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Vitamin D regulates claudin-2 and claudin-4 expression in active ulcerative colitis by p-Stat-6 and Smad-7 signaling

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International Journal of Colorectal Disease Aims and scope Submit manuscript

Abstract

Purpose

The tight junctions (TJ) responsible for the integrity of the intestinal barrier are altered in patients with inflammatory bowel disease (IBD), but the physiopathological mechanisms that lead to this alteration are not yet clear. The aim of this study was to determine whether vitamin D, which regulates the integrity of the epithelial barrier by expressing TJ proteins, reduces claudin-2 (Cl-2) levels by inhibiting Stat-6 phosphorylation and whether it increases claudin-4 (Cl-4) levels by blocking Smad-7 activity.

Methods

Biopsies were obtained from inflamed and non-inflamed tracts of the right side colon (caecum or ascending colon) from the same patient with active UC. All the patients were affected by a recent flare-up of ulcerative rectocolitis (RCU), with no previous biologic or immunosuppressive therapy, and all the biopsies were obtained before any treatments. The biopsies were cultured in the presence or not of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3). We also used T84 cells as an in vitro model to perform transfection experiments with Stat-6 and Smad-7.

Results

Our results indicate that 1,25(OH)2D3 is able to regulate CL-2 and CL-4 protein levels, which are increased and reduced in the intestinal mucosa of UC patients, respectively. In the biopsies obtained from UC patients 1,25(OH)2D3 reduces Cl-2 levels by blocking Stat-6 phosphorylation and increases Cl-4 levels by blocking Smad-7 activity. T84 cells, transfected with siRNA of Stat-6 and Smad-7, showed reduced Cl-2 levels and increased Cl-4 levels, confirming that 1,25(OH)2D3 regulates Cl-2 and Cl-4 by decreasing p-Stat-6 and Smad-7 levels.

Conclusions

Our results indicate that the effects of vitamin D on Cl-2 and Cl-4 are mediated by p-Stat-6 and Smad-7 signal, respectively. The study suggests that vitamin D administration to UC patients could be a useful therapeutic intervention, given that vitamin D deficiency is found in these patients.

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References

  1. Khor B, Gardet A, Xavier RJ (2011) Genetics and pathogenesis of inflammatory bowel disease. Nature 474:307–317

    CAS  Google Scholar 

  2. Atreya R, Neurath MF (2015) IBD pathogenesis in 2014: molecular pathways controlling barrier function in IBD. Nat Rev Gastroenterol Hepatol 2:67–68

    Google Scholar 

  3. Peterson LW, Artis D (2014) Intestinal epithelial cells: regulators of barrier function and immune homeostasis. Nat Rev Immunol 14:141–153

    CAS  Google Scholar 

  4. Hooper LV (2015) Epithelial cell contributions to intestinal immunity. Adv Immunol 126:129–172

    CAS  Google Scholar 

  5. Danese S (2012) New therapies for inflammatory bowel disease: from the bench to the bedside. Gut 61:918–932

    CAS  Google Scholar 

  6. Feldman GJ, Mullin JM, Ryan MP (2005) Occludin: structure, function and regulation. Adv Drug Deliv Rev 57:883–917

    CAS  Google Scholar 

  7. Mandell KJ, Parkos CA (2005) The JAM family of proteins. Adv Drug Deliv Rev 57:857–867

    CAS  Google Scholar 

  8. Furuse M, Fujita K, Hiiragi T et al (1998) Claudin-1 and -2, novel integral membrane proteins localizing at tight junctions with no sequence similarity to occludin. J Cell Biol 114:1539–1550

    Google Scholar 

  9. Tsukita S, Furuse M, Itoh M (2001) Multifunctional strands in tight junctions. Nat Rev Mol Cell Biol 2:285–293

    CAS  Google Scholar 

  10. Oshima T, Miwa H, Jon T (2008) Changes in the expression of claudins in active ulcerative colitis. Gastroenterology 23:S146–S150

    CAS  Google Scholar 

  11. Amasheh S, Meiri N, Gitter AH, Schöneberg T, Mankertz J, Schulzke JD, Fromm M (2002) Claudin-2 expression induces cation-selective channels in tight junctions of epithelial cells. J Cell Sci 115:4969–4976

    CAS  Google Scholar 

  12. König J, Wells J, Cani PD et al (2016) Human intestinal barrier function in health and disease. Clin Transl Gastroenterol 7:e196

    Google Scholar 

  13. Schmitz H, Barmeyer C, Fromm M, Runkel N, Foss HD, Bentzel CJ, Riecken EO, Schulzke JD (1999) Altered tight junction structure contributes to the impaired epithelial barrier function in ulcerative colitis. Gastroenterology 116:301–309

    CAS  Google Scholar 

  14. Madara JL, Stafford J (1989) Interferon-gamma directly affects barrier function of cultured intestinal epithelial monolayers. J Clin Invest 83:724–727

    CAS  Google Scholar 

  15. Prasad S, Mingrino R, Kaukinen K, Hayes KL, Powell RM, MacDonald T, Collins JE (2005) Inflammatory processes have differential effects on claudins 2, 3 and 4 in colonic epithelial cells. Lab Investig 85:1139–1162

    CAS  Google Scholar 

  16. Schulzke JD, Ploeger S, Amasheh M, Fromm A, Zeissig S, Troeger H, Richter J, Bojarski C, Schumann M, Fromm M (2009) Epithelial tight junctions in intestinal inflammation. Ann N Y Acad Sci 1165:294–300

    Google Scholar 

  17. Stio M, Retico L, Annese V et al (2016) Vitamin D regulates the tight-junction protein expression in active ulcerative colitis. Scand J Gastroenterol 51:193–199

    Google Scholar 

  18. Hewison M (2012) An update on vitamin D and human immunity. Clin Endocrinol 76:315–325

    CAS  Google Scholar 

  19. Chun RF, Liu PT, Modlin RL et al (2014) Impact of vitamin D on immune function: lessons learned from genome-wide analysis. Front Physiol 5:151–166

    Google Scholar 

  20. Reich KM, Fedorak RN, Madsen K, Kroeker KI (2014) Vitamin D improves inflammatory bowel disease outcomes: basic science and clinical review. World J Gastroenterol 20:4934–4947

    Google Scholar 

  21. Nielsen OH, Lars Rejnmark L, Moss AC (2018) Role of vitamin D in the natural history of inflammatory bowel disease. J Crohn's Colitis 12:742–752

    Google Scholar 

  22. Mouli VP, Ananthakrishnan AN (2014) Review article: vitamin D and inflammatory bowel diseases. Aliment Pharmacol Ther 39:125–136

    CAS  Google Scholar 

  23. Laudisi F, Dinallo V, Di Fusco D et al (2016) Smad7 and its potential as therapeutic target in inflammatory bowel diseases. Curr Drug Metab 17:303–306

    CAS  Google Scholar 

  24. Lennard-Jones JE (1989) Classification of inflammatory bowel disease. Scand J Gastroenterol 24:2–6

    Google Scholar 

  25. Feakins RM (2013) British society of gastroenterology. Inflammatory bowel disease biopsies: updated British Society of Gastroenterology reporting guidelines. J Clin Pathol 66:1005–1026

    Google Scholar 

  26. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein- dye binding. Anal Biochem 72:248–254

    CAS  Google Scholar 

  27. Laukoetter MG, Bruewer M, Nusrat A (2006) Regulation of the intestinal epithelial barrier by the apical junction complex. Curr Opin Gastroenterol 22:85–89

    Google Scholar 

  28. Monteleone G, Kumberova A, Croft NM, McKenzie C, Steer HW, MacDonald T (2001) Blocking Smad7 restores TGF-beta1 signaliNG in chronic inflammatory bowel disease. J Clin Investig 108:601–609

    CAS  Google Scholar 

  29. Rosen MJ, Frey R, Washington MK et al (2011) STAT-6 activation in ulcerative colitis: a new target for prevention of IL-13-induced colon epithelial cell dysfunction. Inflamm Bowel Dis 17:2224–2234

    Google Scholar 

  30. Hershey GK (2003) IL-13 receptors and signaling pathways: an evolving web. J Allergy Clin Immunol 111:677–690

    CAS  Google Scholar 

  31. Rosen MJ, Chaturvedi R, Washington MK et al (2013) STAT6 deficiency ameliorates severity of oxazolone colitis by decreasing expression of claudin-2 and Th2-inducing cytokines. J Immunol 190:1849–1858

    CAS  Google Scholar 

  32. Madden KB, Whitman L, Sullivan C et al (2002) Role of STAT6 and mast cells in IL-4- and IL-13-induced alterations in murine intestinal epithelial cell function. J Immunol 169:4417–4422

    CAS  Google Scholar 

  33. Wu D, Ahrens R, Osterfeld H, Noah TK, Groschwitz K, Foster PS, Steinbrecher KA, Rothenberg ME, Shroyer NF, Matthaei KI, Finkelman FD, Hogan SP (2011) Interleukin-13 (IL-13)/IL-13 receptor alpha1 (IL-13Ralpha1) signaling regulates intestinal epithelial cystic fibrosis transmembrane conductance regulator channel-dependent cl- secretion. J Biol Chem 286:13357–13369

    CAS  Google Scholar 

  34. Fuss IJ, Heller F, Boirivant M, Leon F, Yoshida M, Fichtner-Feigl S, Yang Z, Exley M, Kitani A, Blumberg RS, Mannon P, Strober W (2004) Nonclassical CD1d-restricted NKT cells that produce IL-13 characterize an atypical Th2 response in ulcerative colitis. J Clin Invest 113:1490–1497

    CAS  Google Scholar 

  35. Kadivar K, Ruchelli ED, Markowitz JE, Defelice ML, Strogatz ML, Kanzaria MM, Reddy KP, Baldassano RN, von Allmen D, Brown KA (2004) Intestinal interleukin-13in pediatric inflammatory bowel disease patients. Inflamm Bowel Dis 10:593–598

    Google Scholar 

  36. Vainer B, Nielsen OH, Hendel J, Horn T, Kirman I (2000) Colonic expression and synthesis of interleukin 13 and interleukin 15 in inflammatory bowel disease. Cytokine 12:1531–1536

    CAS  Google Scholar 

  37. Heller F, Florian P, Bojarski C, Richter J, Christ M, Hillenbrand B, Mankertz J, Gitter AH, Bürgel N, Fromm M, Zeitz M, Fuss I, Strober W, Schulzke JD (2005) Interleukin-13 is the key effector Th2 cytokine in ulcerative colitis that affects epithelial tight junctions, apoptosis, and cell restitution. Gastroenterology 129:550–564

    CAS  Google Scholar 

  38. Stio M, Treves C, Martinesi M et al (2005) Biochemical effects of KH 1060 and anti-TNF monoclonal antibody on human peripheral blood mononuclear cells. Int Immunopharmacol 5:649–569

    CAS  Google Scholar 

  39. Bradley JR (2008) TNF-mediated inflammatory disease. J Pathol 214:149–160

    CAS  Google Scholar 

  40. Monteleone G, Boirivant M, Pallone F et al (2008) TGF-beta1 and Smad7 in the regulation of IBD. Mucosal Immunol Suppl 1:S50–S53

    Google Scholar 

  41. Stio M, Martinesi M, Bruni S, Treves C, Mathieu C, Verstuyf A, d'Albasio G, Bagnoli S, Bonanomi AG (2007) The vitamin D analogue TX 527 blocks NF-kappaB activation in peripheral blood mononuclear cells of patients with Crohn's disease. J Steroid Biochem Mol Biol 103:51–60

    CAS  Google Scholar 

  42. Liu W, Chen Y, Golan MA et al (2013) Intestinal epithelial vitamin D receptor signaling inhibits experimental colitis. Clin Invest 123:3983–3996

    CAS  Google Scholar 

  43. Loftus EV Jr (2004) Clinical epidemiology of inflammatory bowel disease: incidence, prevalence, and environmental influence. Gastroenterology 126:1504–1517

    Google Scholar 

  44. Zhu Y, Mahon BD, Froicu M, Cantorna MT (2005) Calcium and 1 alpha,25 Dihydroxyvitamin D3 target the TNF-alpha pathway to suppress experimental inflammatory bowel disease. Eur J Immunol 35:217–224

    CAS  Google Scholar 

  45. Monteleone G, Caruso R, Pallone F (2012) Role of Smad7 in inflammatory bowel diseases. World J Gastroenterol 18:5664–5668

    CAS  Google Scholar 

  46. Monteleone G, Del Vecchio Blanco G, Monteleone I et al (2005) Post-transcriptional regulation of Smad7 in the gut of patients with inflammatory bowel disease. Gastroenterology 129:1420–1429

    CAS  Google Scholar 

  47. Harries AD, Brown R, Heatley RV, Williams LA, Woodhead S, Rhodes J (1985) Vitamin D status in Crohn’s disease: association with nutrition and disease activity. Gut 26:1197–1203

    CAS  Google Scholar 

  48. Ponce de León-Rodríguez M, Guyot JP, Laurent-Babot C (2019) Intestinal in vitro cell culture models and their potential to study the effect of food components on intestinal inflammation. Crit Rev Food Sci Nutr 59:2166–2168

    Google Scholar 

Download references

Acknowledgments

We thank Novella Mugnai for her substantial contribution to the experiments during the preparation of her thesis, and PhD Giulia Lori for her contribution to transfection experiments.

Funding

This study was supported by grants from MIUR [RICATEN18].

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Authors and Affiliations

  1. Department of Biomedical Experimental and Clinical Sciences “Mario Serio”, University of Florence, Viale Morgagni 50, 50134, Florence, Italy

    Vladana Domazetovic, Teresa Iantomasi & Maria Stio

  2. Gastroenterology Unit, Azienda Ospedaliero-Universitaria Careggi, Florence, Italy

    Andrea Giovanni Bonanomi

Authors
  1. Vladana Domazetovic
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  2. Teresa Iantomasi
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  3. Andrea Giovanni Bonanomi
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  4. Maria Stio
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Contributions

Maria Stio designed the study, collected and analyzed the data, and wrote the paper. Vladana Domazetovic performed the majority of experiments. Teresa Iantomasi collected and analyzed the data. Andrea Giovanni Bonanomi provided the collection of all the biopsies that were taken from patients with UC at routine diagnostic colonoscopies and revised the final version of the article, which was approved by all of the authors.

Corresponding author

Correspondence to Maria Stio.

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Conflict of interest

The authors have no conflict of interest related to the study.

Ethical approval

The study protocol was approved by the Medical Ethics Committee, and with the Helsinki Declaration of 1975, as revised in 1983. CE May 2, 2011, protocol 0016888, rif. 95/10.

Statement of informed consent/Human and animal rights and informed consent

Informed consent was obtained from all patients prior to their inclusion in this study

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Domazetovic, V., Iantomasi, T., Bonanomi, A.G. et al. Vitamin D regulates claudin-2 and claudin-4 expression in active ulcerative colitis by p-Stat-6 and Smad-7 signaling. Int J Colorectal Dis 35, 1231–1242 (2020). https://doi.org/10.1007/s00384-020-03576-0

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