Effects of Acute Heat Stress Duration on Hepatic HSP70, GR, and AMPKα Gene Expression in Texas Quail

Öz

Heat stress is a significant environmental factor in poultry production, negatively impacting growth performance, egg yield, and mortality rates. The present work examines how short- and long-term thermal exposure modulates liver expression patterns of heat shock protein 70 (HSP70), glucocorticoid receptor (GR), and AMP-activated protein kinase alpha (AMPKα) genes in the liver tissue of 6-week-old male Texas quail (Coturnix japonica). A total of 24 quail randomly assigned into three experimental groups: a thermoneutral control group (24 ± 1 °C), a group maintained under heat-stress conditions for an 8 hour duration (34 ± 1 °C), and maintained under heat-stress conditions for a 24 hours (34 ± 1 °C). Liver tissues were collected after heat exposure, and cDNA synthesis was performed following total RNA isolation. SYBR Green-based qRT-PCR was employed to quantify gene expression levels. o compare the experimental groups, a one-way ANOVA was used, and pairwise comparisons were conducted with the Tukey test. Compared with the control group, both heat-stressed groups exhibited a significant elevation in HSP70 expression (p < 0.001). GR expression was similar in the control group and the 8-hour heat stress group, while it was showed a significantly higher level in the 24-hour heat stress group (p = 0.003). AMPKα expression was similarly higher in both heat stress groups. In conclusion, it was shown that the liver tissue of Texas quail develops a significant molecular response to acute heat stress. The expression profiles of HSP70, GR, and AMPKα genes suggest that they can be used as potential molecular biomarkers in the assessment of heat stress in poultry.

Anahtar Kelimeler

• Gene regulation
• Heat shock protein 70
• Japanese quail
• Thermal stress response

Teksas Bıldırcınlarında Akut Isı Stresi Süresinin Karaciğer HSP70, GR ve AMPKα Gen Ekspresyonuna Etkisi

Öz

Isı stresi, kümes hayvanı üretiminde önemli bir çevresel faktördür ve büyüme performansını, yumurta verimini ve ölüm oranlarını olumsuz etkiler. Bu çalışma, kısa ve uzun süreli ısıya maruz kalmanın, 6 haftalık erkek Teksas bıldırcınlarının (Coturnix japonica) karaciğer dokusunda ısı şok proteini 70 (HSP70), glukokortikoid reseptörü (GR) ve AMP-aktive protein kinaz alfa (AMPKα) genlerinin ekspresyon paternlerini nasıl modüle ettiğini incelemektedir. Toplam 24 bıldırcın rastgele üç deney grubuna ayrılmıştır: termonötr kontrol grubu (24 ± 1 °C), 8 saat süreyle ısı stresi koşullarında tutulan grup (34 ± 1 °C) ve 24 saat süreyle ısı stresi koşullarında tutulan grup (34 ± 1 °C). Isıya maruz kalmanın ardından karaciğer dokuları toplanmış ve toplam RNA izolasyonunun ardından cDNA sentezi yapılmıştır. Gen ekspresyon seviyelerini ölçmek için SYBR Green bazlı qRT-PCR kullanılmıştır. Deneysel grupları karşılaştırmak için tek yönlü ANOVA kullanıldı ve ikili karşılaştırmalar Tukey testi ile yapıldı. Kontrol grubuyla karşılaştırıldığında, her iki ısı stresi grubunda da HSP70 ekspresyonunda anlamlı bir artış gözlendi (p < 0.001). GR ekspresyonu kontrol grubu ve 8 saatlik ısı stresi grubunda benzerdi, ancak 24 saatlik ısı stresi grubunda anlamlı derecede daha yüksek bir seviye gösterdi (p = 0.003). AMPKα ekspresyonu her iki ısı stresi grubunda da benzer şekilde daha yüksekti. Sonuç olarak, Teksas bıldırcınlarının karaciğer dokusunun akut ısı stresine karşı önemli bir moleküler yanıt geliştirdiği gösterilmiştir. HSP70, GR ve AMPKα genlerinin ekspresyon profilleri, bunların kümes hayvanlarında ısı stresinin değerlendirilmesinde potansiyel moleküler biyobelirteçler olarak kullanılabileceğini düşündürmektedir.

Anahtar Kelimeler

• Gen regülasyonu
• Isı şok proteini 70
• Japon bıldırcını
• Termal stres yanıtı

Kaynakça

1. Azad, M.A.K., Kikusato, M., Sudo, S., Amo, T., & Toyomizu, M. (2010). Time course of ROS production in skeletal muscle mitochondria from chronic heat-exposed broiler chicken. Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 157(3), 266-271. https://doi.org/10.1016/j.cbpa.2010.07.011
2. Beuving, G., & Vonder, G.M. (1978). Effect of stressing factors on corticosterone levels in the plasma of laying hens. General and Comparative Endocrinology, 35(2), 153–159. https://doi.org/10.1016/0016-6480(78)90157-0
3. Garcia, D., & Shaw, R.J. (2017). AMPK: mechanisms of cellular energy sensing and restoration of metabolic balance. Molecular Cell, 66(6), 789-800. http://doi.org/10.1016/j.molcel.2017.05.032
4. Gasparino, E., Del Vesco, A.P., Voltolini, D.M., Nascimento, C.D., Batista, E., Khatlab, A. S., ... & Guimarães, S.E.F. (2014). The effect of heat stress on GHR, IGF-I, ANT, UCP and COXIII mRNA expression in the liver and muscle of high and low feed efficiency female quail. British Poultry Science, 55(4), 466-473. https://doi.org/10.1080/00071668.2014.925090
5. Geraert, P.A., Padilha, J.C., & Guillaumin, S. (1996). Metabolic and endocrine changes induced by chronic heat exposure in broiler chickens: growth performance, body composition and energy retention. British Journal of Nutrition, 75(2), 195–204. https://doi.org/10.1017/BJN19960124
6. Habashy, W.S., Milfort, M.C., Fuller, A.L., Attia, Y.A., Rekaya, R., &Aggrey, S.E. (2017). Effect of heat stress on protein utilization and nutrient transporters in meat-type chickens. International Journal of Biometeorology, 61(12), 2111-2118. https://doi.org/10.1007/s00484-017-1414-1
7. Hardie, D.G. (2011). AMP-activated protein kinase—an energy sensor that regulates all aspects of cell function. Genes & Development, 25(18), 1895-1908. https://doi.org/10.1101/gad.17420111
8. Hardie, D.G., Ross, F.A., & Hawley, S.A. (2012). AMPK: a nutrient and energy sensor that maintains energy homeostasis. Nature Reviews Molecular Cell Biology, 13(4), 251-262. https://doi.org/10.1038/nrm3311

9. Hartl, F.U., Bracher, A., & Hayer-Hartl, M. (2011). Molecular chaperones in protein folding and proteostasis. Nature, 475(7356), 324-332. https://doi.org/10.1038/nature10317
10. Kampinga, H.H., & Craig, E.A. (2010). The HSP70 chaperone machinery: J proteins as drivers of functional specificity. Nature Reviews Molecular Cell Biology, 11(8), 579-592. https://doi.org/10.1038/nrm2941
11. Lara, L.J., & Rostagno, M.H. (2013). Impact of heat stress on poultry production. Animals, 3(2), 356-369. https://doi.org/10.3390/ani3020356
12. Liu, L., Fu, C., Yan, M., Xie, H., Li, S., Yu, Q., ... & He, J. (2016). Resveratrol modulates intestinal morphology and HSP70/90, NF-κB and EGF expression in the jejunal mucosa of black-boned chickens on exposure to circular heat stress. Food & Function, 7(3), 1329-1338. https://doi.org/10.1039/C5FO01338K
13. Livak, K. J., & Schmittgen, T. D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method. Methods, 25(4), 402-408. https://doi.org/10.1006/meth.2001.1262
14. Loyau, T., Berri, C., Bedrani, L., Metayer-Coustard, S., Praud, C., Duclos, M.J., ... & Collin, A. (2013). Thermal manipulation of the embryo modifies the physiology and body composition of broiler chickens reared in floor pens without affecting breast meat processing quality. Journal of Animal Science, 91(8), 3674-3685. https://doi.org/10.2527/jas.2013-6445
15. Mashaly, M.M., Hendricks 3rd, G.L., Kalama, M.A., Gehad, A.E., Abbas, A.O., & Patterson, P.H. (2004). Effect of heat stress on production parameters and immune responses of commercial laying hens. Poultry Science, 83(6), 889-894. https://doi.org/10.1093/ps/83.6.889
16. Mayer, M.P., & Bukau, B. (2005). Hsp70 chaperones: cellular functions and molecular mechanism. Cellular and Molecular Life Sciences, 62(6), 670. https://doi.org/10.1007/s00018-004-4464-6
17. Minvielle,, F. (2004). The future of Japanese quail for research and production. World’s Poultry Science Journal, 60(4), 500-507. https://doi.org/10.1079/WPS200433
18. Mujahid, A., Yoshiki, Y., Akiba, Y., & Toyomizu, M. (2005). Superoxide radical production in chicken skeletal muscle induced by acute heat stress. Poultry Science, 84(2), 307–314. https://doi.org/10.1093/ps/84.2.307
19. Oakley, R.H., & Cidlowski, J.A. (2013). The biology of the glucocorticoid receptor: new signaling mechanisms in health and disease. Journal of Allergy and Clinical Immunology, 132(5), 1033-1044. https://doi.org/10.1016/j.jaci.2013.09.007
20. Revollo, J.R., & Cidlowski, J.A. (2009). Mechanisms generating diversity in glucocorticoid receptor signaling. Annals of the New York Academy of Sciences, 1179(1), 167-178. https://doi.org/10.1111/j.1749-6632.2009.04986.x
21. Richter, K., Haslbeck, M., & Buchner, J. (2010). The heat shock response: life on the verge of death. Molecular Cell, 40(2), 253-266. https://doi.org/10.1016/j.molcel.2010.10.006
22. Santana, T.P., Gasparino, E., de Souza Khatlab, A., Pereira, A.M.F.E., Barbosa, L.T., Fernandes, R.P.M., ... & Del Vesco, A.P. (2023). Effects of maternal methionine supplementation on the response of Japanese quail (Coturnix coturnix japonica) chicks to heat stress. Journal of Animal Science, 101, skad042. https://doi.org/10.1093/jas/skad042
23. Scanes, C.G. (2016). Biology of stress in poultry with emphasis on glucocorticoids and the heterophil to lymphocyte ratio. Poultry Science, 95(9), 2208-2215. https://doi.org/10.3382/ps/pew137
24. St-Pierre, N.R., Cobanov, B., & Schnitkey, G. (2003). Economic losses from heat stress by US livestock industries. Journal of Dairy Science, 86, E52-E77. https://doi.org/10.3168/jds.S0022-0302(03)74040-5
25. Thornton, P.K., van de Steeg, J., Notenbaert, A., & Herrero, M. (2009). The impacts of climate change on livestock and livestock systems in developing countries: A review of what we know and what we need to know. Agricultural Systems, 101(3), 113-127. https://doi.org/10.1016/j.agsy.2009.05.002
26. Wolfenson, D., Frei, Y. F., Snapir, N., & Berman, A. (1981). Heat stress effects on capillary blood flow and its redistribution in the laying hen. Pflügers Archiv, 390(1), 86-93. https://doi.org/10.1007/BF00582717
27. Yahav, S., Collin, A., Shinder, D., & Picard, M. (2004). Thermal manipulations during broiler chick embryogenesis: effects of timing and temperature. Poultry Science, 83(12), 1959-1963. https://doi.org/10.1093/ps/83.12.1959
28. Zulkifli, I., Che Norma, M.T., Chong, C.H., & Loh, T.C. (2000). Heterophil to lymphocyte ratio and tonic immobility reactions to preslaughter handling in broiler chickens treated with ascorbic acid. Poultry Science, 79(3), 402–406. https://doi.org/10.1093/ps/79.3.402