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Transcriptional and posttranscriptional regulation of CXCL8/IL-8 gene expression induced by connective tissue growth factor

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Abstract

Connective tissue growth factor (CTGF), a CCN family member, is a secreted protein regulating cellular functions, including fibrosis, apoptosis, adhesion, migration, differentiation, proliferation, angiogenesis, and chondrogenesis. CTGF increases proinflammatory factor production; however, inflammatory cytokine regulation by CTGF is poorly understood. The aim of this study was to identify novel biological functions and elucidate the functional mechanisms of CTGF. Specifically, the study focused on the ability of CTGF-primed monocytes to secrete interleukin 8 (CXCL8/IL-8) and determined the signaling pathways involved in CTGF-induced CXCL8/IL-8 gene regulation during inflammation. We transfected wild-type or mutant CXCL8/IL-8 promoter-derived luciferase reporter constructs into 293T cells to examine the effect of CTGF on the CXCL8/IL-8 promoter. The results showed that the activator protein-1 and nuclear factor κB binding sites of the CXCL8/IL-8 promoter are essential for CTGF-induced CXCL8/IL-8 transcription. Moreover, the CTGF-induced activation of p38 mitogen-activated protein kinase (MAPK), c-Jun-N-terminal kinase, and extracellular signal-regulated kinase (ERK) is involved in this process. In addition, adenosine–uridine-rich elements (AREs) of the CXCL8/IL-8 3′-untranslated region (3′-UTR) reduce CXCL8/IL-8 mRNA stability. To investigate whether CTGF regulates CXCL8/IL-8 gene expression at the posttranscriptional level, we transfected 293 cells with serial luciferase constructs containing different segments of the CXCL8/IL-8 3′-UTR and then stimulated the cells with CTGF. The results suggested that CTGF stabilized luciferase mRNA and increased luciferase activity by regulating the CXCL8/IL-8 3′-UTR. Moreover, the p38 MAPK pathway may contribute to CTGF-induced CXCL8/IL-8 mRNA stabilization.

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Abbreviations

CTGF:

Connective tissue growth factor

IL:

Interleukin

MAPK:

Mitogen-activated protein kinase

JNK:

c-Jun-NH2-terminal kinase

p38 MAPK:

p38 mitogen-activated protein kinase

ERK:

Extracellular regulating kinase

NF-κB:

Nuclear factor κB

AP-1:

Activator protein-1

3′-UTR:

Three prime untranslated region

ARE:

Adenosine–uridine-rich element

References

  1. Ward PA, Hunninghake GW. Lung inflammation and fibrosis. Am J Respir Crit Care Med. 1998;157(4 Pt 2):S123–9.

    Article  CAS  PubMed  Google Scholar 

  2. Nagashima T, et al. Connective tissue growth factor is required for normal follicle development and ovulation. Mol Endocrinol. 2011;25(10):1740–59.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Kubota S, Takigawa M. The role of CCN2 in cartilage and bone development. J Cell Commun Signal. 2011;5(3):209–17.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Baguma-Nibasheka M, Kablar B. Pulmonary hypoplasia in the connective tissue growth factor (Ctgf) null mouse. Dev Dyn. 2008;237(2):485–93.

    Article  CAS  PubMed  Google Scholar 

  5. Hall-Glenn F, Lyons KM. Roles for CCN2 in normal physiological processes. Cell Mol Life Sci. 2011;68(19):3209–17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Morrison BL, Jose CC, Cutler ML. Connective Tissue Growth Factor (CTGF/CCN2) enhances lactogenic differentiation of mammary epithelial cells via integrin-mediated cell adhesion. BMC Cell Biol. 2010;11:35.

    Article  PubMed  PubMed Central  Google Scholar 

  7. Shimo T, et al. Expression and roles of CCN2 in dental epithelial cells. Vivo. 2015;29(2):189–95.

    CAS  Google Scholar 

  8. Kubota S, Takigawa M. Cellular and molecular actions of CCN2/CTGF and its role under physiological and pathological conditions. Clin Sci (Lond). 2015;128(3):181–96.

    Article  CAS  Google Scholar 

  9. Ivkovic S, et al. Connective tissue growth factor coordinates chondrogenesis and angiogenesis during skeletal development. Development. 2003;130(12):2779–91.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Moussad EE, Brigstock DR. Connective tissue growth factor: what’s in a name? Mol Genet Metab. 2000;71(1–2):276–92.

    Article  CAS  PubMed  Google Scholar 

  11. Blom IE, et al. In vitro evidence for differential involvement of CTGF, TGFbeta, and PDGF-BB in mesangial response to injury. Nephrol Dial Transplant. 2001;16(6):1139–48.

    Article  CAS  PubMed  Google Scholar 

  12. Dammeier J, et al. Connective tissue growth factor: a novel regulator of mucosal repair and fibrosis in inflammatory bowel disease? Int J Biochem Cell Biol. 1998;30(8):909–22.

    Article  CAS  PubMed  Google Scholar 

  13. Mori T, et al. Role and interaction of connective tissue growth factor with transforming growth factor-beta in persistent fibrosis: a mouse fibrosis model. J Cell Physiol. 1999;181(1):153–9.

    Article  CAS  PubMed  Google Scholar 

  14. Sato S, et al. Serum levels of connective tissue growth factor are elevated in patients with systemic sclerosis: association with extent of skin sclerosis and severity of pulmonary fibrosis. J Rheumatol. 2000;27(1):149–54.

    CAS  PubMed  Google Scholar 

  15. Stratton R, et al. Iloprost suppresses connective tissue growth factor production in fibroblasts and in the skin of scleroderma patients. J Clin Invest. 2001;108(2):241–50.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Ponticos M, et al. Pivotal role of connective tissue growth factor in lung fibrosis: MAPK-dependent transcriptional activation of type I collagen. Arthritis Rheum. 2009;60(7):2142–55.

    Article  CAS  PubMed  Google Scholar 

  17. Wang X, et al. A novel single-chain-Fv antibody against connective tissue growth factor attenuates bleomycin-induced pulmonary fibrosis in mice. Respirology. 2011;16(3):500–7.

    Article  PubMed  Google Scholar 

  18. Yang J, et al. Activated alveolar epithelial cells initiate fibrosis through autocrine and paracrine secretion of connective tissue growth factor. Am J Physiol Lung Cell Mol Physiol. 2014;306(8):L786–96.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Yokoi H, et al. Role of connective tissue growth factor in fibronectin expression and tubulointerstitial fibrosis. Am J Physiol Renal Physiol. 2002;282(5):F933–42.

    Article  CAS  PubMed  Google Scholar 

  20. Mukaida N. Interleukin-8: an expanding universe beyond neutrophil chemotaxis and activation. Int J Hematol. 2000;72(4):391–8.

    CAS  PubMed  Google Scholar 

  21. Mukaida N. Pathophysiological roles of interleukin-8/CXCL8 in pulmonary diseases. Am J Physiol Lung Cell Mol Physiol. 2003;284(4):L566–77.

    Article  CAS  PubMed  Google Scholar 

  22. Detmers PA, et al. Differential effects of neutrophil-activating peptide 1/IL-8 and its homologues on leukocyte adhesion and phagocytosis. J Immunol. 1991;147(12):4211–7.

    CAS  PubMed  Google Scholar 

  23. Baggiolini M, Clark-Lewis I. Interleukin-8, a chemotactic and inflammatory cytokine. FEBS Lett. 1992;307(1):97–101.

    Article  CAS  PubMed  Google Scholar 

  24. Scapini P, et al. The neutrophil as a cellular source of chemokines. Immunol Rev. 2000;177:195–203.

    Article  CAS  PubMed  Google Scholar 

  25. Ou XM, et al. Simvastatin attenuates bleomycin-induced pulmonary fibrosis in mice. Chin Med J (Engl). 2008;121(18):1821–9.

    CAS  Google Scholar 

  26. Kelly M, et al. Re-evaluation of fibrogenic cytokines in lung fibrosis. Curr Pharm Des. 2003;9(1):39–49.

    Article  CAS  PubMed  Google Scholar 

  27. Das MK, et al. Connective tissue growth factor induces tube formation and IL-8 production in first trimester human placental trophoblast cells. Eur J Obstet Gynecol Reprod Biol. 2014;181:183–8.

    Article  CAS  PubMed  Google Scholar 

  28. Seher A, et al. Gene expression profiling of connective tissue growth factor (CTGF) stimulated primary human tenon fibroblasts reveals an inflammatory and wound healing response in vitro. Mol Vis. 2011;17:53–62.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Wang X, et al. Regulation of pro-inflammatory and pro-fibrotic factors by CCN2/CTGF in H9c2 cardiomyocytes. J Cell Commun Signal. 2010;4(1):15–23.

    Article  PubMed  PubMed Central  Google Scholar 

  30. Wang Z, et al. Connective tissue growth factor promotes interleukin-1beta-mediated synovial inflammation in knee osteoarthritis. Mol Med Rep. 2013;8(3):877–82.

    CAS  PubMed  Google Scholar 

  31. Yosimichi G, et al. Roles of PKC, PI3K and JNK in multiple transduction of CCN2/CTGF signals in chondrocytes. Bone. 2006;38(6):853–63.

    Article  CAS  PubMed  Google Scholar 

  32. Smale ST, Fisher AG. Chromatin structure and gene regulation in the immune system. Annu Rev Immunol. 2002;20:427–62.

    Article  CAS  PubMed  Google Scholar 

  33. Tsuchiyama J, Mori M, Okada S. Murine spleen stromal cell line SPY3-2 maintains long-term hematopoiesis in vitro. Blood. 1995;85(11):3107–16.

    CAS  PubMed  Google Scholar 

  34. Yu CC, et al. Thrombin-induced connective tissue growth factor expression in human lung fibroblasts requires the ASK1/JNK/AP-1 pathway. J Immunol. 2009;182(12):7916–27.

    Article  CAS  PubMed  Google Scholar 

  35. Baek YS, et al. Identification of novel transcriptional regulators involved in macrophage differentiation and activation in U937 cells. BMC Immunol. 2009;10:18.

    Article  PubMed  PubMed Central  Google Scholar 

  36. Hu X, et al. The role of ERK and JNK signaling in connective tissue growth factor induced extracellular matrix protein production and scar formation. Arch Dermatol Res. 2013;305(5):433–45.

    Article  CAS  PubMed  Google Scholar 

  37. Liu XC, et al. Role of ERK1/2 and PI3-K in the regulation of CTGF-induced ILK expression in HK-2 cells. Clin Chim Acta. 2007;382(1–2):89–94.

    Article  CAS  PubMed  Google Scholar 

  38. Nagai N et al. CTGF is increased in basal deposits and regulates matrix production through the ERK (p42/p44mapk) MAPK and the p38 MAPK signaling pathways. Invest Ophthalmol Vis Sci. 2008.

  39. Shi L, et al. Activation of JNK signaling mediates connective tissue growth factor expression and scar formation in corneal wound healing. PLoS ONE. 2012;7(2):e32128.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Auwerx J. The human leukemia cell line, THP-1: a multifacetted model for the study of monocyte–macrophage differentiation. Experientia. 1991;47(1):22–31.

    Article  CAS  PubMed  Google Scholar 

  41. Wang XM, et al. Caveolin-1: a critical regulator of lung fibrosis in idiopathic pulmonary fibrosis. J Exp Med. 2006;203(13):2895–906.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Yoshida K, et al. MAP kinase activation and apoptosis in lung tissues from patients with idiopathic pulmonary fibrosis. J Pathol. 2002;198(3):388–96.

    Article  CAS  PubMed  Google Scholar 

  43. Alcorn JF, et al. c-Jun N-terminal kinase 1 is required for the development of pulmonary fibrosis. Am J Respir Cell Mol Biol. 2009;40(4):422–32.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Noble PW, Barkauskas CE, Jiang D. Pulmonary fibrosis: patterns and perpetrators. J Clin Invest. 2012;122(8):2756–62.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Chen Y, et al. Matrix contraction by dermal fibroblasts requires transforming growth factor-beta/activin-linked kinase 5, heparan sulfate-containing proteoglycans, and MEK/ERK: insights into pathological scarring in chronic fibrotic disease. Am J Pathol. 2005;167(6):1699–711.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Leask A. Potential therapeutic targets for cardiac fibrosis: TGFbeta, angiotensin, endothelin, CCN2, and PDGF, partners in fibroblast activation. Circ Res. 2010;106(11):1675–80.

    Article  CAS  PubMed  Google Scholar 

  47. Mott GA, Costales JA, Burleigh BA. A soluble factor from Trypanosoma cruzi inhibits transforming growth factor-ss-induced MAP kinase activation and gene expression in dermal fibroblasts. PLoS ONE. 2011;6(9):e23482.

    Article  PubMed  PubMed Central  Google Scholar 

  48. Pannu J, et al. Transforming growth factor-beta receptor type I-dependent fibrogenic gene program is mediated via activation of Smad1 and ERK1/2 pathways. J Biol Chem. 2007;282(14):10405–13.

    Article  CAS  PubMed  Google Scholar 

  49. Galuppo M, et al. MEK inhibition suppresses the development of lung fibrosis in the bleomycin model. Naunyn Schmiedebergs Arch Pharmacol. 2011;384(1):21–37.

    Article  CAS  PubMed  Google Scholar 

  50. Sonnylal S, et al. Selective expression of connective tissue growth factor in fibroblasts in vivo promotes systemic tissue fibrosis. Arthritis Rheum. 2010;62(5):1523–32.

    Article  PubMed  Google Scholar 

  51. Nakerakanti SS, Bujor AM, Trojanowska M. CCN2 is required for the TGF-beta induced activation of Smad1-Erk1/2 signaling network. PLoS ONE. 2011;6(7):e21911.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Leask A. MEK/ERK inhibitors: proof-of-concept studies in lung fibrosis. J Cell Commun Signal. 2012;6(1):59–60.

    Article  PubMed  PubMed Central  Google Scholar 

  53. Yong HY, Koh MS, Moon A. The p38 MAPK inhibitors for the treatment of inflammatory diseases and cancer. Expert Opin Investig Drugs. 2009;18(12):1893–905.

    Article  CAS  PubMed  Google Scholar 

  54. Mercer BA, D’Armiento JM. Emerging role of MAP kinase pathways as therapeutic targets in COPD. Int J Chron Obstruct Pulmon Dis. 2006;1(2):137–50.

    CAS  PubMed  PubMed Central  Google Scholar 

  55. Chopra P, et al. Therapeutic potential of inhaled p38 mitogen-activated protein kinase inhibitors for inflammatory pulmonary diseases. Expert Opin Investig Drugs. 2008;17(10):1411–25.

    Article  CAS  PubMed  Google Scholar 

  56. Renda T, et al. Increased activation of p38 MAPK in COPD. Eur Respir J. 2008;31(1):62–9.

    Article  CAS  PubMed  Google Scholar 

  57. Mukaida N, Mahe Y, Matsushima K. Cooperative interaction of nuclear factor-kappa B- and cis-regulatory enhancer binding protein-like factor binding elements in activating the interleukin-8 gene by pro-inflammatory cytokines. J Biol Chem. 1990;265(34):21128–33.

    CAS  PubMed  Google Scholar 

  58. Mukaida N, et al. Molecular mechanism of interleukin-8 gene expression. J Leukoc Biol. 1994;56(5):554–8.

    CAS  PubMed  Google Scholar 

  59. Holtmann H, et al. Induction of interleukin-8 synthesis integrates effects on transcription and mRNA degradation from at least three different cytokine- or stress-activated signal transduction pathways. Mol Cell Biol. 1999;19(10):6742–53.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Thompson C, et al. Signaling by the cysteinyl-leukotriene receptor 2. Involvement in chemokine gene transcription. J Biol Chem. 2008;283(4):1974–84.

    Article  CAS  PubMed  Google Scholar 

  61. Spiegelman VS, et al. Induction of beta-transducin repeat-containing protein by JNK signaling and its role in the activation of NF-kappaB. J Biol Chem. 2001;276(29):27152–8.

    Article  CAS  PubMed  Google Scholar 

  62. Lin CW, et al. 12-O-tetradecanoylphorbol-13-acetate-induced invasion/migration of glioblastoma cells through activating PKCalpha/ERK/NF-kappaB-dependent MMP-9 expression. J Cell Physiol. 2010;225(2):472–81.

    Article  CAS  PubMed  Google Scholar 

  63. Matoba K, et al. Rho-kinase regulation of TNF-alpha-induced nuclear translocation of NF-kappaB RelA/p65 and M-CSF expression via p38 MAPK in mesangial cells. Am J Physiol Renal Physiol. 2014;307(5):F571–80.

    Article  CAS  PubMed  Google Scholar 

  64. Rahman A, et al. cAMP targeting of p38 MAP kinase inhibits thrombin-induced NF-kappaB activation and ICAM-1 expression in endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2004;287(5):L1017–24.

    Article  CAS  PubMed  Google Scholar 

  65. Trachootham D, et al. Redox regulation of cell survival. Antioxid Redox Signal. 2008;10(8):1343–74.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Liu S, et al. Oxidative stress and MAPK involved into ATF2 expression in immortalized human urothelial cells treated by arsenic. Arch Toxicol. 2013;87(6):981–9.

    Article  CAS  PubMed  Google Scholar 

  67. Monje P, et al. Regulation of the transcriptional activity of c-Fos by ERK. A novel role for the prolyl isomerase PIN1. J Biol Chem. 2005;280(42):35081–4.

    Article  CAS  PubMed  Google Scholar 

  68. Chalmers CJ, et al. The duration of ERK1/2 activity determines the activation of c-Fos and Fra-1 and the composition and quantitative transcriptional output of AP-1. Cell Signal. 2007;19(4):695–704.

    Article  CAS  PubMed  Google Scholar 

  69. Yokoyama K, et al. C-Fos regulation by the MAPK and PKC pathways in intervertebral disc cells. PLoS ONE. 2013;8(9):e73210.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  70. Winzen R, et al. The p38 MAP kinase pathway signals for cytokine-induced mRNA stabilization via MAP kinase-activated protein kinase 2 and an AU-rich region-targeted mechanism. EMBO J. 1999;18(18):4969–80.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Wang S, et al. Nitric oxide-p38 MAPK signaling stabilizes mRNA through AU-rich element-dependent and -independent mechanisms. J Leukoc Biol. 2008;83(4):982–90.

    Article  CAS  PubMed  Google Scholar 

  72. Dean JL, et al. p38 Mitogen-activated protein kinase stabilizes mRNAs that contain cyclooxygenase-2 and tumor necrosis factor AU-rich elements by inhibiting deadenylation. J Biol Chem. 2003;278(41):39470–6.

    Article  CAS  PubMed  Google Scholar 

  73. Pages G, et al. Stress-activated protein kinases (JNK and p38/HOG) are essential for vascular endothelial growth factor mRNA stability. J Biol Chem. 2000;275(34):26484–91.

    Article  CAS  PubMed  Google Scholar 

  74. Ming XF, Kaiser M, Moroni C. c-Jun N-terminal kinase is involved in AUUUA-mediated interleukin-3 mRNA turnover in mast cells. EMBO J. 1998;17(20):6039–48.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  75. Chaudhary LR, Avioli LV. Regulation of interleukin-8 gene expression by interleukin-1beta, osteotropic hormones, and protein kinase inhibitors in normal human bone marrow stromal cells. J Biol Chem. 1996;271(28):16591–6.

    Article  CAS  PubMed  Google Scholar 

  76. Tebo J, et al. Heterogeneity in control of mRNA stability by AU-rich elements. J Biol Chem. 2003;278(14):12085–93.

    Article  CAS  PubMed  Google Scholar 

  77. Winzen R, et al. Functional analysis of KSRP interaction with the AU-rich element of interleukin-8 and identification of inflammatory mRNA targets. Mol Cell Biol. 2007;27(23):8388–400.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgments

This work was supported by grants from the National Science Council (NSC94-2320-B-038-009, NSC95-2320-B-038-001, NSC96-2320-B-038-007, and NSC96-2320-B-038-030-MY3). The authors would like to thank Dr. N. Mukaida for serial CXCL8/IL-8 promoter constructs in a luciferase reporter, Drs. Robert L. Danner and Shuibang Wang for generously providing pGL3-IL-8 3′-UTR reporter constructs, Dr. Ming-Liang Kuo for CTGF cDNA construct, and Meng-Pei Wu and Yu-Ching Lin for research assistance.

Conflict of interest

The authors declare that there is no conflict of interests regarding the publication of this paper.

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Authors and Affiliations

  1. Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan

    Chien-Huang Lin, Yu-Wen Chen, Yu-Liang Lin & Mei-Chieh Chen

  2. Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan

    Yuan-Hung Wang

  3. Department of Medical Research, Shuang Ho Hospital, Taipei Medical University, New Taipei City, Taiwan

    Yuan-Hung Wang

  4. School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan

    Bing-Chang Chen

  5. Department of Microbiology and Immunology, School of Medicine, College of Medicine, Taipei Medical University, No. 250 Wu-Hsing Street, Taipei, 110, Taiwan

    Mei-Chieh Chen

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  1. Chien-Huang Lin
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Lin, CH., Wang, YH., Chen, YW. et al. Transcriptional and posttranscriptional regulation of CXCL8/IL-8 gene expression induced by connective tissue growth factor. Immunol Res 64, 369–384 (2016). https://doi.org/10.1007/s12026-015-8670-0

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