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Inhibition of Nucleostemin Upregulates CDX2 Expression in HT29 Cells in Response to Bile Acid Exposure: Implications in the Pathogenesis of Barrett’s Esophagus

  • Original Article
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Journal of Gastrointestinal Surgery

Abstract

Background

Barrett’s esophagus (BE), a squamous-to-columnar metaplasia, may originate from growth-promoting mutations in metaplastic stem cells. Nucleostemin is a protein highly expressed in undifferentiated embryonic stem cells. The objectives of this study were to explore the potential role of nucleostemin in the pathogenesis of BE

Methods

The expression profiles of 30,968 genes were compared between BE and normal esophageal tissues (n = 6 in each group) by using oligo microarray. Three siRNA plasmid expression vectors against nucleostemin, pRNAi-1, pRNAi-2, and pRNAi-3, were constructed and transfected into HT29 cells. In addition, HT29 cells were exposed to 100–1,000 μM chenodeoxycholic acid (CDC), a bile acid, for 2, 12, and 24 h, and then messenger RNA and protein expressions of nucleostemin and CDX2 were determined by reverse-transcriptase polymerase chain reaction and Western blotting.

Results

Four hundred and twenty-six differentially expressed genes were detected in BE; 142 were upregulated and 284 downregulated. Nucleostemin was downregulated while CDX2 was upregulated. In vitro, all the recombinant plasmids inhibited the nucleostemin expression in transfected HT29 cells, with pRNAi-1 being the most effective. CDX2 expression was significantly increased in pRNAi-1-transfected HT29 cells, compared with that in the empty plasmid (pRNAT-U6.1/Neo) transfected or untransfected HT29 cells. In addition, CDX2 expression was increased whereas nucleostemin expression was decreased in a dose- and time-dependent manner in HT29 cells treated with CDC.

Conclusion

These findings suggest that the inhibition of nucleostemin expression in “esophageal stem cells” in response to bile acid exposure may be involved in the pathogenesis of BE through upregulating CDX2 expression.

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References

  1. Wild CP, Hardle LJ. Reflux, Barrett’s oesophagus and adenocarcinoma: burning questions. Nat Rev Cancer 2003;3:676–684. doi:10.1038/nrc1166.

    Article  CAS  PubMed  Google Scholar 

  2. Vizcaino AP, Moreno V, Lambert R, Perkin DM. Time trends incidence of both major histologic types of esophageal carcinomas in selected countries, 1973–1995. Int J Cancer 2002;99:860–868. doi:10.1002/ijc.10427.

    Article  CAS  PubMed  Google Scholar 

  3. Paulson TG, Reid BJ. Focus on Barrett’s esophagus and esophageal adenocarcinoma. Cancer Cell 2004;6:1–16. doi:10.1016/j.ccr.2004.06.021.

    Article  Google Scholar 

  4. Lagergren J, Bergstrom R, Lindgren A, Nyrén O. Symptomatic gastroesophageal reflux as a risk factor for esophageal adenocarcinoma. N Engl J Med 1999;340:825–831. doi:10.1056/NEJM199903183401101.

    Article  CAS  PubMed  Google Scholar 

  5. Tsai RY, McKay RD. A nucleolar mechanism controlling cell proliferation in stem cells and cancer cells. Genes Dev 2002;16:2991–3003. doi:10.1101/gad.55671.

    Article  CAS  PubMed  Google Scholar 

  6. Kafienah W, Mistry S, Williams C, Hollander AP. Nucleostemin is a marker of proliferating stromal stem cells in adult human bone marrow. Stem Cells 2006;24:1113–1120. doi:10.1634/stemcells.2005-0416.

    Article  PubMed  Google Scholar 

  7. Zhang GY, Yin L, Li SL, **ng WY, Zhao QM, Le XP, Gao DL, Chen KS, Zhang YH, Zhang QX. Expression of nucleostemin mRNA and protein in the esophageal squamous cell carcinoma. Zhonghua Zhong Liu Za Zhi 2008;30:125–128. (Chinese).

    CAS  PubMed  Google Scholar 

  8. Fitzgerald RC, Omary MB, Triadafilopoulos G. Acid modulation of HT29 cell growth and differentiation an in vitro model for Barrett’s esophagus. J Cell Sci 1997;110:663–671.

    CAS  PubMed  Google Scholar 

  9. White NM, Gabril M, Ejeckam G, Mathews M, Fardy J, Kamel F, Doré J, Yousef GM. Barrett’s esophagus and cardiac intestinal metaplasia: two conditions within the same spectrum. Can J Gastroenterol 2008;22:369–375.

    PubMed  Google Scholar 

  10. Silberg DG, Swain GP, Suh ER, Traber PG. CDX1 and CDX2 expression during intestinal development. Gastroenterology 2000;119:961–971. doi:10.1053/gast.2000.18142.

    Article  CAS  PubMed  Google Scholar 

  11. Guo RJ, Suh ER, Lynch JP. The role of CDX proteins in intestinal development and cancer. Cancer Biol Ther 2004;3:593–601.

    Article  CAS  PubMed  Google Scholar 

  12. Chen X, Yang CS. Esophageal adenocarcinoma: a review and perspectives on the mechanism of carcinogenesis and chemoprevention. Carcinogenesis 2001;22:1119–1129. doi:10.1093/carcin/22.8.1119.

    Article  CAS  PubMed  Google Scholar 

  13. Nishijima K, Miwa K, Miyashita T, Kinami S, Ninomiya I, Fushida S, Fujimura T, Hattori T. Impact of the biliary diversion procedure on carcinogenesis in Barrett’s esophagus surgically induced by duodenoesophageal reflux in rats. Ann Surg 2004;240:57–67. doi:10.1097/01.sla.0000130850.31178.8c.

    Article  PubMed  Google Scholar 

  14. Fitzgerald RC, Omary MB, Triadafilopoulos G. Dynamic effects of acid on Barrett’s esophagus. An ex vivo proliferation and differentiation model. J Clin Invest 1996;98:2120–2128. doi:10.1172/JCI119018.

    Article  CAS  PubMed  Google Scholar 

  15. Fein M, Ireland AP, Ritter MP, Peteres JH, Hagen JA, Bremner CG, DeMeester TR. Duodenogastric reflux potentiates the injurious effects of gastroesophageal reflux. J Gastrointest Surg 1997;1:27–32. discussion 33doi:10.1007/s11605-006-0006-x.

    Article  CAS  PubMed  Google Scholar 

  16. Kauer WK, Peters JH, DeMeester TR, Ireland AP, Bremner CG, Hagen JA. Mixed reflux of gastric and duodenal juices is more harmful to the esophagus than gastric juice alone; The need for surgical therapy re-emphasized. Ann Surg 1995;222:525–531. discussion 531–533. doi:10.1097/00000658-199522240-00010.

    CAS  PubMed  Google Scholar 

  17. Odze RD. Unraveling the mystery of the gastroesophageal junction: a pathologist’s perspective. Am J Gastroenterol 2005;100:1853–1067. doi:10.1111/j.1572-0241.2005.50096.x.

    Article  PubMed  Google Scholar 

  18. Jiang Y, Jahagirdar BN, Reinhardt RL, Schwartz RE, Keene CD, Ortiz-Gonzalez XR, Reyes M, Lenvik T, Lund T, Blackstad M, Du J, Aldrich S, Lisberg A, Low WC, Largaespada DA, Verfaillie CM. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 2002;418:41–49. doi:10.1038/nature00870.

    Article  CAS  PubMed  Google Scholar 

  19. Houghton J, Stoicov C, Nomura S, Rogers AB, Carlson J, Li H, Cai X, Fox JG, Goldenring JR, Wang TC. Gastric cancer originating from bone marrow-derived cells. Science 2004;306:1568–1571. doi:10.1126/science.1099513.

    Article  CAS  PubMed  Google Scholar 

  20. Sarosi G, Brown G, Jaiswal K, Feagins LA, Lee E, Crook TW, Souza RF, Zou YS, Shay JW, Spechler SJ. Bone marrow progenitor cells contribute to esophageal regeneration and metaplasia in a rat model of Barrett’s esophagus. Dis Esophagus 2008;21:43–50.

    CAS  PubMed  Google Scholar 

  21. Glickman JN, Chen YY, Wang HH, Antonioli DA, Odze RD. Phenotypic characteristics of a distinctive multilayered epithelium suggests that it is a precursor in the development of Barrett’s esophagus. Am J Surg Pathol 2001;25:569–578. doi:10.1097/00000478-200105000-00002.

    Article  CAS  PubMed  Google Scholar 

  22. Boulton RA, Usselmann B, Mohammed I, Jankowski J. Barrett’s esophagus: environmental influences in the progression of dysplasia. World J Surg 2003;27:1014–1017. doi:10.1007/s00268-003-7054-0.

    Article  PubMed  Google Scholar 

  23. Spechler SJ, Souza RF. Stem cells in Barrett’s esophagus: HALOs or horns? Gastrointest Endosc 2008;68:41–43. doi:10.1016/j.gie.2008.02.080.

    Article  PubMed  Google Scholar 

  24. Souza RF, Krishnan K, Spechler SJ. Acid, bile, and CDX: the ABCs of making Barrett’s metaplasia. Am J Physiol Gastrointest Liver Physiol 2008;295:G211–G218. doi:10.1152/ajpgi.90250.2008.

    Article  CAS  PubMed  Google Scholar 

  25. Seery JP. Stem cells of the oesophageal epithelium. J Cell Sci 2002;115:1783–1789.

    PubMed  Google Scholar 

  26. Shields HM, Rosenberg SJ, Zwas FR, Ransil BJ, Lembo AJ, Odze R. Prospective evaluation of multilayered epithelium in Barrett’s esophagus. Am J Gastroenterol 2001;96:3268–3273. doi:10.1111/j.1572-0241.2001.05324.x.

    Article  CAS  PubMed  Google Scholar 

  27. Suh E, Traber PG. An intestine-specific homeobox gene regulates proliferation and differentiation. Mol Cell Biol 1996;16:619–625.

    CAS  PubMed  Google Scholar 

  28. Yamamoto H, Bai YQ, Yuasa Y. Homeodomain protein CDX2 regulates goblet-specific MUC2 gene expression. Biochem Biophys Res Commun 2003;300:813–818. doi:10.1016/S0006-291X(02)02935-2.

    Article  CAS  PubMed  Google Scholar 

  29. Mesquita P, Jonckheere N, Almeida R, Ducourouble MP, Serpa J, Silva E, Pigny P, Silva FS, Reis C, Silberg D, Van Seuningen I, David L. Human MUC2 mucin gene is transcriptionally regulated by Cdx homeodomain proteins in gastrointestinal carcinoma cell lines. J Biol Chem 2003;278:51549–51556. doi:10.1074/jbc.M309019200.

    Article  CAS  PubMed  Google Scholar 

  30. Groisman GM, Amar M, Meir A. Expression of the intestinal marker CDX2 in the columnar-lined esophagus with and without intestinal (Barrett’s) metaplasia. Mod Pathol 2004;17:1282–1288. doi:10.1038/modpathol.3800182.

    Article  CAS  PubMed  Google Scholar 

  31. Phillips RW, Frierson HF Jr, Moskaluk CA. CDX2 as a marker of epithelial intestinal differentiation in the esophagus. Am J Surg Pathol 2003;27:1442–1447. doi:10.1097/00000478-200311000-00006.

    Article  PubMed  Google Scholar 

  32. Vallböhmer D, DeMeester SR, Peters JH, Oh DS, Kuramochi H, Shimizu D, Hagen JA, Danenberg KD, Danenberg PV, DeMeester TR, Chandrasoma PT. Cdx-2 expression in squamous and metaplastic columnar epithelia of the esophagus. Dis Esophagus 2006;19:260–266. doi:10.1111/j.1442-2050.2006.00586.x.

    Article  PubMed  Google Scholar 

  33. Kaimaktchiev V, Terracciano L, Tornillo L, Spichtin H, Stoios D, Bundi M, Korcheva V, Mirlacher M, Loda M, Sauter G, Corless C. The homeobox intestinal differentiation factor CDX2 is selectively expressed in gastrointestinal adenocarcinomas. Mod Pathol 2004;17:1392–1399. doi:10.1038/modpathol.3800205.

    Article  CAS  PubMed  Google Scholar 

  34. Werling RW, Yaziji H, Bacchi CE, Gown AM. CDX2, a highly sensitive and specific marker of adenocarcinomas of intestinal origin: an immunohistochemical survey of 476 primary and metastatic carcinomas. Am J Surg Pathol 2003;27:303–310. doi:10.1097/00000478-200303000-00003.

    Article  PubMed  Google Scholar 

  35. Keller MS, Ezaki T, Guo RJ, Lynch JP. Cdx1 or Cdx2 expression activates E-cadherin-mediated cell–cell adhesion and compaction in human COLO 205 cells. Am J Physiol Gastrointest Liver Physiol 2004;287:G104–G114. doi:10.1152/ajpgi.00484.2003.

    Article  CAS  PubMed  Google Scholar 

  36. Shi XY, Bhagwandeen B, Leong AS. CDX2 and villin are useful markers of intestinal metaplasia in the diagnosis of Barrett esophagus. Am J Clin Pathol 2008;129:571–577. doi:10.1309/UWK3NAHV31GFHM3J.

    Article  PubMed  Google Scholar 

  37. Kumble S, Omary MB, Fajardo LF, Triadafilopoulos G. Multifocal heterogeneity in villin and Ep-cam expression in Barrett’s esophagus. Int J Cancer 1996;66:48–54. doi:10.1002/(SICI)1097-0215(19960328)66:1<48::AID-IJC9>3.0.CO;2-Z.

    Article  CAS  PubMed  Google Scholar 

  38. Zweibaum A, Pinto M, Chevalier G, Dussaulx E, Triadou N, Lacroix B, Haffen K, Brun JL, Rousset M. Enterocytic differentiation of a subpopulation of the human colon tumor cell line HT-29 selected for growth in sugar-free medium and its inhibition by glucose. J Cell Physiol 1985;122:21–29. doi:10.1002/jcp.1041220105.

    Article  CAS  PubMed  Google Scholar 

  39. Serakinci N, Guldberg P, Burns JS, Abdallah B, Schrødder H, Jensen T, Kassem M. Adult human mesenchymal stem cell as a target for neoplastic transformation. Oncogene 2004;23:5095–5098. doi:10.1038/sj.onc.1207651.

    Article  CAS  PubMed  Google Scholar 

  40. Zhang J, Mayer AN. Possible role for nucleostemin in intestinal progenitor cells. Gastroenterology 2007;132:A629–A629.

    Google Scholar 

  41. Kazumori H, Ishihara S, Rumi MA, Kadowaki Y, Kinoshita Y. Bile acids directly augment caudal related homeobox gene CDX2 expression in oesophageal keratinocytes in Barrett’s epithelium. Gut 2006;55:6–25. doi:10.1136/gut.2005.066209.

    Article  CAS  Google Scholar 

  42. Soubeyran P, Mallo GV, Moucadel V, Dagorn JC, Iovanna JL. Overexpression of Cdx1 and Cdx2 homeogenes enhances expression of the HLA-I in HT-29 cells. Mol Cell Biol Res Commun 2000;3:271–276. doi:10.1006/mcbr.2000.0226.

    Article  CAS  PubMed  Google Scholar 

  43. Domon-Dell C, Wang Q, Kim S, Kedinger M, Evers BM, Freund JN. Stimulation of the intestinal Cdx2 homeobox gene by butyrate in colon cancer cells. Gut 2006;50:525–529. doi:10.1136/gut.50.4.525.

    Article  Google Scholar 

  44. Kim S, Domon-Dell C, Wang QD, Chung DH, Cristofano AD, Pandolfi PP, Freund JN, Evers BM. PTEN and TNF-a regulation of the intestinal specific Cdx-2 homeobox gene through a PI3K, PKB/Akt, and NF-κB-dependent pathway. Gastroenterology 2006;123:1163–1178. doi:10.1053/gast.2002.36043.

    Article  CAS  Google Scholar 

  45. Marchetti M, Caliot E, Pringault E. Chronic acid exposure leads to activation of the Cdx2 intestinal homeobox gene in a long-term culture of mouse esophageal keratinocytes. J Cell Sci 2006;116:1429–1436. doi:10.1242/jcs.00338.

    Article  CAS  Google Scholar 

  46. Debruyne PR, Witek M, Gong L, Birbe R, Chervoneva I, ** T, Domon-Cell C, Palazzo JP, Freund JN, Li P, Pitari GM, Schulz S, Waldman SA. Bile acids induce ectopic expression of intestinal guanylyl cyclase C Through nuclear factor kappa B and CDX2 in human esophageal cells. Gastroenterology 2006;130:1191–1206. doi:10.1053/j.gastro.2005.12.032.

    Article  CAS  PubMed  Google Scholar 

  47. Souza RF, Shewmake K, Terada LS, Spechler SJ. Acid exposure activates the mitogen-activated protein kinase pathways in Barrett’s esophagus. Gastroenterology 2002;122:299–307. doi:10.1053/gast.2002.30993.

    Article  CAS  PubMed  Google Scholar 

  48. Houde M, Laprise P, Jean D, Blais M, Asselin C, Rivard N. Intestinal epithelial cell differentiation involves activation of p38 mitogen-activated protein kinase that regulates the homeobox transcription factor CDX2. J Biol Chem 2001;276:21885–21894. doi:10.1074/jbc.M100236200.

    Article  CAS  PubMed  Google Scholar 

  49. Hu Y, Jones C, Gellersen O, Williams VA, Watson TJ, Peters JH. Pathogenesis of Barrett esophagus: deoxycholic acid up-regulates goblet-specific gene MUC2 in concert with CDX2 in human esophageal cells. Arch Surg 2007;142:540–544. doi:10.1001/archsurg.142.6.540.

    Article  CAS  PubMed  Google Scholar 

  50. Capello A, Moons LM, Van de Winkel A, Siersema PD, van Dekken H, Kuipers EJ, Kusters JG. Bile acid-stimulated expression of the farnesoid X receptor enhances the immune response in Barrett esophagus. Am J Gastroenterol 2008;103:1510–1516. doi:10.1111/j.1572-0241.2008.01908.x.

    Article  CAS  PubMed  Google Scholar 

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  1. Department of Gastroenterology, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China

    Yong-Gang Sun, **ng-Wei Wang, Shi-Ming Yang, Gang Zhou, Wei-Qiang Wang, Hong-Bin Wang, Rong-Quan Wang & Dian-Chun Fang

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Sun, YG., Wang, XW., Yang, SM. et al. Inhibition of Nucleostemin Upregulates CDX2 Expression in HT29 Cells in Response to Bile Acid Exposure: Implications in the Pathogenesis of Barrett’s Esophagus. J Gastrointest Surg 13, 1430–1439 (2009). https://doi.org/10.1007/s11605-009-0899-2

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