Animal Bioscience

Asian Australasian Association of Animal Production Societies (아세아태평양축산학회)
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Regulation of toll-like receptors expression in muscle cells by exercise-induced stress

  • Park, Jeong-Woong (Department of Animal Science, College of Natural Resources and Life Sciences, Pusan National University) ;
  • Kim, Kyung-Hwan (Department of Animal Science, College of Natural Resources and Life Sciences, Pusan National University) ;
  • Choi, Joong-Kook (Division of Biochemistry, College of Medicine, Chungbuk National University) ;
  • Park, Tae Sub (Institute of Green-Bio Science and Technology, Seoul National University) ;
  • Song, Ki-Duk (The Animal Molecular Genetics and Breeding Center, Jeonbuk National University) ;
  • Cho, Byung-Wook (Department of Animal Science, College of Natural Resources and Life Sciences, Pusan National University)
  • Received : 2020.07.13
  • Accepted : 2020.12.01
  • Published : 2021.10.01
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Abstract

Objective: This study investigates the expression patterns of toll-like receptors (TLRs) and intracellular mediators in horse muscle cells after exercise, and the relationship between TLRS expression in stressed horse muscle cells and immune cell migration toward them. Methods: The expression patterns of the TLRs (TLR2, TLR4, and TLR8) and downstream signaling pathway-related genes (myeloid differentiation primary response 88 [MYD88]; activating transcription factor 3 [ATF3]) are examined in horse tissues, and horse peripheral blood mononuclear cells (PBMCs), polymorphonuclear cells (PMNs) and muscles in response to exercise, using the quantitative reverse transcription-polymerase chain reaction (qPCR). Expressions of chemokine receptor genes, i.e., C-X-C motif chemokine receptor 2 (CXCR2) and C-C motif chemokine receptor 5 (CCR5), are studied in PBMCs and PMNs. A horse muscle cell line is developed by transfecting SV-T antigen into fetal muscle cells, followed by examination of muscle-specific genes. Horse muscle cells are treated with stressors, i.e., cortisol, hydrogen peroxide (H2O2), and heat, to mimic stress conditions in vitro, and the expression of TLR4 and TLR8 are examined in stressed muscle cells, in addition to migration activity of PBMCs toward stressed muscle cells. Results: The qPCR revealed that TLR4 message was expressed in cerebrum, cerebellum, thymus, lung, liver, kidney, and muscle, whereas TLR8 expressed in thymus, lung, and kidney, while TLR2 expressed in thymus, lung, and kidney. Expressions of TLRs, i.e., TLR4 and TLR8, and mediators, i.e., MYD88 and ATF3, were upregulated in muscle, PBMCs and PMNs in response to exercise. Expressions of CXCR2 and CCR5 were also upregulated in PBMCs and PMNs after exercise. In the muscle cell line, TLR4 and TLR8 expressions were upregulated when cells were treated with stressors such as cortisol, H2O2, and heat. Migration of PBMCs toward stressed muscle cells was increased by exercise and oxidative stresses, and combinations of these. Treatment with methylsulfonylmethane (MSM), an antioxidant on stressed muscle cells, reduced migration of PBMCs toward stressed muscle cells. Conclusion: In this study, we have successfully cultured horse skeletal muscle cells, isolated horse PBMCs, and established an in vitro system for studying stress-related gene expressions and function. Expression of TLR4, TLR8, CXCR2, and CCR5 in horse muscle cells was higher in response to stressors such as cortisol, H2O2, and heat, or combinations of these. In addition, migration of PBMCs toward muscle cells was increased when muscle cells were under stress, but inhibition of reactive oxygen species by MSM modulated migratory activity of PBMCs to stressed muscle cells. Further study is necessary to investigate the biological function(s) of the TLR gene family in horse muscle cells.

Keywords

Acknowledgement

This work was supported by grants from the Individual Basic Science & Engineering Research Program (2017R1D1A1B03036432), the National Research Foundation of Korea, and the Next-Generation BioGreen 21 Program (Project No. PJ01325701), Rural Development Administration, Republic of Korea. Finally, we would like to thank the Writing Center at Jeonbuk National University for their language assistance, which we think readers will agree has greatly enhanced the readability of the manuscript.

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