*Department of Women Health and Territorial Medicine, University of Rome La Sapienza, Rome
†Neonatal Unit, AORN “V. Monaldi,” Naples
‡Department of Pediatrics, University of Naples “Federico II,” Naples, Italy.
Address correspondence and reprint requests to Gianluca Terrin, MD, PhD, Department of Women's Health and Territorial Medicine, University of Rome “La Sapienza” via di Grottarossa 1035/1039 Rome, Italy (e-mail: email@example.com).
Received 9 November, 2011
Accepted 10 December, 2011
The authors report no conflicts of interest.
See “Methotrexate Modulates Tight Junctions Through NF-κB, MEK, and JNK Pathways” by Beutheu Youmba et al on page 463.
Alteration in intestinal permeability is involved in the pathogenesis of several diseases (1). Intestinal permeability depends on the integrity of the function of the intestinal barrier. The intestinal barrier consists of epithelial cells, tight junctions (TJs), adherens junctions, and luminal secretions (1,2). The TJs and adherens junctions are collectively referred to as the apical junctional complexes (AJCs), which are involved in multiple functions (1–5). They are dynamic structures that normally regulate the trafficking of nutrients, compounds, and fluids between the intestinal lumen and the submucosa. These structures also act as a boundary within the plasma membrane itself, separating the apical and basolateral cell surface domains. They play an important role in intestinal morphogenesis, differentiation, and wound healing (1,2). Finally, AJCs participate in signal transduction across epithelial cells in both directions and in the regulation of many cytoskeleton functions (1,2). Major proteins in the AJCs are transmembrane proteins, occludins, claudins, junctional adhesion molecules, coxsackie adenovirus receptor, cytoplasmic plaque proteins, zonula occludens (ZOs), zonulin receptors, cingulin, cytoplasmic TJ-associated protein (7H6), and E-cadherin (2,6). The ZOs are the best-characterized proteins of the AJC. The ZO-1 and ZO-2 are involved in the organization of proteins within the tight junctional plaque so that signaling events can be propagated (7). Occludin is a transmembrane protein that serves as cell-to-cell adhesion molecule and maintains the cellular polarization and intramembrane fence that restricts the diffusion of lipids in the outer leaflet of the plasma membrane (8). Claudins interact with ZO-1, ZO-2, and ZO-3 and are able to polymerize and form TJ strands in the absence of the ZO-binding region (7). Components of AJCs interact with each other, and their function can be modulated by intrinsic or extrinsic stimuli with consequent modification of intestinal permeability (5).
In this issue of JPGN, Youmba et al (9) report a dose-dependent effect of methotrexate (MTX) on intestinal permeability. Results of the present study showed that MTX regulates expression and cellular distribution of several TJ proteins such as occludins, claudins, and ZO-1. In the present article, the authors demonstrated that the intracellular effects of MTX involve mitogen-activated protein kinases (MAPK) and nuclear factor kappa light-chain enhancer of activated B cells (NF-κB), according to studies that have suggested an important role for these pathways in the regulation of intestinal barrier functions and wound healing (10). MTX is an inhibitor of dihydrofolate reductase and DNA synthesis that is widely used, at high doses, in cancer chemotherapy, and at lower doses, as anti-inflammatory agents, in chronic inflammatory diseases such as inflammatory bowel diseases (11,12). MTX may determine intestinal injury by inducing apoptosis, hypoproliferation, inflammation, and bacterial colonization (10,11). A further effect of MTX on intestinal TJs increases the concerns regarding the use of this chemotherapeutic agent in children. The effect of MTX on intestinal permeability may result in an increased transport of xenobiotics and pathogens across the epithelial barrier with further risk of infections in immunocompromised subjects (13). In addition, MTX may have potential effects on the brain barrier (14). At the same time, the manipulation of intestinal permeability could represent an interesting therapeutic opportunity to limit the adverse effects of MTX and other chemotherapeutic agents. In the last decades, many drugs have been evaluated for their ability to modify intestinal permeability, acting on TJ. A new molecule, namely AT1001, entered clinical trial in humans (15,16). AT1001 is a zonulin peptide inhibitor that competitively blocks the apical zonulin receptor and prevents the opening of TJ (15,16). This peptide has been tested in subjects with celiac disease (15). More recently in animal models of inflammatory bowel disease, the AT1001 significantly attenuates colitis, reducing intestinal permeability (16). In the near future, the use of this new molecule may help to minimize the deleterious consequences of chemotherapy on intestinal permeability; however, an increasing number of molecules have been proposed as enhancers of drug paracellular delivery via TJs modulation (7,17–19). Manipulation of paracellular transport represents an interesting field of research. Modification of paracellular permeability may be used to increase oral drug delivery and replace parenteral with enteral route for the administration of therapeutic compounds. In contrast to strategies targeting transepithelial/transendothelial pathways, modulators acting on TJs could enhance paracellular permeability of biological barriers for a large number and variety of drugs, including hydrophylic compounds and biopharmaceuticals such as peptides, proteins, nucleic acids, and viral vectors, without the need to modify the drugs (7). Unfortunately, the use of the absorption enhancers proposed so far is limited because of scarce selectivity and potential toxicity (7); however, interesting consequences may derive from trials with PN159 and YY3–36 used in patients with type 2 diabetes mellitus to enhance insulin absorption (7). In this scenario, it is reasonable to speculate that MTX could be responsible for the enhanced intestinal absorption of itself and of other chemotherapeutic or immunosuppressant drugs (20). In this way, MTX may improve the clinical efficacy of the therapeutic strategy based on the use of multiple drugs simultaneously (10–21). Thus, a careful analysis of the effects on intestinal permeability should be included in the pharmacodynamic assessment of old and new chemotherapeutic and immunosuppressant drugs (22).
Controlled and reversible opening of the TJs by safe and effective molecules is a fascinating challenge for pediatric gastroenterology in the next decade (23). Although new TJ regulators are under development, many drugs presently used in children with specific indications may have additional effects on intestinal permeability that remain to be defined (24). Thus, aspects that were considered the “dark side” of such therapy could represent an opportunity to improve the efficacy of several therapeutic strategy in the near future.
1. Chiba H, Osanai M, Murata M, et al. Transmembrane proteins of tight junctions. Biophys Acta 2008; 1778:729–756.
2. Laukoetter MG, Bruewer M, Nusrat A. Regulation of the intestinal epithelial barrier by the apical junctional complex. Curr Opin Gastroenterol 2006; 22:85–89.
3. Laukoetter MG, Nava P, Nusrat A. Role of the intestinal barrier in inflammatory bowel disease. World J Gastroenterol 2008; 14:401–407.
4. Ivanov AI, Nusrat A, Parkos CA. Endocytosis of the apical junctional complex: mechanisms and possible roles in regulation of epithelial barriers. Bioassays 2005; 27:27–31.
5. Iizuka M, Konno S. Wound healing of intestinal epithelial cells. World J Gastroenterol 2011; 17:2161–2171.
6. Fasano A. Zonulin and its regulation of intestinal barrier function: the biological door to inflammation, autoimmunity, and cancer. Physiol Rev 2011; 91:151–175.
7. Deli MA. Potential use of tight junction modulators to reversibly open membranous barriers and improve drug delivery. Biochim Biophys Acta 2009; 1788:892–910.
8. Balda MS, Whitney JA, Flores C, et al. Functional dissociation of paracellular permeability and transepithelial electrical resistance and disruption of the apical-basolateral intramembrane diffusion barrier by expression of a mutant tight junction membrane protein. J Cell Biol 1996; 134:1031–1049.
9. Youmba SB, Belmonte L, Galas L, et al. Methotrexate modulates tight junctions through NF-κB, MEK and JNK pathways. J Pediatr Gastroenterol Nutr 2012;54:463–70.
10. Dabrowska M, Skoneczny M, Rode W. Functional gene expression profile underlying methotrexate-induced senescence in human colon cancer cells. Tumour Biol 2011; 32:965–976.
11. Bertino JR. Cancer research: from folate antagonism to molecular targets. Best Pract Res Clin Haematol 2009; 22:577–582.
12. Khan KJ, Dubinsky MC, Ford AC, et al. Efficacy of immunosuppressive therapy for inflammatory bowel disease: a systematic review and meta-analysis. Am J Gastroenterol 2011; 106:630–642.
13. Song D, Shi B, Xue H, et al. Confirmation and prevention of intestinal barrier dysfunction and bacterial translocation caused by methotrexate. Dig Dis Sci 2006; 51:1549–1556.
14. Froklage FE, Reijnebeld JC, Heimans JJ. Central neurotoxicity in cancer chemotherapy: pharmacogenetic insights. Pharmacogenomics 2011; 12:379–395.
15. Paterson BM, Lammers KM, Arrieta MC, et al. The safety, tolerance, pharmacokinetic and pharmacodynamic effects of single doses of AT-1001 in coeliac disease subjects: a proof of concept study. Aliment Pharmacol Ther 2007; 26:757–766.
16. Arrieta MC, Madsen K, Doyle J, et al. Reducing small intestinal permeability attenuates colitis in the IL10 gene-deficient mouse. Gut 2009; 58:41–48.
17. Kondoh M, Yagi K. Tight junction modulators: promising candidates for drug delivery. Curr Med Chem 2007; 14:2482–2488.
18. Matsuhisa K, Kondoh M, Takahashi A, et al. Tight junction modulator and drug delivery. Expert Opin Drug Deliv 2009; 6:509–515.
19. Salama NN, Eddington ND, Fasano A. Tight junction modulation and its relationship to drug delivery. Adv Drug Deliv Rev 2006; 58:15–28.
20. Innocenti F, Danesi R, Di Paolo A, et al. Clinical and experimental pharmacokinetic interaction between 6-mercaptopurine and methotrexate. Cancer Chemother Pharmacol 1996; 37:409–414.
21. Rath T, Rubbert A. Drug combinations with methotrexate to treat rheumatoid arthritis. Clin Exp Rheumatol 2010; 28:S52–S57.
22. Shugarts S, Benet LZ. The role of transporters in the pharmacokinetics of orally administered drugs. Pharm Res 2009; 26:2039–2054.
23. Fasano A, Shea-Donohue T. Mechanisms of disease: the role of intestinal barrier function in the pathogenesis of gastrointestinal autoimmune diseases. Nat Clin Pract Gastroenterol Hepatol 2005; 2:416–422.
24. Lennernäs H, Knutson L, Knutson T, et al. The effect of amiloride on the in vivo effective permeability of amoxicillin in human jejunum: experience from a regional perfusion technique. Eur J Pharm Sci 2002; 15:271–277.