Investigation on the Expression Profile of Immune-Related Genes in Response to Lipopolysaccharide and Lipoteichoic Acid in Alveolar Macrophages of Akkaraman and Romanov Lambs

Year 2022, Volume: 5 Issue: 1, 7 - 23, 27.06.2022

Ahmed Qasim Naji Md Mahmodul Hasan Sohel Saif Adil Abbood Al-janabi Ghulam Asghar Sajid Mehmet Ulaş Çınar

https://doi.org/10.51970/jasp.1050658

Abstract

The alveolar macrophages (AMs) are frontier of defense against foreign materials that initiate immune response in lungs. Knowledge of the expression dynamics of major immune-related genes in the alveolar macrophages in response to lipopolysaccharide (LPS) and lipoteichoic acid (LTA) challenge can help to understand disease mechanism involved in several respiratory diseases. The aim of this study was to investigate the mRNA expression of selected immune-related genes in response to lipopolysaccharide (LPS) and lipoteichoic acid (LTA) challenge in sheep alveolar macrophages in vivo. Results revealed that Romanov lambs exhibited higher mRNA expression of TLR2, TLR4, NF-ĸβ, TNFα, IL-1β, IL-6, IL-8, and IL-10 genes as compared to Akkaraman lambs along with the control of all treatments. Moreover, the expression of TLR2, TLR4, NF-ĸβ, TNFα, IL-1β, IL-6, IL-8, and IL-10 genes was higher in combine treatment of LPS and LTA as compared to separate treatments of LPS and LTA in both breeds. The results showed that the mRNA expression of immune-related genes was significantly increased in the sheep AMs in response to LPS and LTA treatment whereas a synergistic effect was observed in LPS+LTS treatment. Also, breed comparison showed that the native Akkaraman was more resistant to disease compared to exotic Romanov.

Keywords

Cytokines, Immunogenetics, mRNA expression, Respiratory diseases, Ovine, TLRs pathway

Supporting Institution

Erciyes University Scientific Research Projects Unit

Project Number

FDK-2017-7657

Thanks

Authors are indebted to Agricultural Research-Application (ERÜTAM) Center at Erciyes University for providing animal facilities and Betül-Ziya Eren Genome and Stem Cell Research Center at Erciyes University (GENKÖK) for providing laboratory space and equipment. The authors are also thankful to the staff of the Department of Veterinary Genetics at Erciyes University for their help during animal experiments and sample collection. This project was supported by Erciyes University Scientific Research Projects Unit under the code of FDK-2017-7657.

Akkaraman ve Romanov Kuzularının Alveolar Makrofajlarında Lipopolisakkarit ve Lipoteikoik Asite Yanıtta İmmün İlişkili Genlerin Ekspresyon Profilinin Araştırılması

Abstract

Alveolar makrofajlar (AM'lar), akciğerlerde bağışıklık tepkisini başlatan yabancı maddelere karşı savunma bariyeridir. Lipopolisakkarit (LPS) ve lipoteikoik asit (LTA) tehdidine yanıt olarak alveolar makrofajlardaki majör immün bağlantılı genlerin ekspresyon dinamiklerinin bilinmesi, çeşitli solunum yolu hastalıklarında yer alan hastalık mekanizmasının anlaşılmasına yardımcı olabilir. Bu çalışmanın amacı, koyun alveolar makrofajlarında lipopolisakkarit (LPS) ve lipoteikoik asit (LTA) stimülasyonuna yanıt olarak seçilen bağışıklıkla ilgili genlerin mRNA ekspresyonunu in vivo araştırmaktır. Sonuçlar, Romanov kuzularının TLR2, TLR4, NF-ĸβ, TNFa, IL-1β, IL-6, IL-8 ve IL-10 genlerinin mRNA ekspresyonunu Akkaraman kuzularına kıyasla kontrol grubuna göre daha yüksek saptandığını ortaya koymuştur. Ayrıca, her iki ırkta LPS ve LTA'nın kombine uygulanmasında TLR2, TLR4, NF-ĸβ, TNFa, IL-1β, IL-6, IL-8 ve IL-10 genlerinin ekspresyonu, LPS ve LTA'nın ayrı uygulamalarına kıyasla daha yüksektir cinste. Çalışmanın sonuçları, LPS ve LTA uygulamalarına yanıt olarak koyun AM'larında bağışıklıkla ilgili genlerin mRNA ekspresyonunun önemli ölçüde arttığını, LPS+LTS uygulamasında ise sinerjik bir etkinin gözlemlendiğini gösterdi. Ayrıca, ırk karşılaştırması, yerli Akkaraman'ın Romanov'a kıyasla hastalığa karşı daha dirençli olduğunu gösterdi.

Keywords

sitokinler, immunogenetik, mRNA ekspresyonu, solunum yolu hastalıkları, koyun, TLRs yolu

Project Number

FDK-2017-7657

References

• Akira, S., Uematsu, S., Takeuchi, O., 2006. Pathogen recognition and innate immunity. Cell. 124(4), 783-801.
• Aksel, E. G., Akyüz, B., 2021. Effect of LPS and LTA stimulation on the expression of TLR-pathway genes in PBMCs of Akkaraman lambs in vivo. Tropical Animal Health and Production. 53(1), 1-9.
• Barbalat, R., Lau, L., Locksley R. M., Barton, G. M., 2009. Toll-like receptor 2 on inflammatory monocytes induces type I interferon in response to viral but not bacterial ligands. Nature Immunology, 10(11), 1200-1207.
• Choussat, R., Montalescot, G., Collet, J. P., Jardel, C., Ankri, A., Fillet, A. M., … Thomas, D., 2000. Effect of prior exposure to Chlamydia pneumoniae, Helicobacter pylori, or cytomegalovirus on the degree of inflammation and one-year prognosis of patients with unstable angina pectoris or non–q-wave acute myocardial infarction. The American Journal of Cardiology, 86(4), 379-384.
• Cinar, M. U., Islam, M. A., Uddin, M. J., Tholen, E., Tesfaye, D., Looft, C., Schellander, K., 2012. Evaluation of suitable reference genes for gene expression studies in porcine alveolar macrophages in response to LPS and LTA. BMC Research Notes, 5(1), 1-14.
• Cohen, S. B. A., Crawley, J. B., Kahan, M. C., Feldmann, M., Foxwell, B. M. J., 1997. Interleukin‐10 rescues T cells from apoptotic cell death: association with an upregulation of Bcl‐2. Immunology, 92(1), 1-5.
• Daly, K. A., Mailer, S. L., Digby, M. R., Lefévre, C., Thomson, P., Deane, E., Williamson, P., 2009. Molecular analysis of tammar (Macropus eugenii) mammary epithelial cells stimulated with lipopolysaccharide and lipoteichoic acid. Veterinary Immunology and Immunopathology, 129(1-2), 36-48.
• Dempsey, P. W., Vaidya, S. A., Cheng, G., 2003. The art of war: Innate and adaptive immune responses. Cellular and Molecular Life Sciences CMLS, 60(12), 2604-2621.
• Donaldson, L., Vuocolo, T., Gray, C., Strandberg, Y., Reverter, A., McWilliam, S., Tellam, R., 2005. Construction and validation of a bovine innate immune microarray. BMC Genomics, 6(1), 1-22.
• Duffield, J. S., Lupher, M., Thannickal, V. J., Wynn, T. A., 2013. Host responses in tissue repair and fibrosis. Annual Review of Pathology: Mechanisms of Disease, 8, 241-276.
• Gao, Y., Flori, L., Lecardonnel, J., Esquerré, D., Hu, Z. L., Teillaud, A., Lemonnier, G., Lefèvre, F., Oswald, I. P., Rogel, G. C. 2010. Transcriptome analysis of porcine PBMCs after in vitro stimulation by LPS or PMA/ionomycin using an expression array targeting the pig immune response. BMC Genomics, 11:292.
• González, J. M., 2015. Factores que condicionan la supervivencia de los corderos tipo Ternasco. Estudio del complejo respiratorio ovino: presentación, formas lesionales, agentes implicados y serotipificacion de Pasterella haemolytica. Ph. D. Thesis, University of Zaragoza, Spain.
• Goodwin, K. A., Jackson, R., Brown, C., Davies, P. R., Morris, R. S., Perkins, N. R., 2004. Pneumonic lesions in lambs in New Zealand: patterns of prevalence and effects on production. New Zealand Veterinary Journal, 52(4), 175-179.
• Hack, C. E., De Groot, E. R., Felt-Bersma, R. J., Nuijens, J. H., Strack, Van, R. J., Eerenberg-Belmer, A. J., Thijs, L. G, Aarden, L. A., 1989. Increased plasma levels of interleukin-6 in sepsis. Blood, 74(5):1704-10.
• Hoebe, K., Janssen, E., Beutler, B., 2004. The interface between innate and adaptive immunity. Nature Immunology. 5(10):971-4.
• Hoshino, K., Takeuchi, O., Kawai, T., Sanjo, H., Ogawa, T., Takeda, Y., Takeda, K. Akira S., 1999. Cutting edge: Toll-like receptor 4 (TLR4)-deficient mice are hyporesponsive to lipopolysaccharide: evidence for TLR4 as the Lps gene product. The Journal of Immunology, 162(7), 3749-3752.
• Hu, G., Christman, J. W., 2019. Alveolar macrophages in lung inflammation and resolution. Frontiers in Immunology, 10, 2275.
• Hussell, T., Bell T. J., 2014. Alveolar macrophages: plasticity in a tissue-specific context. Nature Reviews Immunology. 14(2):81-93.
• Ishida, H., Muchamuel, T., Sakaguchi, S., Andrade, S., Menon, S., Howard, M., 1994. Continuous administration of anti-interleukin 10 antibodies delays onset of autoimmunity in NZB/W F1 mice. The Journal of Experimental Medicine. 179(1), 305-310.
• Islam, M. A., Cinar, M. U., Uddin, M. J., Tholen, E., Tesfaye, D., Looft, C. Schellander, K., 2012b. Expression of Toll-like receptors and downstream genes in lipopolysaccharide-induced porcine alveolar macrophages. Veterinary Immunology and Immunopathology. 146(1), 62-73.
• Islam, M. A., Pröll, M., Hölker, M., Tholen, E., Tesfaye, D., Looft, C., Schellander, K., Cinar, M. U., 2013. Alveolar macrophage phagocytic activity is enhanced with LPS priming, and combined stimulation of LPS and lipoteichoic acid synergistically induce pro-inflammatory cytokines in pigs. Innate Immunity. 19(6), 631-643.
• Islam, M. A., Uddin, M. J., Tholen, E., Tesfaye, D., Looft, C., Schellander, K., Cinar, M. U., 2012a. Age-related changes in phagocytic activity and production of pro-inflammatory cytokines by lipopolysaccharide stimulated porcine alveolar macrophages. Cytokine. 60(3), 707-717.
• Koressaar T, Remm M., 2007. Enhancements and modifications of primer design program Primer3. Bioinformatics. 23(10), 1289-1291.
• Lacasta, D., González, J. M., Navarro, T., Saura, F., Acín, C., Vasileiou, N. G. C., 2019. Significance of respiratory diseases in the health management of sheep. Small Ruminant Research. 181, 99-102.
• Luzón, J., 1999. Influencia de las Afecciones Respiratorias en los Principales Parámetros Productivos de los Corderos Tipo Ternasco (Influence of Respiratory Diseases on the Main Productive Parameters of Ternasco-Type Lambs) (Doctoral dissertation, University of Zaragoza).
• McRae, K. M., Baird, H. J., Dodds, K. G., Bixley, M. J., Clarke, S. M., 2016. Incidence and heritability of ovine pneumonia, and the relationship with production traits in New Zealand sheep. Small Ruminant Research. 145, 136-141.
• Medzhitov, R., 2007. Recognition of microorganisms and activation of the immune response. Nature. 449, 819–826.
• Micera, A., Stampachiacchiere, B., Normando, E. M., Lambiase, A., Bonini, S., Bonini, S., 2009. Nerve growth factor modulates toll-like receptor (TLR) 4 and 9 expression in cultured primary VKC conjunctival epithelial cells. Molecular vision. 15, 2037.
• Mosser, D. M., Edwards, J. P., 2008. Exploring the full spectrum of macrophage activation. Nature Reviews Immunology. 8(12), 958-969.
• Murphy, K., Weaver, C., 2016. Janeway's Immunobiology. Garland science.
• Nettle, R., Paine, M., Penry J., 2010. Aligning farm decision making and genetic information systems to improve animal production: methodology and findings from the Australian dairy industry. Animal Production Science. 50(6), 429-434.
• Perkins, D. J., Vogel, S. N., 2015. Space and time: New considerations about the relationship between Toll-like receptors (TLRs) and type I interferons (IFNs). Cytokine. 74(2), 171-4.
• Pinilla-Vera, M., Xiong, Z., Zhao, Y., Zhao, J., Donahoe, M. P., Barge, S., Horne, W. T., Kolls, J. K., McVerry, B. J., Birukova, A, Tighe, R. M., 2016. Full spectrum of LPS activation in alveolar macrophages of healthy volunteers by whole transcriptomic profiling. PloS One. 11(7), p.e0159329.
• Raymond, C. R., Wilkie, B. N., 2005. Toll-like receptor, MHC II, B7 and cytokine expression by porcine monocytes and monocyte-derived dendritic cells in response to microbial pathogen-associated molecular patterns. Veterinary immunology and immunopathology, 107(3-4), pp.235-247.
• Rock, J. R., Hogan, B. L., 2011. Epithelial progenitor cells in lung development, maintenance, repair, and disease. Annual review of cell and developmental biology. 27, 493-512. Schwandner, R., Dziarski, R., Wesche, H., Rothe, M., Kirschning C. J., 1999. Peptidoglycan-and lipoteichoic acid-induced cell activation is mediated by toll-like receptor 2. Journal of Biological Chemistry. 274(25), 17406-17409.
• Sohel, M. M. H., Konca, Y., Akyuz, B., Arslan, K., Sariozkan, S., Cinar, M. U., 2017. Concentration dependent antioxidative and apoptotic effects of sulforaphane on bovine granulosa cells in vitro. Theriogenology. 97, 17-26.
• Sriskandan, S., Cohen, J., 1999. Gram-positive sepsis. Mechanisms and differences from gram-negative sepsis. Infectious Disease Clinics of North America. 13(2):397-412.
• Su, S. C., Hua, K. F., Lee, H., Chao, L. K., Tan, S. K., Yang, S. F., Hsu, H. Y., 2006. LTA and LPS mediated activation of protein kinases in the regulation of inflammatory cytokines expression in macrophages. Clinica Chimica Acta. 374(1-2), 106-115.
• Surh, Y. J., Chun, K. S., Cha, H. H., Han, S. S., Keum, Y. S., Park, K. K., Lee, S. S., 2001. Molecular mechanisms underlying chemopreventive activities of anti-inflammatory phytochemicals: down-regulation of COX-2 and iNOS through suppression of NF-κB activation. Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis. 480, 243-268.
• Thompson, S. L., Dhar, V., Bond, M. W., Mosmann, T. R., Moore, K. W., Rennick, D. M., (1991). Interleukin 10: a novel stimulatory factor for mast cells and their progenitors. Journal of Experimental Medicine, 173(2):507-10.
• Thorley, A. J., Ford, P. A., Giembycz, M. A., Goldstraw, P., Young, A., Tetley, T. D., 2007. Differential regulation of cytokine release and leukocyte migration by lipopolysaccharide-stimulated primary human lung alveolar type II epithelial cells and macrophages. The Journal of Immunology. 178(1), 463-473.
• Twigg, H. L., 2004. Macrophages in innate and acquired immunity. Thieme. 25(1):21-31.
• Uebelhoer, M., Bewig, B., Sternberg, K., Rabe, K., Nowak, D., Magnussen, H., Barth, J., 1995. Alveolar macrophages from bronchoalveolar lavage of patients with pulmonary histiocytosis X: determination of phenotypic and functional changes. Lung. 173(3), 187-195.
• Wang, C., Deng, L., Hong, M., Akkaraju, G. R., Inoue, J. I., Chen, Z. J., 2001. TAK1 is a ubiquitin-dependent kinase of MKK and IKK. Nature. 412(6844), 346-351.
• Wang, T., Lafuse, W. P., Zwilling, B. S., 2000. Regulation of toll-like receptor 2 expression by macrophages following Mycobacterium avium infection. The Journal of Immunology. 165(11), 6308-6313.
• Wickham, H., 2009. ggplot2: elegant graphics for data analysis. Springer New York.
• Yang, Q., Pröll, M. J., Salilew-Wondim, D., Zhang, R., Tesfaye, D., Fan, H., Cinar, M. U., Große-Brinkhaus, C., Tholen, E., Islam, M. A., Hölker, M., 2016. LPS-induced expression of CD14 in the TRIF pathway is epigenetically regulated by sulforaphane in porcine pulmonary alveolar macrophages. Innate Immunity. 22(8), 682-695.
• Yoshimura, A., Lien, E., Ingalls, R. R., Tuomanen, E., Dziarski, R., Golenbock, D., 1999. Cutting edge: recognition of Gram-positive bacterial cell wall components by the innate immune system occurs via Toll-like receptor 2. The Journal of Immunology. 163(1), 1-5.
• Zhang, M., Jin, X., Yang Y. F., 2019. β-Glucan from Saccharomyces cerevisiae induces SBD-1 production in ovine ruminal epithelial cells via the Dectin-1–Syk–NF-κB signaling pathway. Cellular Signaling. 53, 304-315.
• Zhou, Y., Murthy, J. N., Zeng, D., Belardinelli, L., Blackburn, M. R., 2010. Alterations in adenosine metabolism and signaling in patients with chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis. PloS One. 5(2), e9224.