IGF-2 gene expression in liver and muscle tissue of fattening lambs under the influence of organic zinc-methionine supplement and calcium salt of flaxseed oil supplement in the diet

Introduction: Insulin-like growth factors (IGFs) are known as regulators of cell growth and development. This protein plays an essential role in growth and development before birth. Studies suggest that insulin-like growth factor 2 (IGF-2) promotes the growth and division (proliferation) of cells in many different tissues. Zinc is one of the most limiting trace mineral elements and is required for body growth, structure, hormonal and enzyme activity, nutrient metabolism, cell division, and the immune system. It has been found that zinc deficiency is associated with reduced food intake and reduced growth. Zinc is one of the important factors in the regulation of IGF family gene expression in many tissues. Also, zinc is effective in desaturating linoleic acid. Zinc can prevent lipid peroxidation. In general, there is a significant relationship between fat metabolism and zinc. On the other hand, flaxseed oil, also known as flax oil or linseed oil, is made from flax seeds that have been ground and pressed to release their natural oil. Flaxseed oil contains both omega-3 and omega-6 fatty acids, which are needed for health. Flaxseed oil contains the essential fatty acid alpha-linolenic acid (ALA), which the body converts into eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are the omega-3 fatty acids found in fish oil. Omega-3 fatty acids are essential to health and have been associated with benefits like reduced inflammation, improved heart health, and the protection of the brain against aging. Some researchers showed that the essential fatty acid alpha-linolenic acid (omega-3) is related to the functions of the IGF-2 in the body. Because of the adverse effects of unprotected fatty acids (FAs) on the rumen environment through altering the direct pathway of rumen biohydrogenation and altering the FAs profile in the fore-stomach, methods should be used to protect polyunsaturated fatty acids in the rumen. Protection methods include either encapsulating unsaturated FAs inside a microbial-resistant shell (such as lipid encapsulation) or modifying the FAs' structure by blocking the carboxyl group (such as calcium salts or fatty amides) to resist microbial enzymes. Because of the commercial availability of calcium salt, researchers have used this form of protection in assessing the flow of FAs in the duodenum. This research was conducted to investigate the effect of adding zinc-methionine organic supplement to diets with and without calcium salt of flaxseed oil on the IGF-2 gene expression in liver and muscle tissue of fattening lambs.
Materials and methods: In this research, 44 Arab male lambs were used with a 2×2 factorial experiment in a completely randomized design with four treatments and 11 replications. The four experimental diets were: 1) Basal diet without Ca-salt of flaxseed oil supplement and zinc-methionine supplement (CON), 2) Basal diet without Ca-salt of flaxseed oil supplement containing 0.083% zinc-methionine supplement (equivalent to 120 mg of zinc) per kg of dry matter, ZM), 3) Basal diet containing 3% Ca-salt of flaxseed oil supplement without zinc-methionine supplement (CFO), and 4) Basal diet containing 3% Ca-salt of flaxseed oil supplement plus 0.083% zinc-methionine supplement (CFO+ ZM). After the fattening period, three heads of lambs from each treatment were slaughtered. The liver and muscle tissues were taken to the laboratory in liquid nitrogen. After extracting RNA and measuring its quality, cDNA synthesis was performed. Finally, the expression of the IGF-2 gene was evaluated using the real-time polymerase chain reaction (RT-PCR) method. In this method, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was used as a housekeeping gene to normalize the gene expression data in the quantitative RT-PCR.
Results and discussion: The presence of only one peak in the melting curves of the IGF-2 and GAPDH genes in the RT-PCR reaction confirmed the production of a specific product in this reaction. The observation of a single band in the range of 203 bp for the IGF-2 gene and the range of 78 pairs of nucleotides for the GAPDH gene on gel electrophoresis indicated the correctness of the test and amplification of the desired fragment by the PCR. The results of this study showed that adding zinc supplementation significantly increased the IGF-2 gene expression in the liver (P<0.01). In comparison, this effect was not significant in the muscle. The effect of calcium salt of flaxseed oil supplementation on the IGF-2 gene expression was significant in both tissues. The interaction effects of zinc-methionine supplementation and calcium salt of flaxseed oil supplementation were non-significant. Adding organic zinc-methionine supplement to diets containing calcium salt of flaxseed oil supplementation increased the IGF-2 gene expression. IGF-2 is known as a key growth factor in metabolic processes and tissue growth. This gene is known as an important factor in tissue growth and development, especially in liver and muscle tissue, and plays a vital role in regulating fat and protein metabolism. In recent years, attention has increased to the effect of nutritional supplements in improving the performance and health of livestock, as well as improving gene expression in various tissues. One of the supplements of interest in this field is zinc-methionine, which is known as an organic-mineral compound. Several experiments have been conducted in this field, which have shown that feeding diets with sufficient zinc increases the expression of IGF-2 compared to diets with zinc deficiency. Incorporating these supplements into the diet can directly affect protein and amino acid metabolism, thereby increasing the expression of growth-related genes.
Conclusions: Adding a zinc-methionine organic supplement to diets containing calcium salt of flaxseed oil increased the IGF-2 gene expression. This will likely increase growth and ultimately production.