بیان ژ‌‌ن IGF-2 در بافت کبد و ماهیچه بره‌های پرواری تحت تاثیر مکمل آلی روی-متیونین و مکمل نمک کلسیمی روغن کتان در جیره

نوع مقاله : مقاله پژوهشی

نویسندگان

گروه علوم دامی، دانشکده علوم دامی و صنایع غذایی، دانشگاه علوم کشاورزی و منابع طبیعی خوزستان

چکیده

هدف از این پژوهش، بررسی همزمان اثر تغذیه‌ای مکمل آلی روی-متیونین و مکمل چربی غیراشباع (مکمل نمک کلسیمی روغن کتان) بر بیان ژن‌ IGF-2 در بافت کبد و ماهیچه بره‌های نر پرواری بود. در این مطالعه از 44 راس بره نر عربی با آزمایش فاکتوریل 2×2 در قالب طرح کاملاً تصادفی با چهار تیمار و 11 تکرار استفاده شد. چهار جیره‌ آزمایشی عبارت بودند از: 1) جیره پایه بدون مکمل کلسیمی روغن کتان و بدون مکمل روی-متیونین (گروه شاهد)، 2) جیره پایه بدون مکمل کلسیمی روغن کتان حاوی 08/0 درصد مکمل روی-متیونین (معادل 120 میلی‌گرم روی در کیلوگرم ماده خشک، 3) جیره پایه حاوی سه درصد مکمل کلسیمی روغن کتان بدون مکمل روی-متیونین، و 4) جیره پایه حاوی سه درصد مکمل کلسیمی روغن کتان بعلاوه 08/0 درصد مکمل روی-متیونین .پس از پایان دوره پروار، سه راس بره از هر تیمار کشتار شد و نمونه‌های بافت ماهیچه و کبد بلافاصله در ازت مایع ذخیره شده و به آزمایشگاه منتقل شدند. پس از استخراج RNA و اندازه‌گیری کیفیت آن، ساخت cDNA انجام شد. در نهایت، بیان ژن IGF-2 با استفاده از روش Real time PCR مورد ارزیابی قرار گرفت. نتایج حاصل از این پژوهش نشان داد که افزودن مکمل روی منجر به افزایش معنی‌دار بیان ژن IGF-2 در کبد شد (01/0>P)، در حالی که این تغییر در ماهیچه معنی­دار نبود. اثر مکمل نمک کلسیمی روغن کتان بر بیان ژن IGF-2 در کبد و ماهیچه معنی­دار شد. آثار متقابل مکمل روی-متیونین و مکمل نمک کلسیمی روغن کتان معنی‌دار نشد. افزودن مکمل آلی روی-متیونین به جیره‌های حاوی مکمل نمک کلسیمی روغن کتان سبب افزایش بیان ژن‌ IGF-2 می­شود. افزایش بیان ژن‌ IGF-2 در کبد و ماهیچه بره­ها احتمالاً منجر به افزایش رشد و در نهایت، افزایش تولید دام خواهد شد.

کلیدواژه‌ها

موضوعات


عنوان مقاله [English]

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

نویسندگان [English]

  • M. Nazari
  • Z. Alipoor
  • S. Rostami
  • G. Mohammadi Ahvazi
Department of Animal Science, Faculty of Animal Science and Food Technology, Agricultural Science and Natural Resources University of Khuzestan, Mollasani, Iran
چکیده [English]

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.

کلیدواژه‌ها [English]

  • Fattening lamb
  • Gene expression
  • Zinc
  • Flaxseed oil
  • IGF-2 gene
Abareghi, F., Mohammadabadi, M., Ayatollahi Mehrjardi, A., Khezri, A., Shaban Jorjandy, D., Askari-Hesni, M., & Salemi, S. (2023). Examining of IGF2 gene expression in muscular and back fat tissues of Kermani sheep. Breeding and Improvement of Livestock, 3(2), 5-17. [In Persian]
Ahmad, S. S., Chun, H. J., Ahmad, K., Shaikh, S., Lim, J. H., Ali, S., & Choi, I. (2023). The roles of growth factors and hormones in the regulation of muscle satellite cells for cultured meat production. Journal of Animal Science and Technology, 65(1), 16-31. doi: 10.5187/jast.2022.e114
Beletskiy, A., Chesnokova, E., & Bal, N. (2021). Insulin-like growth factor 2 as a possible neuroprotective agent and memory enhancer—its comparative expression, processing and signaling in mammalian CNS. International Journal of Molecular Sciences, 22(4), 1849. doi: 10.3390/ijms22041849
Coelho, M. C., Malcata, F. X., & Silva, C. C. (2022). Lactic acid bacteria in raw-milk cheeses: From starter cultures to probiotic functions. Foods, 11(15), 2276. doi: 10.3390/foods11152276
Coyne, G. S., Kenny, D. A., & Waters, S. M. (2011). Effect of dietary n-3 polyunsaturated fatty acid supplementation on bovine uterine endometrial and hepatic gene expression of the insulin-like growth factor system. Theriogenology, 75(3), 500-512. doi: 10.1016/j.theriogenology.2010.09.018
De Souza, J., & Lock, A. L. (2018). Long-term palmitic acid supplementation interacts with parity in lactating dairy cows: Production responses, nutrient digestibility, and energy partitioning. Journal of Dairy Science, 101(4), 3044-3056. doi: 10.3168/jds.2017-13946
Deng, K., Li, X., Liu, Z., Su, Y., Sun, X., Wei, W., & Wang, F. (2024). IGF2BP2 regulates the proliferation and migration of endometrial stromal cells through the PI3K/AKT/mTOR signaling pathway in Hu sheep. Journal of Animal Science, 102, skae129. doi: 10.1093/jas/skae129
Gardner, S., Alzhanov, D., Knollman, P., Kuninger, D., & Rotwein, P. (2011). TGF-β inhibits muscle differentiation by blocking autocrine signaling pathways initiated by IGF-II. Molecular Endocrinology, 25(1), 128-137.‏ doi: 10.1210/me.2010-0292
Hao, K. L., Zhai, Q. C., Gu, Y., Chen, Y. Q., Wang, Y. N., Liu, R., & Hu, S. J. (2023). Disturbance of suprachiasmatic nucleus function improves cardiac repair after myocardial infarction by IGF2-mediated macrophage transition. Acta Pharmacologica Sinica, 44(8), 1612-1624.‏ doi: 10.1038/s41401-023-01059-w
Jafarpour, N., Khorvash, M., Rahmani, H.R., Pezeshki, A., & Hosseini Ghaffari, M. (2015). Dose–responses of zinc–methionine supplements on growth, blood metabolites and gastrointestinal development in sheep. Journal of Animal Physiology and Animal Nutrition, 99, 668-675. doi: 10.1111/jpn.12286
Kent, L. N., Ohboshi, S., & Soares, M. J. (2012). Akt1 and insulin-like growth factor 2 (Igf2) regulate placentation and fetal/postnatal development. The International journal of Developmental Biology, 56(4), 255. doi: 10.1387/ijdb.113407lk
Mahmoudi, T., Nouri, S., Zarei, F., Najafabadi, Z. N., Sanei, M., Sayedsalehi, S., & Zali, M. R. (2023). Insulin-like growth factor binding protein 3 promoter variant (rs2854744) is associated with nonalcoholic fatty liver disease. Archives of Endocrinology and Metabolism, 68, e230017. doi: 10.20945/2359-4292-2023-0017
Markljung, E., Jiang, L., Jaffe, J. D., Mikkelsen, T. S., Wallerman, O., Larhammar, M., & Andersson, L. (2009). ZBED6, a novel transcription factor derived from a domesticated DNA transposon regulates IGF2 expression and muscle growth. PLoS Biology, 7(12), e1000256. doi: 10.1371/journal.pbio.1000256
McCarthy, M. M., Mann, S., Nydam, D. V., Overton, T. R., & McArt, J. A. A. (2015). Concentrations of nonesterified fatty acids and β-hydroxybutyrate in dairy cows are not well correlated during the transition period. Journal of Dairy Science, 98(9), 6284-6290. doi: 10.3168/jds.2015-9446
Ndandala, C. B., Zhou, Q., Li, Z., Guo, Y., Li, G., & Chen, H. (2024). Identification of Insulin-like Growth Factor (IGF) family genes in the Golden Pompano, Trachinotus ovatus: Molecular cloning, characterization and gene expression. International Journal of Molecular Sciences, 25(5), 2499. doi: 10.3390/ijms25052499‏
Nazari, M., Salari, S., & Ghorbani, M. R. (2017). Effects of zinc supplementation and betaine substitution to methionine on hepatic betaine - homocysteine methyltransferase and lipogenic genes expression in laying hens under heat stress. Agricultural Biotechnology Journal, 9(1), 95-110. [In Persian]
Nazari, M., Sallari, S., & Ghorbani, M. R. (2020). Effect of Zinc supplementation and Betaine substitution to methionine on performance and blood parameters of laying hens under heat stress. Veterinary Research & Biological Products, 33(1), 61-70. [In Persian]
Oh, Y. S., & Choi, C. B. (2004). Effects of zinc on lipogenesis of bovine intramuscular adipocytes. Asian-Australasian Journal of Animal Sciences, 17(10), 1378-1382. doi: 10.5713/ajas.2004.1378
Palmquist, D.  L., & Jenkins, T. C. (2017). A 100-Year Review: Fat feeding of dairy cows. Journal of Dairy Science, 100(12), 10061-10077. doi: 10.3168/jds.2017-12924
Pavkovych, S., Vovk, S., & Kruzhel, B. (2015). Protected lipids and fatty acids in cattle feed rations. Acta Scientiarum Polonorum. Zootechnica, 14(3), 3-4.
Pewan, S. B., Otto, J. R., Huerlimann, R., Budd, A. M., Mwangi, F. W., Edmunds, R. C., & Malau-Aduli, A. E. O. (2020). Genetics of omega-3 long-chain polyunsaturated fatty acid metabolism and meat eating quality in Tattykeel Australian White lambs. Genes, 11(5), 587. doi: 10.3390/genes11050587
Pfaffl, M. W., Horgan, G. W., & Dempfle, L. (2002). Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Research, 30(9), e36-e36. doi: 10.1093/nar/30.9.e36
Salabi, F., Boujarpoor, M., Fayazi, J., Salari, S., & Nazari, M. (2011). Effects of different levels of zinc on the performance and carcass characteristics of broiler reared under heat stress condition. Journal of Animal and Veterinary Advances, 10(10), 1332-1335. doi: 10.3923/javaa.2011.1332.1335
Salabi, F., Nazari, M., Chen, Q., Nimal, J., Tong, J., & Cao, W. (2014). Myostatin knockout using zinc-finger nucleases promotes proliferation of ovine primary satellite cells in vitro. Journal of Biotechnology, 192, 268-280. doi: 10.1016/j.jbiotec.2014.10.038
Sloup, V., Jankovská, I., Nechybová, S., Peřinková, P., & Langrová, I. (2017). Zinc in the animal organism: a review. Scientia Agriculturae Bohemica, 48(1), 13-21. doi: 10.1515/sab-2017-0003
Sobeková, A., Piešová, E., Maková, Z., Szabóová, R., Sopková, D., Andrejčáková, Z., & Faixová, Z. (2023). Duration of the flaxseed supplementation affects antioxidant defense mechanisms and the oxidative stress of fattening pigs. Veterinary Sciences, 10(9), 586. doi: 10.3390/vetsci10090586.
Spears, J. W. (1989). Zinc methionine for ruminants: relative bioavailability of zinc in lambs and effects of growth and performance of growing heifers. Journal of Animal Science, 67(3), 835-843.‏ doi: 10.2527/jas1989.673835x
Vierboom, M. M., Engle, T. E., & Kimberling, C. V. (2003). Effects of gestational status on apparent absorption and retention of copper and zinc in mature Angus cows and Suffolk ewes. Asian-Australasian Journal of Animal Sciences, 16(4), 515-518. doi: 10.5713/ajas.2003.515
Wang, X., Lin, L., Lan, B., Wang, Y., Du, L., Chen, X., & Wang, Y. (2020). IGF2R-initiated proton rechanneling dictates an anti-inflammatory property in macrophages. Science Advances, 6(48), eabb7389. doi: 10.1126/sciadv.abb7389
Wei, C., Wu, M., Wang, C., Liu, R., Zhao, H., Yang, L., Liu, J., Wang, Y., Zhang, S., Yuan, Z., Liu, Z., Hu, S., Chu, M., Wang, X., & Du, L. (2018). Long noncoding RNA Lnc-SEMT modulates IGF2 expression by sponging miR-125b to promote sheep muscle development and growth. Cellular Physiology and Biochemistry, 49(2), 447-462. doi: 10.1159/000492979
White, V., Jawerbaum, A., Mazzucco, M. B., Gauster, M., Desoye, G., & Hiden, U. (2018). IGF2 stimulates fetal growth in a sex-and organ-dependent manner. Pediatric Research, 83(1), 183-189. doi: 10.1038/pr.2017.221
Wilson, E. M., & Rotwein, P. (2006). Control of MyoD function during initiation of muscle differentiation by an autocrine signaling pathway activated by insulin-like growth factor-II. Journal of Biological Chemistry, 281, 29962-29971. doi: 10.1074/jbc.M605445200
Wynn, P. C., & Sheehy, P. A. (2012). Minor proteins, including growth factors. In Advanced Dairy Chemistry: Volume 1A: Proteins: Basic Aspects, 4th Edition (pp. 317-335). Boston, MA: Springer US. doi: 10.1007/978-1-4614-4714-6_11
Yang, H., Zhang, F., Sun, S., Li, H., Li, L., Xu, H., & Lyu, F. (2023). Brushite-coated Mg–Nd–Zn–Zr alloy promotes the osteogenesis of vertebral laminae through IGF2/PI3K/AKT signaling pathway. Biomaterials Advances, 152, 213505. doi: 10.1016/j.bioadv.2023.213505
Yi, T., Wang, T., Shi, Y., Peng, X., Tang, S., Zhong, L., & Li, Q. (2020). Long noncoding RNA 91H overexpression contributes to the growth and metastasis of HCC by epigenetically positively regulating IGF2 expression. Liver International, 40(2), 456-467. doi: 10.1111/liv.14300
Younis, S., Schönke, M., Massart, J., Hjortebjerg, R., Sundström, E., Gustafson, U., & Andersson, L. (2018). The ZBED6–IGF2 axis has a major effect on growth of skeletal muscle and internal organs in placental mammals. Proceedings of the National Academy of Sciences, 115(9), E2048-E2057. doi: 10.1073/pnas.1719278115.
Yu, Z. P., Le, G. W., & Shi, Y. H. (2005). Effect of zinc sulphate and zinc methionine on growth, plasma growth hormone concentration, growth hormone receptor and insulin-like growth factor-I gene expression in mice. Clinical and Experimental Pharmacology and Physiology, 32(4), 273-278. doi: 10.1111/j.1440-1681.2005.04183.x