Inflammation Is Present, Persistent and More Sensitive to Proinflammatory Triggers in Celiac Disease Enterocytes
Abstract
:1. Introduction
2. Results
2.1. In CD Biopsies, the Inflammatory Markers IL-1β and IL-6 Are Increased in Enterocytes
2.2. The Inflammatory Markers pNF-κB, pERK, IL-1β, and IL-6 Were Increased and Persistent in CD Organoids
2.3. 3D and 2D Organoids from CD Patients Had Increased Inflammatory Markers Compared to Those from CTR Patients
2.4. Organoids from CD Patients Were More Sensitive to P31-43
2.5. Organoids from CD Patients Were More Sensitive to Lox
3. Discussion
4. Materials and Methods
4.1. Organoids
4.2. Culture Medium to Maintain Organoids (CM-S)
4.3. Fixing of Organoids, OCT Embedding, and Cryosectioning
4.4. Immunostaining
4.5. Western Blot
4.6. ELISA
4.7. RNAscope to Detect IL-1β and IL-6 mRNA
4.8. PCR
4.9. Statical Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
- Sollid, L.M.; Jabri, B. Triggers and drivers of autoimmunity: Lessons from coeliac disease. Nat. Rev. Immunol. 2013, 13, 294–302. [Google Scholar] [CrossRef] [Green Version]
- Abenavoli, L.; Dastoli, S.; Bennardo, L.; Boccuto, L.; Passante, M.; Silvestri, M.; Proietti, I.; Potenza, C.; Luzza, F.; Nisticò, S.P. The Skin in Celiac Disease Patients: The Other Side of the Coin. Medicina 2019, 55, 578. [Google Scholar] [CrossRef] [Green Version]
- Lindfors, K.; Ciacci, C.; Kurppa, K.; Lundin, K.E.A.; Makharia, G.K.; Mearin, M.L.; Murray, J.A.; Verdu, E.F.; Kaukinen, K. Coeliac disease. Nat. Rev. Dis. Primers 2019, 5, 3. [Google Scholar] [CrossRef]
- Bouziat, R.; Hinterleitner, R.; Brown, J.J.; Stencel-Baerenwald, J.E.; Ikizler, M.; Mayassi, T.; Meisel, M.; Kim, S.M.; Discepolo, V.; Pruijssers, A.J.; et al. Reovirus infection triggers inflammatory responses to dietary antigens and development of celiac disease. Science 2017, 356, 44–50. [Google Scholar] [CrossRef] [Green Version]
- Barone, M.V.; Troncone, R.; Auricchio, S. Gliadin peptides as triggers of the proliferative and stress/innate immune response of the celiac small intestinal mucosa. Int. J. Mol. Sci. 2014, 15, 20518–20537. [Google Scholar] [CrossRef] [Green Version]
- Chirdo, F.G.; Auricchio, S.; Troncone, R.; Barone, M.V. The gliadin p31-43 peptide: Inducer of multiple proinflammatory effects. Int. Rev. Cell Mol. Biol. 2021, 358, 165–205. [Google Scholar]
- Junker, Y.; Zeissig, S.; Kim, S.J.; Barisani, D.; Wieser, H.; Leffler, D.A.; Zevallos, V.; Libermann, T.A.; Dillon, S.; Freitag, T.L.; et al. Wheat amylase trypsin inhibitors drive intestinal inflammation via activation of toll-like receptor 4. J. Exp. Med. 2012, 209, 2395–2408. [Google Scholar] [CrossRef]
- Barroso, M.; Beth, S.A.; Voortman, T.; Jaddoe, V.W.V.; van Zelm, M.C.; Moll, H.A.; Kiefte-de Jong, J.C. Dietary Patterns after the Weaning and Lactation Period Are Associated with Celiac Disease Autoimmunity in Children. Gastroenterology 2018, 154, 2087–2096.e7. [Google Scholar] [CrossRef] [Green Version]
- Barone, M.V.; Auricchio, S. A Cumulative Effect of Food and Viruses to Trigger Celiac Disease (CD): A Commentary on the Recent Literature. Int. J. Mol. Sci. 2021, 22, 2027. [Google Scholar] [CrossRef]
- Deets, K.A.; Vance, R.E. Inflammasomes and adaptive immune responses. Nat. Immunol. 2021, 22, 412–422. [Google Scholar] [CrossRef]
- Stamnaes, J.; Stray, D.; Stensland, M.; Sarna, V.K.; Nyman, T.A.; Lundin, K.E.A.; Sollid, L.M. In Well-Treated Celiac Patients Low-Level Mucosal Inflammation Predicts Response to 14-day Gluten Challenge. Adv. Sci. 2021, 8, 2003526. [Google Scholar] [CrossRef]
- Dotsenko, V.; Oittinen, M.; Taavela, J.; Popp, A.; Peräaho, M.; Staff, S.; Sarin, J.; Leon, F.; Isola, J.; Mäki, M.; et al. Genome-Wide Transcriptomic Analysis of Intestinal Mucosa in Celiac Disease Patients on a Gluten-Free Diet and Postgluten Challenge. Cell Mol. Gastroenterol. Hepatol. 2021, 11, 13–32. [Google Scholar] [CrossRef]
- Hayden, M.S.; Ghosh, S. NFkBin immunobiology. Cell Res. 2011, 21, 223–244. [Google Scholar] [CrossRef] [Green Version]
- Trynka, G.; Zhernakova, A.; Romanos, J.; Franke, L.; Hunt, K.A.; Turner, G.; Bruinenberg, M.; Heap, G.A.; Platteel, M.; Ryan, A.W.; et al. Coeliac disease-associated risk variants in TNFAIP3 and REL implicate altered NF-kappaB signalling. Gut 2009, 58, 1078–1083. [Google Scholar] [CrossRef] [Green Version]
- Fernandez-Jimenez, N.; Castellanos-Rubio, A.; Plaza-Izurieta, L.; Irastorza, I.; Elcoroaristizabal, X.; Jauregi-Miguel, A.; Lopez-Euba, T.; Tutau, C.; de Pancorbo, M.M.; Vitoria, J.C.; et al. Coregulation and modulation of NFκB-related genes in celiac disease: Uncovered aspects of gut mucosal inflammation. Hum. Mol. Genet. 2014, 23, 1298–1310. [Google Scholar] [CrossRef]
- Castellanos-Rubio, A.; Bilbao, J.R. Profiling Celiac Disease-Related Transcriptional Changes. Int. Rev. Cell Mol. Biol. 2018, 336, 149–174. [Google Scholar]
- Castellanos-Rubio, A.; Ghosh, S. Disease-Associated SNPs in Inflammation-Related lncRNAs. Front. Immunol. 2019, 10, 420. [Google Scholar] [CrossRef] [Green Version]
- Lania, G.; Nanayakkara, M.; Maglio, M.; Auricchio, R.; Porpora, M.; Conte, M.; De Matteis, M.A.; Rizzo, R.; Luini, A.; Discepolo, V.; et al. Constitutive alterations in vesicular trafficking increase the sensitivity of cells from celiac disease patients to gliadin. Commun. Biol. 2019, 2, 190. [Google Scholar] [CrossRef]
- Barone, M.V.; Zanzi, D.; Maglio, M.; Nanayakkara, M.; Santagata, S.; Lania, G.; Miele, E.; Ribecco, M.T.; Maurano, F.; Auricchio, R.; et al. Gliadin-mediated proliferation and innate immune activation in celiac disease are due to alterations in vesicular trafficking. PLoS ONE 2011, 6, e17039. [Google Scholar] [CrossRef] [Green Version]
- Pietz, G.; De, R.; Hedberg, M.; Sjöberg, V.; Sandström, O.; Hernell, O.; Hammarström, S.; Hammarström, M.L. Immunopathology of childhood celiac disease-Key role of intestinal epithelial cells. PLoS ONE 2017, 12, e0185025. [Google Scholar] [CrossRef]
- Fantini, M.C.; Pallone, F. Cytokines: From gut inflammation to colorectal cancer. Curr. Drug Targets 2008, 9, 375–380. [Google Scholar] [CrossRef]
- Karin, M.; Greten, F.R. NF-kappaB: Linking inflammation and immunity to cancer development and progression. Nat. Rev. Immunol. 2005, 5, 749–759. [Google Scholar] [CrossRef]
- Andrews, C.; McLean, M.H.; Durum, S.K. Cytokine Tuning of Intestinal Epithelial Function. Front. Immunol. 2018, 9, 1270. [Google Scholar] [CrossRef]
- Barone, M.V.; Zimmer, K.P. Endocytosis and transcytosis of gliadin peptides. Mol. Cell Pediatr. 2016, 3, 8. [Google Scholar] [CrossRef] [Green Version]
- Nanayakkara, M.; Lania, G.; Maglio, M.; Auricchio, R.; De Musis, C.; Discepolo, V.; Miele, E.; Jabri, B.; Troncone, R.; Auricchio, S.; et al. P31-43, an undigested gliadin peptide, mimics and enhances the innate immune response to viruses and interferes with endocytic trafficking: A role in celiac disease. Sci. Rep. 2018, 8, 10821. [Google Scholar] [CrossRef] [Green Version]
- Cario, E.; Podolsky, D.K. Differential alteration in intestinal epithelial cell expression of toll-like receptor 3 (TLR3) and TLR4 in inflammatory bowel disease. Infect. Immun. 2000, 68, 7010–7017. [Google Scholar] [CrossRef] [Green Version]
- Strober, W.; Fuss, I.; Mannon, P. The fundamental basis of inflammatory bowel disease. J. Clin. Investig. 2007, 117, 514–521. [Google Scholar] [CrossRef] [Green Version]
- Rose-John, S. Interleukin-6 Family Cytokines. Cold Spring Harb. Perspect. Biol. 2018, 10, a028415. [Google Scholar] [CrossRef] [Green Version]
- Arnauts, K.; Verstockt, B.; Ramalho, A.S.; Vermeire, S.; Verfaillie, C.; Ferrante, M. Ex Vivo Mimicking of Inflammation in Organoids Derived From Patients With Ulcerative Colitis. Gastroenterology 2020, 159, 1564–1567. [Google Scholar] [CrossRef]
- Li, V.S.W. Modelling intestinal inflammation and infection using ’mini-gut’ organoids. Nat. Rev. Gastroenterol. Hepatol. 2021, 18, 89–90. [Google Scholar] [CrossRef]
- Dieterich, W.; Neurath, M.F.; Zopf, Y. Intestinal ex vivo organoid culture reveals altered programmed crypt stem cells in patients with celiac disease. Sci. Rep. 2020, 10, 3535. [Google Scholar] [CrossRef] [Green Version]
- Amaral, G.A.; Alves, J.D.; Honorio-França, A.C.; Fagundes, D.L.; Araujo, G.G.; Lobato, N.S.; Lima, V.V.; Giachini, F.R. Interleukin 1-beta is Linked to Chronic Low-Grade Inflammation and Cardiovascular Risk Factors in Overweight Adolescents. Endocr. Metab. Immune Disord. Drug Targets 2020, 20, 887–894. [Google Scholar] [CrossRef]
- Freire, R.; Ingano, L.; Serena, G.; Cetinbas, M.; Anselmo, A.; Sapone, A.; Sadreyev, R.I.; Fasano, A.; Senger, S. Human gut derived-organoids provide model to study gluten response and effects of microbiota-derived molecules in celiac disease. Sci. Rep. 2019, 9, 7029. [Google Scholar] [CrossRef] [Green Version]
- Galatola, M.; Izzo, V.; Cielo, D.; Morelli, M.; Gambino, G.; Zanzi, D.; Strisciuglio, C.; Sperandeo, M.P.; Greco, L.; Auricchio, R. Gene expression profile of peripheral blood monocytes: A step towards the molecular diagnosis of celiac disease? PLoS ONE 2013, 8, e74747. [Google Scholar] [CrossRef] [Green Version]
- Auricchio, R.; Galatola, M.; Cielo, D.; Amoresano, A.; Caterino, M.; De Vita, E.; Illiano, A.; Troncone, R.; Greco, L.; Ruoppolo, M. A Phospholipid Profile at 4 Months Predicts the Onset of Celiac Disease in at-Risk Infants. Sci. Rep. 2019, 9, 14303. [Google Scholar] [CrossRef]
- Auricchio, R.; Stellato, P.; Bruzzese, D.; Cielo, D.; Chiurazzi, A.; Galatola, M.; Castilljeo, G.; Crespo Escobar, P.; Gyimesi, J.; Hartman, C.; et al. Growth rate of coeliac children is compromised before the onset of the disease. Arch. Dis. Child. 2020, 105, 964–968. [Google Scholar] [CrossRef]
- Bernardo, D.; Garrote, J.A.; Allegretti, Y.; León, A.; Gómez, E.; Bermejo-Martin, J.F.; Calvo, C.; Riestra, S.; Fernández-Salazar, L.; Blanco-Quirós, A.; et al. Higher constitutive IL15R alpha expression and lower IL-15 response threshold in coeliac disease patients. Clin. Exp. Immunol. 2008, 154, 64–73. [Google Scholar] [CrossRef]
- Wang, Y.; DiSalvo, M.; Gunasekara, D.B.; Dutton, J.; Proctor, A.; Lebhar, M.S.; Williamson, I.A.; Speer, J.; Howard, R.L.; Smiddy, N.M.; et al. Self-renewing Monolayer of Primary Colonic or Rectal Epithelial Cells. Cell Mol. Gastroenterol. Hepatol. 2017, 4, 165–182.e7. [Google Scholar] [CrossRef] [Green Version]
- Sato, T.; Vries, R.G.; Snippert, H.J.; van de Wetering, M.; Barker, N.; Stange, D.E.; van Es, J.H.; Abo, A.; Kujala, P.; Peters, P.J.; et al. Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche. Nature 2009, 459, 262–265. [Google Scholar] [CrossRef]
- VanDussen, K.L.; Marinshaw, J.M.; Shaikh, N.; Miyoshi, H.; Moon, C.; Tarr, P.I.; Ciorba, M.A.; Stappenbeck, T.S. Development of an enhanced human gastrointestinal epithelial culture system to facilitate patient-based assays. Gut 2015, 64, 911–920. [Google Scholar] [CrossRef] [Green Version]
Patients | Range Age (Years) | Sex | Biopsy (Marsh Classification *) | Serum AntiTG2 (U/mL) | Anti-Endomysial Antibody (EMA) |
---|---|---|---|---|---|
Controls (N =8) | 10–18 | M = 4, F = 4 | 8 = T0 | 0–1.5 | Negative |
GCD-CD (N = 8) | 3–15 | M = 3, F = 5 | 10 = T3 c 2 = T3 c/b | >50 | Positive |
GFD-CD (N = 5) | 5–18 | F = 5 | 3 = T0 2 = T1 | 0–1.5 | Negative |
POT-CD (N = 4) | 7–12 | M = 2, F = 2 | 3 = M0 1 = T1 | 9.3–57 | 3 = positive 1 = ND |
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Porpora, M.; Conte, M.; Lania, G.; Bellomo, C.; Rapacciuolo, L.; Chirdo, F.G.; Auricchio, R.; Troncone, R.; Auricchio, S.; Barone, M.V.; et al. Inflammation Is Present, Persistent and More Sensitive to Proinflammatory Triggers in Celiac Disease Enterocytes. Int. J. Mol. Sci. 2022, 23, 1973. https://doi.org/10.3390/ijms23041973
Porpora M, Conte M, Lania G, Bellomo C, Rapacciuolo L, Chirdo FG, Auricchio R, Troncone R, Auricchio S, Barone MV, et al. Inflammation Is Present, Persistent and More Sensitive to Proinflammatory Triggers in Celiac Disease Enterocytes. International Journal of Molecular Sciences. 2022; 23(4):1973. https://doi.org/10.3390/ijms23041973
Chicago/Turabian StylePorpora, Monia, Mariangela Conte, Giuliana Lania, Claudia Bellomo, Luciano Rapacciuolo, Fernando Gabriel Chirdo, Renata Auricchio, Riccardo Troncone, Salvatore Auricchio, Maria Vittoria Barone, and et al. 2022. "Inflammation Is Present, Persistent and More Sensitive to Proinflammatory Triggers in Celiac Disease Enterocytes" International Journal of Molecular Sciences 23, no. 4: 1973. https://doi.org/10.3390/ijms23041973
APA StylePorpora, M., Conte, M., Lania, G., Bellomo, C., Rapacciuolo, L., Chirdo, F. G., Auricchio, R., Troncone, R., Auricchio, S., Barone, M. V., & Nanayakkara, M. (2022). Inflammation Is Present, Persistent and More Sensitive to Proinflammatory Triggers in Celiac Disease Enterocytes. International Journal of Molecular Sciences, 23(4), 1973. https://doi.org/10.3390/ijms23041973