Colon layer and histological phenotype influence mRNA and miRNA expression in colon samples of the DSS mouse model

Unkovič, Ana (author), Belič, Aleš (author), Boštjančič, Emanuela (author), Mrak, Tanja (author), Perše, Martina (author)

Spatial heterogeneity is increasingly recognized as a major determinant of molecular responses in inflammatory bowel disease, yet molecular analyses in dextran sulfate sodium (DSS)-induced colitis commonly rely on whole-colon measurements that may obscure localized biological processes. We investigated how colon layer, histological phenotype, and colitis phase influence RNA expression by combining archived formalin-fixed paraffin-embedded (FFPE) colon tissue, histology-guided laser microdissection (LMD), targeted RT-qPCR, and multivariate modelling. Expression of five mRNAs (Tnf, Tnfr1, Tnfr2, Duox2, and iNos) and three miRNAs (miR-223-3p, miR-181a-5p, and miR-3473b) was analyzed in whole-colon sections, individual colon-wall layers, and histologically defined mucosal microenvironments. RNA expression varied according to anatomical layer, histological phenotype, and colitis phase, with the most pronounced molecular changes observed in erosive mucosa. Although several RNAs showed similar overall trends, whole-section measurements did not consistently reflect the magnitude or spatial distribution of RNA expression observed in individual layers and histological microenvironments. Multivariate modelling further revealed context-dependent interrelationships among the analyzed mRNAs and miRNAs and enabled these molecular patterns to be evaluated in relation to histological, anatomical, and temporal variables. These findings demonstrate that spatial and histological context can substantially influence the interpretation of targeted molecular measurements in DSS colitis and that whole-colon analysis may miss spatially restricted molecular changes. Histology-guided analysis of archived FFPE tissue, combined with multivariate modelling, therefore provides a complementary approach for resolving tissue heterogeneity and extracting additional biological information from existing animal tissue repositories, potentially reducing the need for additional animal experiments when appropriate archived material is available.

• Language: English
• Keywords: DSS-induced colitis, formalin-fixed paraffin-embedded tissue, laser microdissection, spatial heterogeneity, histology-guided analysis, multivariate modelling, gene expression, miRNA
• Work type: Article
• Typology: 1.01 - Original Scientific Article
• Organization: MF - Faculty of Medicine
• Publication status: Published
• Publication version: Version of Record
• Year: 2026
• Number of pages: Str. 1-25
• Numbering: Vol. 15, iss. 8, art. 1677
• PID: 20.500.12556/RUL-188845
• UDC: 616-092
• ISSN on article: 2073-4409
• DOI: 10.3390/cells15181677
• COBISS.SI-ID: 292287235
• Publication date in RUL: 29.09.2026

Record is a part of a journal

• Title: Cells
• Shortened title: Cells
• Publisher: MDPI
• ISSN: 2073-4409
• COBISS.SI-ID: 519958809

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.

Secondary language

• Language: Slovenian
• Keywords: z DSS izzvani kolitis, FFPE tkivo, laserska mikrodisekcija, prostorska heterogenost, histološko vodena analiza, multivariatno modeliranje, mikroRNA