Biological Trace Element Research

, Volume 189, Issue 1, pp 209–223 | Cite as

Transcriptional Study Revealed That Boron Supplementation May Alter the Immune-Related Genes Through MAPK Signaling in Ostrich Chick Thymus

  • Ke Xiao
  • Keli Yang
  • Jing Wang
  • Pengpeng Sun
  • Haibo Huang
  • Haseeb Khaliq
  • Muhammad Ahsan Naeem
  • Juming Zhong
  • Kemei PengEmail author


The objective of this study is to construct a digital gene expression tag profile to identify genes potentially related to immune response in the ostrich. Exposure to boron leads to an immune response in the ostrich, although the underlying mechanism remains obscure. Thus, a dire need of biological resource in the form of transcriptomic data for ostriches arises to key out genes and to gain insights into the function of boron on the immune response of thymus. For this purpose, RNA-Seq analysis was performed using the Illumina technique to investigate differentially expressed genes in ostrich thymuses treated with different boric acid concentrations (0, 80, and 640 mg/L). Compared with the control group, we identified 309 upregulated and 593 downregulated genes in the 80 mg/L treated sample and 228 upregulated and 1816 downregulated genes in 640 mg/L treated sample, respectively. Trend analysis of these differentially expressed genes uncovers three statistically significant trends. Functional annotation analysis of the differentially expressed genes verifies multiple functions associated with immune response. When ostrich thymuses were treated with boron, expression changes were observed in genes predominantly associated with MAPK and calcium signaling pathways. The results of this study provide all-inclusive information on gene expression at the transcriptional level that further enhances our apprehension for the molecular mechanisms of boron on the ostrich immune system. The calcium and MAPK signaling pathways might play a pivotal role in regulating the immune response of boron-treated ostriches.


Ostrich thymus Boron Transcriptome Immune response MAPK 



Illumina RNA sequencing


differentially expressed genes


gene ontology


Kyoto Encyclopedia of Genes and Genomes


80 mg/L boric acid


640 mg/L boric acid


quantitative reverse transcription PCR


mitogen-activated protein kinase


extracellular signal-related kinase


phospho-extracellular signal-related kinase


Jun amino-terminal kinases


phospho-Jun amino-terminal kinase


p38 mitogen-activated protein kinase


phospho-p38 mitogen-activated protein kinase


protein phosphatase 3, catalytic subunit, alpha isozyme


protein phosphatase 2B regulatory subunit 1


nuclear factor of activated T cells


myocyte enhancer factor 2C


polyvinylidene fluoride membranes


Funding information

This study was supported by the National Natural Science Foundation Projects of China (No. 31672504), the National Natural Youth Science Foundation of China (No. 31702196), and the National Science Foundation for Post-doctoral Scientists of China (No. 2017M612482).

Compliance with Ethical Standards

The present study was approved by the Ethics Committee of Huazhong Agricultural University. Protocols of animals were performed according to the No. 5 Proclamation of the standing Committee of Hubei People’s Congress, People’s Republic of China.

Conflict of Interest

The authors declare that they have no competing interest.

Supplementary material

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12011_2018_1441_MOESM3_ESM.tif (8.6 mb)
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12011_2018_1441_MOESM4_ESM.doc (30 kb)
ESM 4 (DOC 29 kb)


  1. 1.
    Johnston P (2011) New morphological evidence supports congruent phylogenies and Gondwana vicariance for palaeognathous birds. Zool J Linn Soc-Lond 163(3):959–982CrossRefGoogle Scholar
  2. 2.
    Mitchell KJ, Llamas B, Soubrier J, Rawlence NJ, Worthy TH, Wood J (2014) Ancient DNA reveals elephant birds and kiwi are sister taxa and clarifies ratite bird evolution. Science 344(6186):898–900CrossRefGoogle Scholar
  3. 3.
    Alnasser A, Alkhalaifa H, Holleman K, Al-Ghalaf W (2003) Ostrich production in the arid environment of Kuwait. J Arid Environ 54(1):219–224CrossRefGoogle Scholar
  4. 4.
    Bejaei M, Cheng KM (2014) A survey of current ostrich handling and transport practices in North America with reference to ostrich welfare and transportation guidelines set up in other countries. Poultry Sci 93(2):296–306CrossRefGoogle Scholar
  5. 5.
    Pittaway T, Niekerk PV (2015) Horizon-scanning the ostrich industry with bibliometric indicators. AfJARE 10:64–71Google Scholar
  6. 6.
    Bonato M, Evans MR, Hasselquist D, Sherley RB, Cloete SWP, Cherry MI (2013) Ostrich chick humoral immune responses and growth rate are predicted by parental immune responses and paternal colouration. Behav Ecol Sociobiol 67(12):1891–1901CrossRefGoogle Scholar
  7. 7.
    Kabu M, Akosman MS (2013) Biological effects of boron. Reviews of environmental contamination and toxicology. Springer, New York, pp 57–75CrossRefGoogle Scholar
  8. 8.
    Kabu M, Civelek T (2012) Effects of propylene glycol, methionine and sodium borate on metabolic profile in dairy cattle during periparturient period. Rev Med Vet-Toulouse 163(8):419–430Google Scholar
  9. 9.
    Jin E, Gu Y, Wang J, Jin G, Li S (2014) Effect of supplementation of drinking water with different levels of boron on performance and immune organ parameters of broilers. Ital J Anim Sci 13(2):124CrossRefGoogle Scholar
  10. 10.
    Xiao K, Ansari AR, Rehman Z, Khaliq H, Song H, Tang J, Wang J, Wang W, Sun PP, Zhong J, Peng KM (2015) Effect of boric acid supplementation of ostrich water on the expression of Foxn1 in thymus. Histol Histopathol 30(11):1367–1378Google Scholar
  11. 11.
    Jing W, Zhong JM, Sun PP, Ke X, Tang J, Wei W, Peng KM (2015) Effect of boron administration on the morphology of ostrich chick kidney tissue. Pak Vet J 35(4):489–493Google Scholar
  12. 12.
    Cheng J, Peng KM, Jin E, Zhang Y, Liu Y, Zhang N (2011) Effect of additional boron on tibias of African ostrich chicks. Biol Trace Elem Res 144(1–3):538–549CrossRefGoogle Scholar
  13. 13.
    Sun PP, Luo Y, Wu XT, Ansari AR, Wang J, Yang KL, Zhong JM, Peng KM (2016) Effects of supplemental boron on intestinal proliferation and apoptosis in African ostrich chicks. Int J Morphol 34(3):830–835CrossRefGoogle Scholar
  14. 14.
    Tang J, Zheng XT, Xiao K, Wang KL, Wang J, Wang YX, Zhong JM, Peng KM (2016) Effect of boric acid supplementation on the expression of BDNF in African ostrich chick brain. Biol Trace Elem Res 170(1):1–8CrossRefGoogle Scholar
  15. 15.
    Haseeb K, Wang J, Xiao K, Yang KL, Sun PP, Wu XT, Zhong JM, Peng KM (2017) Effects of boron supplementation on expression of Hsp70 in the spleen of African ostrich. Biol Trace Elem Res 182(4):1–11Google Scholar
  16. 16.
    Krishna M, Narang H (2008) The complexity of mitogen-activated protein kinases (MAPKs) made simple. Cell Mol Life Sci 65(22):3525–3544CrossRefGoogle Scholar
  17. 17.
    Bubici C, Papa S (2014) JNK signalling in cancer: in need of new, smarter therapeutic targets. Brit J Pharmacol 171(1):24–37CrossRefGoogle Scholar
  18. 18.
    Huang G, Shi LZ, Chi H (2009) Regulation of jnk and p38 MAPK in the immune system: signal integration, propagation and termination. Cytokine 48(3):161–169CrossRefGoogle Scholar
  19. 19.
    Kaminska B (2005) MAPK signalling pathways as molecular targets for anti-inflammatory therapy from molecular mechanisms to therapeutic benefits. BBA-Proteins Proteom 1754(1–2):253–262CrossRefGoogle Scholar
  20. 20.
    Bedognetti D, Roelands J, Decock J, Wang E, Hendrickx W (2017) The MAPK hypothesis: immune-regulatory effects of MAPK-pathway genetic dysregulations and implications for breast cancer immunotherapy 1(5): 429–445Google Scholar
  21. 21.
    Hsu SC, Wu CC, Han J, Lai MZ (2003) Involvement of p38 mitogen-activated protein kinase in different stages of thymocyte development. Blood 101(3):970–976CrossRefGoogle Scholar
  22. 22.
    Crompton T, Gilmour KC, Owen MJ (1996) The MAP kinase pathway controls differentiation from double-negative to double-positive thymocyte. Cell 86(2):243–251CrossRefGoogle Scholar
  23. 23.
    Tournier C (2013) The 2 faces of JNK signaling in cancer. Genes Cancer 4(9–10):397–400CrossRefGoogle Scholar
  24. 24.
    Hamdi M, Kool J, Cornelissen-Steijger P, Carlotti F, Popeijus HE, van der Burgt C, Janssen JM, Yasui A, Hoeben RC (2005) DNA damage in transcribed genes induces apoptosis via the JNK pathway and the JNK-phosphatase MKP-1. Oncogene 24(48):7135–7144CrossRefGoogle Scholar
  25. 25.
    Kyriakis JM, Avruch J (2001) Mammalian mitogen-activated protein kinase signal transduction pathways activated by stress and inflammation. Physiol Rev 81(2):807–869CrossRefGoogle Scholar
  26. 26.
    Roux PP, Blenis J (2004) ERK and p38 MAPK-activated protein kinases: a family of protein kinases with diverse biological functions. Microbiol Mol Biol Rev 68(2):320–344CrossRefGoogle Scholar
  27. 27.
    Cook R, Wu CC, Kang YJ, Han J (2006) The role of the p38 pathway in adaptive immunity. Cell Mol Immunol 4:253–259Google Scholar
  28. 28.
    Dodeller F, Schulze-Koops H (2006) The p38 mitogen-activated protein kinase signaling cascade in CD4 T cells. Arthritis Res Ther 8:205CrossRefGoogle Scholar
  29. 29.
    Zhang JX, Wu KL, Zeng SJ, da Silva JAT, Zhao XL, Tian CE, Xia HQ, Duan J (2013) Transcriptome analysis of cymbidium sinense and its application to the identification of genes associated with floral development. BMC Genomics 14(1):279CrossRefGoogle Scholar
  30. 30.
    Mortazavi A, Williams BA, McCue K, Schaeffer L, Wold B (2008) Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat Methods 5:621–628CrossRefGoogle Scholar
  31. 31.
    Wang Z, Gerstein M, Snyder M (2009) RNA-Seq: a revolutionary tool for transcriptomics. Nat Rev Genet 10:57–63CrossRefGoogle Scholar
  32. 32.
    Polato NR, Vera JC, Baums IB (2011) Gene discovery in the threatened elkhorn coral: 454 sequencing of the Acropora palmata transcriptome. PLoS One 6(12):e28634CrossRefGoogle Scholar
  33. 33.
    Wan L, Han J, Sang M, Li A, Wu H, Yin S, Zhang C (2012) De novo transcriptomic analysis of an oleaginous microalga: pathway description and gene discovery for production of next-generation biofuels. PLoS One 7(6):e35142CrossRefGoogle Scholar
  34. 34.
    Lu X, Kim H, Zhong S, Chen H, Hu Z, Zhou B (2014) De novo transcriptome assembly for rudimentary leaves in litchi chinesis sonn and identification of differentially expressed genes in response to reactive oxygen species. BMC Genomics 15(1):805CrossRefGoogle Scholar
  35. 35.
    Zhao X, Mo D, Li A, Gong W, Xiao S, Zhang Y, Qiu L, Guo Y, Liu X, Cong P, He Z, Wang C, Li J, Chen Y (2011) Comparative analyses by sequencing of transcriptomes during skeletal muscle development between pig breeds differing in muscle growth rate and fatness. PLoS One 6(5):e19774CrossRefGoogle Scholar
  36. 36.
    Yang C, Jiang M, Wen H, Tian J, Liu W, Wu F, Guo G (2015) Analysis of differential gene expression under low-temperature stress in Nile tilapia (oreochromis niloticus) using digital gene expression. Gene 564(2):134–140CrossRefGoogle Scholar
  37. 37.
    Wang W, Xiao K, Zheng X, Zhu D, Yang Z, Tang J, Sun P, Wang J, Peng KM (2014) Effects of supplemental boron on growth performance and meat quality in African ostrich chicks. J Agric Food Chem 62(46):11024–11029CrossRefGoogle Scholar
  38. 38.
    Audic S, Claverie JM (1997) The significance of digital gene expression profiles. Genome Res 7(10):986–995CrossRefGoogle Scholar
  39. 39.
    Ernst J, Bar-Joseph Z (2006) Stem: a tool for the analysis of short time series gene expression data. BMC Bioinformatics 7(1):191CrossRefGoogle Scholar
  40. 40.
    Kanehisa M, Araki M, Goto S, Hattori M, Hirakawa M, Itoh M, Katayama T, Kawashima S, Okuda S, Tokimatsu T, Yamanishi Y (2008) KEGG for linking genomes to life and the environment. Nucleic Acids Res 36:480–484CrossRefGoogle Scholar
  41. 41.
    Monticelli S, Rao A (2002) NFAT1 and NFAT2 are positive regulators of IL-4 gene transcription. Eur J Immunol 32(10):2971–2978CrossRefGoogle Scholar
  42. 42.
    Khaliq H, Juming Z, Kemei P (2018) The physiological role of boron on health. Biol Trace Elem Res 2:1–21Google Scholar
  43. 43.
    Hu Q, Li S, Qiao E, Tang Z, Jin E, Jin G et al (2014) Effects of boron on structure and antioxidative activities of spleen in rats. Biol Trace Elem Res 158(1):73–80CrossRefGoogle Scholar
  44. 44.
    Olgun O, Yazgan O, Cufadar Y (2013) Effect of supplementation of different boron and copper levels to layer diets on performance, egg yolk and plasma cholesterol. J Trace Elem Med Biol 27(2):132–136CrossRefGoogle Scholar
  45. 45.
    Li SH, Zhu HG, Wang J, Jin GM, Gu YF, Liu DY (2009) Effect of environmental estrogen boron on microstructure of thymus in rats. Journal of Anhui Science & Technology University (6):1–5Google Scholar
  46. 46.
    Henderson K, Stella SL, Kobylewski S, Eckhert CD (2009) Receptor activated Ca(2+) release is inhibited by boric acid in prostate cancer cells. PLoS One 4(6):e6009CrossRefGoogle Scholar
  47. 47.
    Huang HB, Xiao K, Lu S, Yang KL, Ansari AR, Haseeb K, Song H, Zhong JM, Liu HZ, Peng KM (2015) Increased thymic cell turnover under boron stress may bypass TLR3/4 pathway in African ostrich. PLoS One 10(6):e0129596CrossRefGoogle Scholar
  48. 48.
    Lee MD, Bingham KN, Mitchell TY, Meredith JL, Rawlings JS (2015) Calcium mobilization is both required and sufficient for initiating chromatin decondensation during activation of peripheral T-cells. Mol Immunol 63(2):540–549CrossRefGoogle Scholar
  49. 49.
    Li S (2005) Effect of boron on the growth, hematology and development of immune organ in Gushi Chickens (in chinese). Journal of Northwest A&F University 37(2):52–58Google Scholar
  50. 50.
    Goihl J (2002) More research needed on boron supplementation of swine diets. Feedstuffs 74:10–27Google Scholar
  51. 51.
    Macian F (2005) NFAT proteins: key regulators of T-cell development and function. Nat Rev Immunol 5(6):472–484CrossRefGoogle Scholar
  52. 52.
    Mcneil LK, Starr TK, Hogquist KA (2005) A requirement for sustained ERK signaling during thymocyte positive selection in vivo. P Natl Acad Sci USA 102(38):13574–13579CrossRefGoogle Scholar
  53. 53.
    Khiem D, Cyster JG, Schwarz JJ, Black BL (2008) A p38 MAPK-MEF2C pathway regulates β-cell proliferation. P Natl Acad Sci USA 105(44):17067–17072CrossRefGoogle Scholar
  54. 54.
    Barranco WT, Kim DH, Jr SS, Eckhert CD (2009) Boric acid inhibits stored Ca2+ release in DU-145 prostate cancer cells. Cell Biol Toxicol 25(4):309–320CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Ke Xiao
    • 1
  • Keli Yang
    • 1
  • Jing Wang
    • 1
  • Pengpeng Sun
    • 1
  • Haibo Huang
    • 1
  • Haseeb Khaliq
    • 1
  • Muhammad Ahsan Naeem
    • 1
  • Juming Zhong
    • 2
  • Kemei Peng
    • 1
    Email author
  1. 1.College of Veterinary MedicineHuazhong Agricultural UniversityWuhanPeople’s Republic of China
  2. 2.College of Veterinary MedicineAuburn UniversityAuburnUSA

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