Impaired gap junctions in human hepatocellular carcinoma limit intrinsic oxaliplatin chemosensitivity: A key role of connexin 26

  • Authors:
    • Yan Yang
    • Jian Zhu
    • Na Zhang
    • Yu Zhao
    • Wan-Yun Li
    • Fu-You Zhao
    • Yu-Rong Ou
    • Shu-Kui Qin
    • Qiong Wu
  • View Affiliations

  • Published online on: November 26, 2015     https://doi.org/10.3892/ijo.2015.3266
  • Pages: 703-713
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Abstract

Hepatocellular carcinoma (HCC) is generally believed to have low sensitivity to chemotherapeutic agents including oxaliplatin (OXA). Studies have demonstrated that gap junctions (GJs) composed of connexin (Cx) proteins have the potential to modulate drug chemosensitivity in multiple tumor cells. In the present study, we investigated the characteristics of Cx and GJs in HCC at both histologic and cytologic levels, and the effects of GJ and its effective components on OXA cytotoxicity in HCC cells in vitro. Immunohistochemistry was performed in 76 HCCs and 20 normal liver tissues to detect and locate the expression of Cx26, Cx32 and Cx43. At cytologic levels, the expression and localization of Cxs were evaluated by RT-PCR, western blot and immunofluorescence assay, respectively. The GJ function between adjacent cells was detected using dye transfer assay. The role of GJs in the modulation of OXA toxicity in HCC cells was explored using pharmacologic and molecular biologic methods. We found that Cx expression in HCC tissues was significantly lower than in normal liver tissues, and the ‘internalization’ from cell membrane to cytoplasm was remarkable. In vitro experiments revealed the presence of functional GJs in the SMMC-7721 HCC cells due to a small amount of Cx protein along the plasma membrane at cell-cell contacts. Regulation of this part of GJs positively influenced OXA cytotoxicity. Using RNA interference, only specific inhibition of Cx26 but not Cx32 or Cx43 reduced OXA cytotoxicity. Conversely, Cx26 overexpression by transfection of Cx26 plasmid DNA enhanced OXA cytotoxicity. This study demonstrated that during hepatocarcinogenesis, the reduced expression and internalization of Cx proteins impaired the GJ function, which further attenuated OXA cytotoxicity. Impaired GJ function may contribute to low intrinsic chemosensitivity of HCC cells to OXA, mediated by Cx26.

References

1 

Wu Q, Qin SK, Teng FM, Chen CJ and Wang R: Lobaplatin arrests cell cycle progression in human hepatocellular carcinoma cells. J Hematol Oncol. 3:432010. View Article : Google Scholar : PubMed/NCBI

2 

Zaanan A, Williet N, Hebbar M, Dabakuyo TS, Fartoux L, Mansourbakht T, Dubreuil O, Rosmorduc O, Cattan S, Bonnetain F, et al: Gemcitabine plus oxaliplatin in advanced hepatocellular carcinoma: A large multicenter AGEO study. J Hepatol. 58:81–88. 2013. View Article : Google Scholar

3 

Qin S, Bai Y, Lim HY, Thongprasert S, Chao Y, Fan J, Yang TS, Bhudhisawasdi V, Kang WK, Zhou Y, et al: Randomized, multicenter, open-label study of oxaliplatin plus fluorouracil/leucovorin versus doxorubicin as palliative chemotherapy in patients with advanced hepatocellular carcinoma from Asia. J Clin Oncol. 31:3501–3508. 2013. View Article : Google Scholar : PubMed/NCBI

4 

Petrelli F, Coinu A, Borgonovo K, Cabiddu M, Ghilardi M, Lonati V and Barni S: Oxaliplatin-based chemotherapy: A new option in advanced hepatocellular carcinoma. a systematic review and pooled analysis. Clin Oncol (R Coll Radiol). 26:488–496. 2014. View Article : Google Scholar

5 

Abdel-Rahman O: Revisiting oxaliplatin-based regimens for advanced hepatocellular carcinoma. Curr Oncol Rep. 16:3942014. View Article : Google Scholar : PubMed/NCBI

6 

Miao J, Chen GG, Chun SY, Chak EC and Lai PB: Bid sensitizes apoptosis induced by chemotherapeutic drugs in hepatocellular carcinoma. Int J Oncol. 25:651–659. 2004.PubMed/NCBI

7 

Vinken M, Vanhaecke T, Papeleu P, Snykers S, Henkens T and Rogiers V: Connexins and their channels in cell growth and cell death. Cell Signal. 18:592–600. 2006. View Article : Google Scholar

8 

Brockmeyer P, Jung K, Perske C, Schliephake H and Hemmerlein B: Membrane connexin 43 acts as an independent prognostic marker in oral squamous cell carcinoma. Int J Oncol. 45:273–281. 2014.PubMed/NCBI

9 

Krutovskikh VA, Piccoli C and Yamasaki H: Gap junction inter-cellular communication propagates cell death in cancerous cells. Oncogene. 21:1989–1999. 2002. View Article : Google Scholar : PubMed/NCBI

10 

Jensen R and Glazer PM: Cell-interdependent cisplatin killing by Ku/DNA-dependent protein kinase signaling transduced through gap junctions. Proc Natl Acad Sci USA. 101:6134–6139. 2004. View Article : Google Scholar : PubMed/NCBI

11 

Tanaka M and Grossman HB: Connexin 26 gene therapy of human bladder cancer: Induction of growth suppression, apoptosis, and synergy with Cisplatin. Hum Gene Ther. 12:2225–2236. 2001. View Article : Google Scholar

12 

Shishido SN and Nguyen TA: Gap junction enhancer increases efficacy of cisplatin to attenuate mammary tumor growth. PLoS One. 7:e449632012. View Article : Google Scholar : PubMed/NCBI

13 

Yoon SY, Robinson CR, Zhang H and Dougherty PM: Spinal astrocyte gap junctions contribute to oxaliplatin-induced mechanical hypersensitivity. J Pain. 14:205–214. 2013. View Article : Google Scholar : PubMed/NCBI

14 

Bai C, Yang M, Fan Z, Li S, Gao T and Fang Z: Associations of chemo- and radio-resistant phenotypes with the gap junction, adhesion and extracellular matrix in a three-dimensional culture model of soft sarcoma. J Exp Clin Cancer Res. 34:582015. View Article : Google Scholar : PubMed/NCBI

15 

Mesnil M: Connexins and cancer. Biol Cell. 94:493–500. 2002. View Article : Google Scholar

16 

Cronier L, Crespin S, Strale PO, Defamie N and Mesnil M: Gap junctions and cancer: New functions for an old story. Antioxid Redox Signal. 11:323–338. 2009. View Article : Google Scholar

17 

Vinken M, De Kock J, Oliveira AG, Menezes GB, Cogliati B, Dagli ML, Vanhaecke T and Rogiers V: Modifications in connexin expression in liver development and cancer. Cell Commun Adhes. 19:55–62. 2012. View Article : Google Scholar : PubMed/NCBI

18 

Zhang JT and Nicholson BJ: The topological structure of connexin 26 and its distribution compared to connexin 32 in hepatic gap junctions. J Membr Biol. 139:15–29. 1994. View Article : Google Scholar : PubMed/NCBI

19 

Maes M, Decrock E, Cogliati B, Oliveira AG, Marques PE, Dagli ML, Menezes GB, Mennecier G, Leybaert L, Vanhaecke T, et al: Connexin and pannexin (hemi)channels in the liver. Front Physiol. 4:4052014. View Article : Google Scholar : PubMed/NCBI

20 

Plante I, Charbonneau M and Cyr DG: Decreased gap junctional intercellular communication in hexachlorobenzene-induced gender-specific hepatic tumor formation in the rat. Carcinogenesis. 23:1243–1249. 2002. View Article : Google Scholar : PubMed/NCBI

21 

Ma XD, Sui YF and Wang WL: Expression of gap junction genes connexin 32, connexin 43 and their proteins in hepatocellular carcinoma and normal liver tissues. World J Gastroenterol. 6:66–69. 2000. View Article : Google Scholar

22 

Nakashima Y, Ono T, Yamanoi A, El-Assal ON, Kohno H and Nagasue N: Expression of gap junction protein connexin32 in chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. J Gastroenterol. 39:763–768. 2004. View Article : Google Scholar : PubMed/NCBI

23 

Bevans CG, Kordel M, Rhee SK and Harris AL: Isoform composition of connexin channels determines selectivity among second messengers and uncharged molecules. J Biol Chem. 273:2808–2816. 1998. View Article : Google Scholar : PubMed/NCBI

24 

Ayad WA, Locke D, Koreen IV and Harris AL: Heteromeric, but not homomeric, connexin channels are selectively permeable to inositol phosphates. J Biol Chem. 281:16727–16739. 2006. View Article : Google Scholar : PubMed/NCBI

25 

Zheng R, Wang J, Wu Q, Wang Z, Ou Y, Ma L, Wang M, Wang J and Yang Y: Expression of ALDH1 and TGFβ2 in benign and malignant breast tumors and their prognostic implications. Int J Clin Exp Pathol. 7:4173–4183. 2014.

26 

He XD, Wang Y, Wu Q, Wang HX, Chen ZD, Zheng RS, Wang ZS, Wang JB and Yang Y: Xuebijing protects rats from sepsis challenged with acinetobacter baumannii by promoting Annexin A1 expression and inhibiting proinflammatory cytokines secretion. Evid Based Complement Alternat Med. 2013:8049402013. View Article : Google Scholar : PubMed/NCBI

27 

Yang Y, Qin SK, Wu Q, Wang ZS, Zheng RS, Tong XH, Liu H, Tao L and He XD: Connexin-dependent gap junction enhancement is involved in the synergistic effect of sorafenib and all-trans retinoic acid on HCC growth inhibition. Oncol Rep. 31:540–550. 2014.

28 

Yang Y, Cao MH, Wang Q, Yuan DD, Li L and Tao L: The effects of 2-aminoethoxydiphenyl borate and diphenylboronic anhydride on gap junctions composed of Connexin43 in TM(4) sertoli cells. Biol Pharm Bull. 34:1390–1397. 2011. View Article : Google Scholar

29 

Eugenín EA, Eckardt D, Theis M, Willecke K, Bennett MV and Saez JC: Microglia at brain stab wounds express connexin 43 and in vitro form functional gap junctions after treatment with interferon-gamma and tumor necrosis factor-alpha. Proc Natl Acad Sci USA. 98:4190–4195. 2001. View Article : Google Scholar : PubMed/NCBI

30 

Garg S, Md Syed M and Kielian T: Staphylococcus aureus-derived peptidoglycan induces Cx43 expression and functional gap junction intercellular communication in microglia. J Neurochem. 95:475–483. 2005. View Article : Google Scholar : PubMed/NCBI

31 

Sáez CG, Velásquez L, Montoya M, Eugenín E and Alvarez MG: Increased gap junctional intercellular communication is directly related to the anti-tumor effect of all-trans-retinoic acid plus tamoxifen in a human mammary cancer cell line. J Cell Biochem. 89:450–461. 2003. View Article : Google Scholar : PubMed/NCBI

32 

Wang J, Dai Y, Huang Y, Chen X, Wang H, Hong Y, Xia J and Cheng B: All-trans retinoic acid restores gap junctional intercellular communication between oral cancer cells with upregulation of Cx32 and Cx43 expressions in vitro. Med Oral Patol Oral Cir Bucal. 18:e569–e577. 2013. View Article : Google Scholar : PubMed/NCBI

33 

Mesnil M, Crespin S, Avanzo JL and Zaidan-Dagli ML: Defective gap junctional intercellular communication in the carcinogenic process. Biochim Biophys Acta. 1719:125–145. 2005. View Article : Google Scholar : PubMed/NCBI

34 

Leithe E, Sirnes S, Omori Y and Rivedal E: Downregulation of gap junctions in cancer cells. Crit Rev Oncog. 12:225–256. 2006. View Article : Google Scholar

35 

Dejana E, Orsenigo F, Molendini C, Baluk P and McDonald DM: Organization and signaling of endothelial cell-to-cell junctions in various regions of the blood and lymphatic vascular trees. Cell Tissue Res. 335:17–25. 2009. View Article : Google Scholar

36 

Lampugnani MG and Dejana E: The control of endothelial cell functions by adherens junctions. Novartis Found Symp. 283:4–13; discussion 13–17, 238–241. 2007. View Article : Google Scholar

37 

Yano T, Hernandez-Blazquez FJ, Omori Y and Yamasaki H: Reduction of malignant phenotype of HEPG2 cell is associated with the expression of connexin 26 but not connexin 32. Carcinogenesis. 22:1593–1600. 2001. View Article : Google Scholar : PubMed/NCBI

38 

Yamasaki H and Naus CC: Role of connexin genes in growth control. Carcinogenesis. 17:1199–1213. 1996. View Article : Google Scholar : PubMed/NCBI

39 

Wang Z, Zhou J, Fan J, Qiu SJ, Yu Y, Huang XW, Sun J, Tan CJ and Dai Z: Oxaliplatin induces apoptosis in hepatocellular carcinoma cells and inhibits tumor growth. Expert Opin Investig Drugs. 18:1595–1604. 2009. View Article : Google Scholar : PubMed/NCBI

40 

Gao J, Wang R, Yang Q, Chen C and Wu Q: Effect of Oxaliplatin on cell cycle of hepatocellular carcinoma cell line HepG2. Zhejiang Da Xue Xue Bao Yi Xue Ban. 42:437–442. 2013.(In Chinese). PubMed/NCBI

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Copy and paste a formatted citation
APA
Yang, Y., Zhu, J., Zhang, N., Zhao, Y., Li, W., Zhao, F. ... Wu, Q. (2016). Impaired gap junctions in human hepatocellular carcinoma limit intrinsic oxaliplatin chemosensitivity: A key role of connexin 26. International Journal of Oncology, 48, 703-713. https://doi.org/10.3892/ijo.2015.3266
MLA
Yang, Y., Zhu, J., Zhang, N., Zhao, Y., Li, W., Zhao, F., Ou, Y., Qin, S., Wu, Q."Impaired gap junctions in human hepatocellular carcinoma limit intrinsic oxaliplatin chemosensitivity: A key role of connexin 26". International Journal of Oncology 48.2 (2016): 703-713.
Chicago
Yang, Y., Zhu, J., Zhang, N., Zhao, Y., Li, W., Zhao, F., Ou, Y., Qin, S., Wu, Q."Impaired gap junctions in human hepatocellular carcinoma limit intrinsic oxaliplatin chemosensitivity: A key role of connexin 26". International Journal of Oncology 48, no. 2 (2016): 703-713. https://doi.org/10.3892/ijo.2015.3266