Document Type : Original Article(s)

Authors

• Hawra Zia Hossein Al-Jamali ¹
• Farshad Sheikhesmaili ²
• Mohammad Moradad ¹
• Amjad Ahmadi ³
• Ali Jalili ⁴
• Bahram Nikkhoo ^{4, 5}
• Mohammad Reza Rahmani ¹
• Shohreh Fakhari ⁵

¹ Department of Immunology, Faculty of Medicine, Kurdistan University of Medical Sciences, Sanandaj, Iran

² Liver and Digestive Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran

³ Health Metrics and Evaluation Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran

⁴ Cancer and Immunology Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran

⁵ Cellular and Molecular Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran

Abstract

Background: Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD). Neutrophil extracellular traps (NETs) have been recognized as contributing to UC progression. This study aimed to identify key genes driving NET formation in UC and evaluate their potential as biomarkers.
Methods: Gene expression profiles from colon biopsies of UC patients and healthy controls were obtained from the GSE224758 dataset. Differentially expressed genes (DEGs) were identified using the GEO2R tool. Hub genes involved in NET formation, including FCGR3B, AQP9, FPR1, FPR2, and NCF2, were validated through reverse transcription-quantitative polymerase chain reaction (RT-qPCR) using colon biopsy samples from healthy controls (n=20), newly diagnosed UC (n=20), and treatment-resistant UC patients (n=20) collected between November 8, 2022, and February 2, 2025, at Tohid Hospital, Kurdistan University of Medical Sciences, Sanandaj, Iran. Data normality was assessed using the Shapiro–Wilk test; Kruskal–Wallis test and Dunn’s 
post hoc test were applied for group comparisons. ROC analysis was applied to determine the potential of genes for the differentiation of healthy people from patients.
Results: RT-qPCR validation confirmed significantly increased expression of AQP9 (P <0.0001), FPR1, and FPR2 (P=0.03 and P=0.02) genes in patients with UC and those with refractory UC compared to healthy controls. Among these, AQP9 exhibited the most significant differential expression, demonstrating high sensitivity and specificity in distinguishing newly diagnosed and treatment-resistant UC patients from healthy individuals (sensitivity 83.49% and 80.98%, and specificity 67.77% and 64.24%, respectively), indicating its potential as a diagnostic biomarker. 
Conclusion: This study identifies NET-associated genes, particularly AQP9, FPR1, and FPR2, as candidate tissue biomarkers for UC, with AQP9 exhibiting the strongest ability to distinguish patients from healthy controls. These findings support the utility of NET-related genes as tissue biomarkers for UC, while emphasizing that any therapeutic targeting of these molecules lies beyond the scope of the present work and will require additional functional and in vivo validation.

Highlights

Hawra Zia Hossein al-Jamali (Google Scholar)   
Shohreh Fakhari (Google Scholar)   

Keywords

• Colitis, ulcerative
• Neutrophil extracellular traps
• Aquaporin 9
• Gene expression profiling