{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,21]],"date-time":"2025-11-21T09:24:39Z","timestamp":1763717079674,"version":"3.45.0"},"reference-count":29,"publisher":"MDPI AG","issue":"23","license":[{"start":{"date-parts":[[2025,11,21]],"date-time":"2025-11-21T00:00:00Z","timestamp":1763683200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Diagnostics"],"abstract":"<jats:p>Objectives: Anti-TNF-\u03b1 therapies have transformed the management of Inflammatory Bowel Disease (IBD), yet a substantial proportion of patients fail to respond, highlighting the urgent need for predictive biomarkers. Mucosal TNF-\u03b1 mRNA quantification in fresh biopsies has shown promise but showed several problems for routine clinical use. This study aimed to validate a clinically feasible method for TNF-\u03b1 quantification in formalin-fixed paraffin-embedded (FFPE) tissues and to assess its correlation with established measures of disease activity. Methods: FFPE and matched fresh-frozen biopsies from 54 ulcerative colitis patients were analyzed. Total RNA was extracted from FFPE sections, and TNF-\u03b1 RNA was quantified by RT-qPCR and compared with fresh tissue expression levels. Molecular data were correlated with clinical (pMayo), endoscopic (Mayo Endoscopic Score, MES), and histological (Geboes) indices. Results: TNF-\u03b1 expression in FFPE samples strongly correlated with fresh tissue levels (r = 0.83, p &lt; 0.0001). High TNF-\u03b1 expression in FFPE tissue was significantly associated with active endoscopic mucosal disease (MES \u2265 1; OR 28, 95% CI 3.31\u2013237; p &lt; 0.0001) and with histological inflammation (Geboes \u2265 3.1; OR 0.12, 95% CI 0.02\u20130.59; p = 0.009). Fresh tissue TNF-\u03b1 levels showed similar associations. Clinical parameters such as age, sex, and pMayo score did not significantly correlate with mucosal TNF-\u03b1 expression. RT-qPCR quantification of TNF-\u03b1 in FFPE tissue is a reliable, cost-effective surrogate for fresh biopsy analysis and correlates strongly with endoscopic and histological disease activity. Conclusions: This method offers a practical approach for integrating molecular biomarkers into routine pathology workflows, supporting the implementation of personalized treatment strategies in IBD.<\/jats:p>","DOI":"10.3390\/diagnostics15232946","type":"journal-article","created":{"date-parts":[[2025,11,21]],"date-time":"2025-11-21T08:59:46Z","timestamp":1763715586000},"page":"2946","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["TNF-\u03b1 Quantification in Formalin-Fixed Paraffin-Embedded Tissues as a Predictive Biomarker in Ulcerative Colitis"],"prefix":"10.3390","volume":"15","author":[{"given":"Anna","family":"Viola","sequence":"first","affiliation":[{"name":"Department of Clinical and Experimental Medicine, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-8530-2474","authenticated-orcid":false,"given":"Walter Giuseppe","family":"Giordano","sequence":"additional","affiliation":[{"name":"Department of Biomedical, Dental and Morphological and Functional Imaging Sciences, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rasmus","family":"Goll","sequence":"additional","affiliation":[{"name":"Division of Gastroenterology, Department of Internal Medicine, University Hospital of North Norway, 9019 Troms\u00f8, Norway"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0008-1905-8700","authenticated-orcid":false,"given":"Emanuela","family":"German\u00e0","sequence":"additional","affiliation":[{"name":"Department of Biomedical, Dental and Morphological and Functional Imaging Sciences, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1132-1761","authenticated-orcid":false,"given":"Vincenzo","family":"Fiorentino","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Valeria","family":"Zuccal\u00e0","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0002-1349-0910","authenticated-orcid":false,"given":"Gabriele","family":"Ricciardi","sequence":"additional","affiliation":[{"name":"Department of Biomedical, Dental and Morphological and Functional Imaging Sciences, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-5638-0508","authenticated-orcid":false,"given":"Mariagiovanna","family":"Ballato","sequence":"additional","affiliation":[{"name":"Department of Biomedical, Dental and Morphological and Functional Imaging Sciences, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4957-3283","authenticated-orcid":false,"given":"Pietro","family":"Tralongo","sequence":"additional","affiliation":[{"name":"Department of Biomedical, Dental and Morphological and Functional Imaging Sciences, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Antonio","family":"Ieni","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Guido","family":"Fadda","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Giuseppe","family":"Giuffr\u00e8","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6260-6310","authenticated-orcid":false,"given":"Maurizio","family":"Martini","sequence":"additional","affiliation":[{"name":"Department of Human Pathology of the Adulthood and Developing Age \u201cGaetano Barresi\u201d, Section of Pathology, University of Messina, 98125 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Walter","family":"Fries","sequence":"additional","affiliation":[{"name":"Department of Clinical and Experimental Medicine, University of Messina, 98124 Messina, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,11,21]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"446","DOI":"10.3390\/gidisord6020030","article-title":"Role of IL-33\/ST2 Pathway in Inflammatory Bowel Disease: An Overview and Future Perspectives","volume":"6","author":"Giordano","year":"2024","journal-title":"Gastrointest. Disord."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Caviglia, G.P., Garrone, A., Bertolino, C., Vanni, R., Bretto, E., Poshnjari, A., Tribocco, E., Frara, S., Armandi, A., and Astegiano, M. (2023). Epidemiology of Inflammatory Bowel Diseases: A Population Study in a Healthcare District of North-West Italy. J. Clin. Med., 12.","DOI":"10.3390\/jcm12020641"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"e383","DOI":"10.1097\/MEG.0000000000002107","article-title":"Population-Based Incidence and Prevalence of In-flammatory Bowel Diseases in Milan (Northern Italy), and Estimates for Italy","volume":"33","author":"Crocetti","year":"2021","journal-title":"Eur. J. Gastroenterol. Hepatol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1270","DOI":"10.1016\/j.dld.2019.05.017","article-title":"Prevalence and Incidence of Inflammatory Bowel Dis-ease in Two Italian Islands, Sicily and Sardinia: A Report Based on Health Information Systems","volume":"51","author":"Macaluso","year":"2019","journal-title":"Dig. Liver Dis."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1016\/j.mayocp.2018.09.013","article-title":"Mechanisms of Disease: Inflammatory Bowel Diseases","volume":"94","author":"Ramos","year":"2019","journal-title":"Mayo Clin. Proc."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"2733","DOI":"10.3748\/wjg.v29.i18.2733","article-title":"Study of tumor necrosis factor receptor in the inflammatory bowel disease","volume":"29","author":"Souza","year":"2023","journal-title":"World J. Gastroenterol."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Jang, D., Lee, A.-H., Shin, H.-Y., Song, H.-R., Park, J.-H., Kang, T.-B., Lee, S.-R., and Yang, S.-H. (2021). The Role of Tumor Necrosis Factor Alpha (TNF-\u03b1) in Autoimmune Disease and Current TNF-\u03b1 Inhibitors in Therapeutics. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22052719"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"17562848211059954","DOI":"10.1177\/17562848211059954","article-title":"Tumour necrosis factor inhibitors in inflammatory bowel disease: The story continues","volume":"14","author":"Sandborn","year":"2021","journal-title":"Ther. Adv. Gastroenterol."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Marsal, J., Barreiro-de Acosta, M., Blumenstein, I., Cappello, M., Bazin, T., and Sebastian, S. (2022). Management of Non-Response and Loss of Response to Anti-Tumor Necrosis Factor Therapy in Inflammatory Bowel Disease. Front. Med., 9.","DOI":"10.3389\/fmed.2022.897936"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Fiorino, G., Ananthakrishnan, A., Cohen, R.D., Cross, R.K., Deepak, P., Farraye, F.A., Halfvarson, J., and Steinhart, A.H. (2025). Accelerating Earlier Access to Anti-TNF-\u03b1 Agents with Biosimilar Medicines in the Management of Inflammatory Bowel Disease. J. Clin. Med., 14.","DOI":"10.3390\/jcm14051561"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"284","DOI":"10.1186\/s12967-024-05058-1","article-title":"Predictive biomarkers for anti-TNF alpha therapy in IBD patients","volume":"22","author":"Kumar","year":"2024","journal-title":"J. Transl. Med."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1456","DOI":"10.1053\/j.gastro.2022.01.024","article-title":"Disease Monitoring in Inflammatory Bowel Disease: Evolving Principles and Possibilities","volume":"162","author":"Plevris","year":"2022","journal-title":"Gastroenterology"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Roseira, J., Estevinho, M.M., Gros, B., Marafini, I., Solitano, V., Sousa, P., Carretero, C., Zou, W., Parsa, N., and Charabaty, A. (2025). Advances in Endoscopy in IBD Diagnostics and Management. Best. Pract. Res. Clin. Gastroenterol., 102055.","DOI":"10.1016\/j.bpg.2025.102055"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"222","DOI":"10.1016\/j.cyto.2009.02.001","article-title":"TNF-alpha gene expression in colorectal mucosa as a predictor of remission after induction therapy with infliximab in ulcerative colitis","volume":"46","author":"Olsen","year":"2009","journal-title":"Cytokine"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1237","DOI":"10.1080\/00365521.2023.2217313","article-title":"Estimating tissue-specific TNF mRNA levels prior to anti-TNF\u03b1 treatment may support therapeutic optimisation in IBD patients","volume":"58","author":"James","year":"2023","journal-title":"Scand. J. Gastroenterol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"90","DOI":"10.1016\/j.cyto.2015.12.021","article-title":"Normalization of mucosal tumor necrosis factor-\u03b1: A new criterion for discontinuing infliximab therapy in ulcerative colitis","volume":"79","author":"Olsen","year":"2016","journal-title":"Cytokine"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1312","DOI":"10.1080\/00365520701409035","article-title":"Tissue levels of tumor necrosis factor-alpha correlates with grade of inflammation in untreated ulcerative colitis","volume":"42","author":"Olsen","year":"2007","journal-title":"Scand. J. Gastroenterol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"S340","DOI":"10.1093\/ecco-jcc\/jjab076.438","article-title":"P314 TNF gene activation used as criterion to discontinue biologic therapy\u2014A patient series introducing the NovaPrime TNF kit","volume":"15","author":"Goll","year":"2021","journal-title":"J. Crohns Colitis"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Gao, X.H., Li, J., Gong, H.F., Yu, G.Y., Liu, P., Hao, L.Q., Liu, L.J., Bai, C.G., and Zhang, W. (2020). Comparison of Fresh Frozen Tissue with Formalin-Fixed Paraffin-Embedded Tissue for Mutation Analysis Using a Multi-Gene Panel in Patients with Colorectal Cancer. Front. Oncol., 10.","DOI":"10.3389\/fonc.2020.00310"},{"key":"ref_20","first-page":"e61060","article-title":"Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples","volume":"160","author":"Levin","year":"2020","journal-title":"J. Vis. Exp."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"487","DOI":"10.1093\/jxb\/ern305","article-title":"Normalization of QRT-PCR Data: The Necessity of Adopting a Systematic, Experimental Conditions-Specific, Validation of References","volume":"60","author":"Mauriat","year":"2009","journal-title":"J. Exp. Bot."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1006\/meth.2001.1262","article-title":"Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method","volume":"25","author":"Livak","year":"2001","journal-title":"Methods"},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Viscido, A., Valvano, M., Stefanelli, G., Capannolo, A., Castellini, C., Onori, E., Ciccone, A., Vernia, F., and Latella, G. (2022). Systematic Review and Meta-Analysis: The Advantage of Endoscopic Mayo Score 0 over 1 in Patients with Ulcerative Colitis. BMC Gastroenterol., 22.","DOI":"10.1186\/s12876-022-02157-5"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Principi, M., Contaldo, A., Bianchi, F.P., Losurdo, G., Iannone, A., Ierardi, E., Tafuri, S., Di Leo, A., and Work, I.E.T. (2020). Inter-Observer Agreement of a New Endoscopic Score for Ulcerative Colitis Activity: Preliminary Experience. Diagnostics, 10.","DOI":"10.3390\/diagnostics10040213"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1136\/gut.47.3.404","article-title":"A reproducible grading scale for histological assessment of inflammation in ulcerative colitis","volume":"47","author":"Geboes","year":"2000","journal-title":"Gut J."},{"key":"ref_26","doi-asserted-by":"crossref","unstructured":"Porcu, C., Sideri, S., Martini, M., Cocomazzi, A., Galli, A., Tarantino, G., and Balsano, C. (2018). Oleuropein Induces AMPK-Dependent Autophagy in NAFLD Mice, Regardless of the Gender. Int. J. Mol. Sci., 19.","DOI":"10.3390\/ijms19123948"},{"key":"ref_27","first-page":"763","article-title":"Personalised Therapy in Inflammatory Bowel Disease","volume":"47","author":"Nos","year":"2024","journal-title":"Gastroenterol. Y Hepatol. (Engl. Ed.)"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"20","DOI":"10.1016\/j.anndiagpath.2018.10.011","article-title":"Localization of TNF alpha in ileocolonic biopsies of patients with inflammatory bowel disease","volume":"38","author":"Villanacci","year":"2019","journal-title":"Ann Diagn Pathol."},{"key":"ref_29","doi-asserted-by":"crossref","unstructured":"Pavel, C., Diculescu, M.M., Ilie, M., Plotogea, O.-M., Sandru, V., Enache, V., Gheonea, D.-I., Jichitu, A., Constantinescu, A., and Serban, R.-E. (2025). Immunohistochemistry Analysis in Inflammatory Bowel Disease-Should We Bring to Light Interleukin-10?. Biomedicines, 13.","DOI":"10.3390\/biomedicines13020406"}],"container-title":["Diagnostics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2075-4418\/15\/23\/2946\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,11,21]],"date-time":"2025-11-21T09:19:39Z","timestamp":1763716779000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2075-4418\/15\/23\/2946"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,11,21]]},"references-count":29,"journal-issue":{"issue":"23","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["diagnostics15232946"],"URL":"https:\/\/doi.org\/10.3390\/diagnostics15232946","relation":{},"ISSN":["2075-4418"],"issn-type":[{"type":"electronic","value":"2075-4418"}],"subject":[],"published":{"date-parts":[[2025,11,21]]}}}