentration, along with a rapid neonatal growth, poses a significant oxidative stress on these newborns and necessitates the development of antioxidant defense systems to cope with the expected increases in free radicals production (Frank and Sosenko, 1987). It has been widely believed that intracellular GPx, SOD and CAT are an important part of these systems (Mishra and Delivoria-Papadopoulos, 1988, Sohal et al., 1990). Numerous studies have revealed biochemical mechanisms and tissue-specific functions of intracellular antioxidant enzymes GPx, CAT and SOD. At present it has not been well studied whether antioxidant enzymes expression in different growing phases pigs is developmentally regulated.

Copper, zinc, manganese, selenium, iron and vitamin E are important and necessary nutrient compositions in the dietary of pigs. They are also the assistant factors which participate in the expression and antioxidant activities of AOEs. Their deficiency and superabundance in dietary of human and animals usually induced a line of diseases. Dietary selenium or vitamin E deficiency in young pigs produces mulberry heart disease, which is similar to the Keshan disease associated with selenium deficiency in children and pregnant women (Lei et al., 1997). Thus, pigs are an excellent model for studying the biochemical mechanism of aging and some disease. In addition, pigs are one of the most important species in animal agriculture worldwide. Abnormality in the antioxidant defense systems causes the pigs to be susceptible to stress such as transportation or exercise (Duthie et al., 1989), and to some extent affect the meat quality. Elucidating regulation of antioxidant enzymes gene expression in pigs will provide a basis for appropriate dietary antioxidant nutrients for efficient pork production that serves for human nutrition. Therefore, the objective of the experiment was to study the developmental regulation and tissue-specific patterns of antioxidant enzymes mRNA and activity in various tissues of different growing phases pigs in an effort to better understand the purpose of these responses during development.

Section snippets

Animals

This experiment was approved by the Institutional Animal Care and Use Committee at Zhejiang University and was conducted in accordance with the National Institutes of Health guidelines for the care and use of experimental animals. The experimental pigs (Sus scrofa) (n = 30, male, Duroc × Landrace × Yorkshire) were from the Swine Research and Teaching Farm at Zhejiang University. Six each of age-stage (1, 42, 84, 126 and 168 days) male pigs were used. The average body mass of experimental pigs were

Developmental pattern of antioxidant enzymes mRNA levels

The developmental patterns of antioxidant enzymes gene expression of liver and muscle tissues in pigs with age of 1, 42, 84, 126, 168 days was evaluated using semi-quantitative RT-PCR, which allowed the assessment of relative antioxidant enzymes mRNA levels in pigs at different growth stages. The post-natal developmental profiles of AOEs mRNA in pigs liver and muscle are shown in Fig. 1, Fig. 2. Relative abundances of antioxidant enzymes GPx, CuZnSOD and CAT mRNA in liver and muscle, normalized

Discussion

In order to cope with an excess of free radicals produced upon oxidative stress, mammals have developed sophisticated mechanisms. Maintenance of redox homeostasis involves many control systems which must be precisely coordinated. It has been widely believed that intracellular antioxidant enzymes, GPx, SOD and CAT are important parts of these systems and play an important role in protecting tissues from ROS reactions (Mishra and Delivoria-Papadopoulos, 1988, Sohal et al., 1990). Numerous studies

Acknowledgement

This work was supported by a grant from the National Basic Research Program of China, Grant No. 2004CB117506.

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