Abstract
Background
Renal interstitial fibrosis (RIF) is a pathological change common to a variety of chronic renal diseases, ultimately progressing to end-stage renal failure. It is believed that epithelial cell phenotype inversion plays an important role in RIF, which is characterized by expression of the mesenchymal maker α-SMA, loss of the epithelial maker E-cadherin, and enhanced secretion of extracellular matrix. IL-17, a newly discovered pro-inflammatory cytokine, has recently been reported to play an important role in tissue fibrosis, involving pulmonary, liver, intestine and skin tissues. This study aimed to investigate whether IL-17A, a member of the IL-17 family, can induce epithelial cell phenotype inversion, and to explore the molecular mechanism of this phenotype inversion, in vitro.
Methods
HK-2 cells were cultured and incubated with IL-17A. Cell proliferation was measured by CCK-8 assay, and the secretion of types I and III collagen was detected by ELISA in dose-dependent and time-dependent experiments. To find out whether IL-17A can induce epithelial cell phenotype inversion, HK-2 cells were stimulated with 80 ng/mL of IL-17A and 10 ng/mL of TGF-β1 as a positive control, for 72 h. To explore the potential signaling pathway, anti-TGF-β1 antibody was added before IL-17A treatment. At the same time, anti-TGF-β1 antibody alone was added to the medium as the negative control group. The expression of types I and III collagen, α-SMA and E-cadherin proteins, and mRNA was measured by real-time PCR, western blotting and immuno-histochemistry.
Results
IL-17A promoted the proliferation of HK-2 cells and secretion of types I and III collagen in a dose-dependent and time-dependent manner. Compared with the normal control, IL-17A could stimulate the expression of α-SMA, types I and III collagen, and suppressed the expression of E-cadherin in HK-2 cells. Incubation of IL-17A with TGF-β1 antibody decreased significantly the expression of α-SMA, but increased the expression of E-cadherin in HK-2 cells.
Conclusion
Our results suggest that IL-17A might promote the proliferation of HK-2 cells and secretion of extracellular matrix, and induce epithelial cell phenotype inversion via a TGF-β1-dependent pathway. Blocking the pro-inflammatory cytokine IL-17A might be a potential target for the treatment of fibrotic kidney disease.
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References
Eddy AA. Molecular basis of renal fibrosis. Pediatr Nephrol 2000; 15: 290–301.
Razzaque MS, Taguchi T. Cellular and molecular events leading to renal tubulointerstitial fibrosis. Med Electron Microsc 2002; 35: 68–80.
Okada H, Kalluri R. Cellular and molecular pathways that lead to progression and regression of renal fibrogenesis. Curr Mol Med 2005; 5: 467–74.
Yang J, Liu Y. Dissection of key events in tubular epithelial to myofibroblast transition and its implications in renal interstitial fibrosis. Am J Pathol 2001; 159: 1465–75.
Zeisberg M, Neilson EG. Mechanisms of tubulointerstitial fibrosis. J Am Soc Nephrol 2010; 21: 1819–34.
Cho ME, Smith DC, Branton MH, Penzak SR, Kopp JB. Pirfenidone slows renal function decline in patients with focal segmental glomerulosclerosis. Clin J Am Soc Nephrol 2007; 2: 906–13.
Conway B, Hughes J. Cellular orchestrators of renal fibrosis. QJM 2012; 105: 611–5.
Boor P, Floege J. The renal (myo-)fibroblast: a heterogeneous group of cells. Nephrol Dial Transplant 2012; 27: 3027–36.
Badid C, Mounier N, Costa AM, Desmouliere A. Role of myofibroblasts during normal tissue repair and excessive scarring: interest of their assessment in nephropathies. Histol Histopathol 2000; 15: 269–80.
Ohtake Y, Tojo H, Seiki M. Multifunctional roles of MT1-MMP in myofiber formation and morphostatic maintenance of skeletal muscle. J Cell Sci 2006; 119: 3822–32.
Lan HY. Tubular epithelial-myofibroblast transdifferentiation mechanisms in proximal tubule cells. Curr Opin Nephrol Hypertens 2003; 12: 25–9.
Zeisberg M, Kalluri R. The role of epithelial-to-mesenchymal transition in renal fibrosis. J Mol Med 2004; 82: 175–81.
Strutz F, Muller GA. Renal fibrosis and the origin of the renal fibroblast. Nephrol Dial Transplant 2006; 21: 3368–70.
Slattery C, Campbell E, McMorrow T, Ryan MP. Cyclosporine Ainduced renal fibrosis: a role for epithelial-mesenchymal transition. Am J Pathol 2005; 167: 395–407.
Lee JM, Dedhar S, Kalluri R, Thompson EW. The epithelialmesenchymal transition: newinsights in signaling, development, and disease. J Cell Biol 2006; 172: 973–81.
Harrington LE, Hatton RD, Mangan PR, et al. Interleukin 17-producing CD4+ effector T cells develop via a lineage distinct from the T helper type 1 and 2 lineages. Nat Immunol 2005; 6: 1123–32.
Park H, Li Z, Yang XO, et al. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat Immunol 2005; 6: 1133–41.
Aggarwal S, Gurney AL. IL-17: prototype member of an emerging cytokine family. J Leukoc Biol 2002; 71: 1–8.
Korn T, Oukka M, Kuchroo V, Bettelli E. Th17 cells: effector T cells with inflammatory properties. Semin Immunol 2007; 19: 362–71.
Kolls JK, Linden A. Interleukin-17 family members and inflammation. Immunity 2004; 21: 467–76.
Manotham K, Tanaka T, Matsumoto M, et al. Transdifferentiation of cultured tubular cells induced by hypoxia. Kidney Int 2004; 65: 871–80.
Fan JM, Ng YY, Hill PA, et al. Transforming growth factor-beta regulates tubular epithelial-myofibroblast transdifferentiation in vitro. Kidney Int 1999; 56: 1455–67.
Dong Z, Yang Y, Zhang T, et al. siRNA-Act1 inhibits the function of IL-17 on lung fibroblasts via the NF-kappaB pathway. Respiration 2013; 86: 332–40.
Du WJ, Zhen JH, Zeng ZQ, et al. Expression of interleukin-17 associated with disease progression and liver fibrosis with hepatitis B virus infection: IL-17 in HBV infection. Diagn Pathol 2013; 8: 40.
Meng F, Wang K, Aoyama T, et al. Interleukin-17 signaling in inflammatory, Kupffer cells, and hepatic stellate cells exacerbates liver fibrosis in mice. Gastroenterology 2012; 143: 765–76, e1-3.
Biancheri P, Pender SL, Ammoscato F, et al. The role of interleukin 17 in Crohn’s disease-associated intestinal fibrosis. Fibrogenesis Tissue Repair 2013; 6: 13.
Okamoto Y, Hasegawa M, Matsushita T, et al. Potential roles of interleukin-17A in the development of skin fibrosis in mice. Arthritis Rheum 2012; 64: 3726–35.
Knight PG, Glister C. TGF-beta superfamily members and ovarian follicle development. Reproduction 2006; 132: 191–206.
Ruiz-Ortega M, Rodriguez-Vita J, Sanchez-Lopez E, Carvajal G, Egido J. TGF-beta signaling in vascular fibrosis. Cardiovasc Res 2007; 74: 196–206.
Wynn TA. Cellular and molecular mechanisms of fibrosis. J Pathol 2008; 214: 199–210.
Sato M, Muragaki Y, Saika S, Roberts AB, Ooshima A. Targeted disruption of TGF-beta1/Smad3 signaling protects against renal tubulo-interstitial fibrosis induced by unilateral ureteral obstruction. J Clin Invest 2003; 112: 1486–94.
Phanish MK, Wahab NA, Colville-Nash P, Hendry BM, Dockrell ME. The differential role of Smad2 and Smad3 in the regulation of pro-fibrotic TGFbeta1 responses in human proximal-tubule epithelial cells. Biochem J 2006; 393: 601–7.
Meng XM, Huang XR, Chung AC, et al. Smad2 protects against TGF-beta/Smad3-mediated renal fibrosis. J Am Soc Nephrol 2010; 21: 1477–87.
Li J, Qu X, Ricardo SD, Bertram JF, Nikolic-Paterson DJ. Resveratrol inhibits renal fibrosis in the obstructed kidney: potential role in deacetylation of Smad3. Am J Pathol 2010; 177: 1065–71.
Mi S, Li Z, Yang HZ, et al. Blocking IL-17A promotes the resolution of pulmonary inflammation and fibrosis via TGF-beta1-dependent and-independent mechanisms. J Immunol 2011; 187: 3003–14.
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Liu, L., Li, Fg., Yang, M. et al. Effect of pro-inflammatory interleukin-17A on epithelial cell phenotype inversion in HK-2 cells in vitro . Eur Cytokine Netw 27, 27–33 (2016). https://doi.org/10.1684/ecn.2016.0373
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DOI: https://doi.org/10.1684/ecn.2016.0373