Tumor Biology

, Volume 36, Issue 12, pp 9723–9732 | Cite as

Knockdown of PLCε inhibits inflammatory cytokine release via STAT3 phosphorylation in human bladder cancer cells

  • Xue Yang
  • Liping Ou
  • Min Tang
  • Yin Wang
  • Xiaorong Wang
  • E Chen
  • Jianjun Diao
  • Xiaohou Wu
  • Chunli Luo
Research Article

Abstract

Phospholipase Cε (PLCε) is a multifunctional enzyme implicated in inflammatory functions. There are limited data, however, on how PLCε can alter inflammatory cytokine by affecting downstream pathways. Recent studies suggest that inflammation is likely to have an important role in transitional cell carcinoma of bladder (TCCB) and cancer disease progression. Here, we showed that PLCε and p-STAT3 expression were both elevated in TCCB tissues compared to adjacent tissues, and the increase of PLCε level was associated with the increase of p-STAT3 level. Then, knockdown of PLCε using adenovirus-shPLCε significantly decreased inflammatory cytokine (IL-6, TNF-α, IL-1β) expression and inflammation-associated gene (TLR4, MyD88, p-STAT3) expression. Furthermore, we demonstrated that PLCε knockdown blocked LPS-induced inflammatory cytokine and p-STAT3 expression. Additionally, we found that combined treatment of STAT3 inhibitor S3I-201 with adenovirus-shPLCε exhibited synergistic inhibitory effects on expression of p-STAT3. Our results suggested that STAT3 phosphorylation is involved in PLCε-mediated inflammatory cytokine release. Our research is of potential importance in drug development programs using PLCε as a therapeutic target for TCCB.

Keywords

TCCB PLCε STAT3 IL-6 TNF-α IL-1β 

Notes

Acknowledgements

This work was supported by the National Natural Science Foundation of China (Grant No. 81072086) and Scientific and Technological Research Program of Chongqing Municipal Education Committee of China (Grant No. KJ110305).

Conflict of interest

None

References

  1. 1.
    Rhee SG. Regulation of phosphoinositide-specific phospholipase C. Annu Rev Biochem. 2001;70:281–312.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Ada-Nguema AS, Xenias H, Hofman JM, Wiggins CH, Sheetz MP, Keely PJ. The small GTPase R-Ras regulates organization of actin and drives membrane protrusions through the activity of PLCepsilon. J Cell Sci. 2006;119:1307–19.CrossRefPubMedGoogle Scholar
  3. 3.
    Edamatsu H, Satoh T, Kataoka T. Ras and Rap1 activation of PLCepsilon lipase activity. Method Enzymol. 2006;407:99–107.CrossRefGoogle Scholar
  4. 4.
    Wing MR, Snyder JT, Sondek J, Harden TK. Direct activation of phospholipase C-epsilon by Rho. J Biol Chem. 2003;278:41253–8.CrossRefPubMedGoogle Scholar
  5. 5.
    Li M, Edamatsu H, Kitazawa R, Kitazawa S, Kataoka T. Phospholipase Cepsilon promotes intestinal tumorigenesis of Apc(Min/+) mice through augmentation of inflammation and angiogenesis. Carcinogenesis. 2009;30:1424–32.CrossRefPubMedGoogle Scholar
  6. 6.
    Harada Y, Edamatsu H, Kataoka T. PLCε cooperates with the NF-κB pathway to augment TNFα-stimulated CCL2/MCP1 expression in human keratinocyte. Biochem Biophys Res Commun. 2011;414:106–11.CrossRefPubMedGoogle Scholar
  7. 7.
    Oka M, Edamatsu H, Kunisada M, Hu L, Takenaka N, Sakaguchi M, et al. Phospholipase Cε has a crucial role in ultraviolet B-induced neutrophil-associated skin inflammation by regulating the expression of CXCL1/KC. Lab Invest. 2011;91:711–8.CrossRefPubMedGoogle Scholar
  8. 8.
    Siegel R, Ma J, Zou Z, Jemal A. Cancer statistics, 2014. CA Cancer J Clin. 2014;64:9–29.CrossRefPubMedGoogle Scholar
  9. 9.
    Zhang Y, Yan L, Zhao Y, Ou L, Wu X, Luo C. Knockdown of phospholipase C-epsilon by short-hairpin RNA-mediated gene silencing induces apoptosis in human bladder cancer cell lines. Cancer Biother Radiopharm. 2013;28(3):233–9.CrossRefPubMedGoogle Scholar
  10. 10.
    Ling Y, Chunli L, Xiaohou W, Qiaoling Z. Involvement of the PLCε/PKCα pathway in human BIU-87 bladder cancer cell proliferation. Cell Biol Int. 2011;35(10):1031–6.CrossRefPubMedGoogle Scholar
  11. 11.
    Du HF, Ou LP, Yang X, Song XD, Fan YR, Tan B, et al. A new PKCα/β/TBX3/E-cadherin pathway is involved in PLCε-regulated invasion and migration in human bladder cancer cells. Cell Signal. 2014;26:580–93.CrossRefPubMedGoogle Scholar
  12. 12.
    Janeway Jr CA, Medzhitov R. Innate immune recognition. Annu Rev Immunol. 2002;20:197–216.CrossRefPubMedGoogle Scholar
  13. 13.
    LaRue H, Ayari C, Bergeron A, Fradet Y. Toll-like receptors in urothelial cells—targets for cancer immunotherapy. Nat Rev Urol. 2013;10:537–45.PubMedGoogle Scholar
  14. 14.
    Kim HM, Park BS, Kim JI, Kim SE, Lee J, Oh SC, et al. Crystal structure of the TLR4-MD-2 complex with bound endotoxin antagonist Eritoran. Cell. 2007;130:906–17.CrossRefPubMedGoogle Scholar
  15. 15.
    Park BS, Song DH, Kim HM, Choi BS, Lee H, Lee JO. The structural basis of lipopolysaccharide recognition by the TLR4-MD-2 complex. Nature. 2009;458:1191–5.CrossRefPubMedGoogle Scholar
  16. 16.
    Lu YC, Yeh WC, Ohashi PS. LPS/TLR4 signal transduction pathway. Cytokine+. 2008;42:145–51.PubMedGoogle Scholar
  17. 17.
    Qian Y, Deng J, Xie H, Geng L, Zhou L, Wang Y, et al. Regulation of TLR4-induced IL-6 response in bladder cancer cells by opposing actions of MAPK and PI3K signaling. J Cancer Res Clin Oncol. 2009;135:379–86.CrossRefPubMedGoogle Scholar
  18. 18.
    Ferguson SD, Srinivasan VM, Heimberger AB. The role of STAT3 in tumor-mediated immune suppression. J Neurooncol. 2015;22.Google Scholar
  19. 19.
    Bromberg JF, Wrzeszczynska MH, Devgan G, Zhao Y, Pestell RG, Albanese C, et al. STAT3 as an oncogene. Cell. 1999;98:295–303.CrossRefPubMedGoogle Scholar
  20. 20.
    Kishimoto T. IL-6: from its discovery to clinical applications. Int Immunol. 2010;22:347–52.CrossRefPubMedGoogle Scholar
  21. 21.
    Yu H, Pardoll D, Jove R. STATs in cancer inflammation and immunity: a leading role for STAT3. Nat Rev Cancer. 2009;9:798–809.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Lee H, Deng J, Kujawski M, Yang C, Liu Y, Herrmann A, et al. STAT3-induced S1PR1 expression is crucial for persistent STAT3 activation in tumors. Nat Med. 2010;16:1421–8.CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Samavati L, Rastogi R, Du W, Hüttemann M, Fite A, Franchi L. STAT3 tyrosine phosphorylation is critical for interleukin 1 beta and interleukin-6 production in response to lipopolysaccharide and live bacteria. Mol Immunol. 2009;46:1867–77.CrossRefPubMedGoogle Scholar
  24. 24.
    Akira S, Uematsu S, Takeuchi O. Pathogen recognition and innate immunity. Cell. 2006;124:783–801.CrossRefPubMedGoogle Scholar
  25. 25.
    Szajnik M, Szczepanski MJ, Czystowska M, Elishaev E, Mandapathil M, Nowak-Markwitz E, et al. TLR4 signaling induced by lipopolysaccharide or paclitaxel regulates tumor survival and chemoresistance in ovarian cancer. Oncogene. 2009;28:4353–63.CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Hua D, Liu MY, Cheng ZD, Qin XJ, Zhang HM, Chen Y, et al. Small interfering RNA-directed targeting of Toll-like receptor 4 inhibits human prostate cancer cell invasion, survival, and tumorigenicity. Mol Immunol. 2009;46:2876–84.CrossRefPubMedGoogle Scholar
  27. 27.
    Szczepanski MJ, Czystowska M, Szajnik M, Harasymczuk M, Boyiadzis M, Kruk-Zagajewska A, et al. Triggering of Toll-like receptor 4 expressed on human head and neck squamous cell carcinoma promotes tumor development and protects the tumor from immune attack. Cancer Res. 2009;69:3105–13.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Ying H, Da L, Yu-xiu S, Yu X, Li-xia L, Li-mei X, et al. TLR4 mediates MAPK–STAT3 axis activation in bladder epithelial cells. Inflammation. 2013;36:1064–74.CrossRefPubMedGoogle Scholar
  29. 29.
    Ikuta S, Edamatsu H, Li M, Hu L, Kataoka T. Crucial role of phospholipase C epsilon in skin inflammation induced by tumor-promoting phorbol ester. Cancer Res. 2008;68:64–72.CrossRefPubMedGoogle Scholar
  30. 30.
    Hu L, Edamatsu H, Takenaka N, Ikuta S, Kataoka T. Crucial role of phospholipase Cepsilon in induction of local skin inflammatory reactions in the elicitation stage of allergic contact hypersensitivity. J Immunol. 2010;184:993–1002.CrossRefPubMedGoogle Scholar
  31. 31.
    Dusaban SS, Purcell NH, Rockenstein E, Masliah E, Cho MK, Smrcka AV, et al. Phospholipase Cε links G protein-coupled receptor activation to inflammatory astrocytic responses. Proc Natl Acad Sci U S A. 2013;110:3609–14.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011;144:646–74.CrossRefPubMedGoogle Scholar
  33. 33.
    Grivennikov SI, Greten FR, Karin M. Immunity, inflammation, and cancer. Cell. 2010;140:883–99.CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Cai T, Nesi G, Boddi V, Mazzoli S, Dal Canto M, Bartoletti R. Prognostic role of the tumor-associated tissue inflammatory reaction in transitional bladder cell carcinoma. Oncol Rep. 2006;16:329–34.PubMedGoogle Scholar
  35. 35.
    Michaud DS. Chronic inflammation and bladder cancer. Urol Oncol. 2007;25:260–8.CrossRefPubMedGoogle Scholar
  36. 36.
    Kagan JC, Su T, Horng T, Chow A, Akira S, Medzhitov R. TRAM couples endocytosis of Toll-like receptor 4 to the induction of interferon-beta. Nat Immunol. 2008;9:361–8.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Tanimura N, Saitoh S, Matsumoto F, Akashi-Takamura S, Miyake K. Roles for LPS-dependent interaction and relocation of TLR4 and TRAM in TRIF-signaling. Biochem Biophys Res Commun. 2008;368:94–9.CrossRefPubMedGoogle Scholar
  38. 38.
    Zanoni I, Ostuni R, Marek LR, Barresi S, Barbalat R, Barton GM, et al. CD14 controls the LPS-induced endocytosis of Toll-like receptor 4. Cell. 2011;147:868–80.CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Nagano T, Edamatsu H, Kobayashi K, Takenaka N, Yamamoto M, Sasaki N, et al. Phospholipase cε, an effector of ras and rap small GTPases, is required for airway inflammatory response in a mouse model of bronchial asthma. PLoS One. 2014;9, e108373.CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Jarnicki A, Putoczki T, Ernst M. STAT3: linking inflammation to epithelial cancer-more than a “gut” feeling? Cell Div. 2010;5:14.CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Garner JM, Fan M, Yang CH, Du Z, Sims M, Davidoff AM, et al. Constitutive activation of signal transducer and activator of transcription 3 (STAT3) and nuclear factor κB signaling in glioblastoma cancer stem cells regulates the Notch pathway. J Biol Chem. 2013;288:26167–76.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  • Xue Yang
    • 1
  • Liping Ou
    • 1
  • Min Tang
    • 1
  • Yin Wang
    • 1
  • Xiaorong Wang
    • 1
  • E Chen
    • 1
  • Jianjun Diao
    • 2
  • Xiaohou Wu
    • 2
  • Chunli Luo
    • 1
  1. 1.The Key Laboratory of Diagnostics Medicine designated by the Ministry of EducationChongqing Medical UniversityChongqingPeople’s Republic of China
  2. 2.Department of Urinary SurgeryThe First Affiliated Hospital of Chongqing Medical UniversityChongqingPeople’s Republic of China

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