Crohn's disease-associated mucosal factors regulate the expression of TNF-like cytokine 1A and its receptors in primary subepithelial intestinal myofibroblasts and intestinal epithelial cells.
Intestinal subepithelial myofibroblasts (SEMFs) exert a profibrotic role in Crohn's disease (CD). Tumor necrosis factor-like cytokine 1A (TL1A) and its receptors, death-domain receptor 3 (DR3) and decoy receptor 3 (DcR3), are mucosal factors with significant involvement in experimental inflammation and CD. We aimed to determine the regulation of expression of this system of proteins in SEMFs and intestinal epithelial cells. The relative amount of mRNA transcripts for TL1A, DR3, and DcR3 was measured by real-time reverse transcription polymerase chain reaction in cultured primary SEMFs, colonic myofibroblast cell line 18CO, and epithelial cell line HT29. Protein expression was determined by immunofluorescence. The effect of various proinflammatory stimuli in mRNA and protein expression was studied. TL1A mRNA and protein expression in primary SEMFs (and 18CO cells) was significantly upregulated after stimulation with interleukin 1-alpha and/or tumor necrosis factor alpha (TNF-α) (32- to 44-fold increase, P < 0.05 vs unstimulated). Following stimulation with interleukin 1-alpha + TNF-α + IFN-γ, HT-29 cells highly expressed DR3 (4.1-fold over unstimulated, P = 0.008) and DcR3 (56-fold, P = 0.009) and secreted soluble factors that led to induction of TL1A mRNA in primary SEMFs (28-fold, P = 0.008). Activated epithelial cells significantly upregulated IL-8 expression in response to stimulation with recombinant TL1A. Supernatants from mucosal cultures of patients with CD were able to stimulate the expression of TL1A in cultured primary SEMFs, in comparison to supernatants from healthy controls (3.8-fold increase, P < 0.05) or culture media alone (P < 0.05). In conclusion, we found that proinflammatory cytokines are important regulators of the expression of TL1A in SEMFs and of its receptors in intestinal epithelial cells. Our results raise the possibility for involvement of TL1A/DR3/DR3-mediated mechanisms in epithelial-mesenchymal interactions and the development of inflammation-induced intestinal fibrosis in CD.
Academic Department of Gastroenterology, Laiko Hospital, Medical School of National and Kapodistrian University of Athens, University of Athens, Athens, Greece. Electronic address: gbamias@gmail.com.
Giorgos
G
Bamias
Laboratory of Pharmacology, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece.
Eirini
E
Filidou
Laboratory of Infectious Diseases, First Department of Internal Medicine, Laiko Hospital, Medical School of National and Kapodistrian University of Athens, University of Athens, Athens, Greece.
Dimitris
D
Goukos
Laboratory of Gastroenterology, Faculty of Medicine, University of Crete, Heraklion, Greece.
Vasilis
V
Valatas
Laboratory of Pharmacology, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece.
Konstantinos
K
Arvanitidis
Laboratory of Pharmacology, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece.
Maria
M
Panagopoulou
Endoscopy Unit, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece.
Georgios
G
Kouklakis
Laboratory of Infectious Diseases, First Department of Internal Medicine, Laiko Hospital, Medical School of National and Kapodistrian University of Athens, University of Athens, Athens, Greece.
George L
GL
Daikos
Academic Department of Gastroenterology, Laiko Hospital, Medical School of National and Kapodistrian University of Athens, University of Athens, Athens, Greece.
Spiros D
SD
Ladas
Laboratory of Pharmacology, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece.
George
G
Kolios
D003424:Crohn Disease / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D019073:HT29 Cells; D006801:Humans; D018836:Inflammation Mediators / Q000494:pharmacology; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D058628:Myofibroblasts / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D053219:Receptors, Tumor Necrosis Factor, Member 25 / Q000378:metabolism*; D053323:Receptors, Tumor Necrosis Factor, Member 6b / Q000378:metabolism*; D012995:Solubility; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15 / Q000235:genetics / Q000378:metabolism
D018836:Inflammation Mediators; D016209:Interleukin-8; D012333:RNA, Messenger; D053219:Receptors, Tumor Necrosis Factor, Member 25; D053323:Receptors, Tumor Necrosis Factor, Member 6b; C506033:TNFRSF25 protein, human; C505911:TNFRSF6B protein, human; C117026:TNFSF15 protein, human; D053380:Tumor Necrosis Factor Ligand Superfamily Member 15