West Asia Organization for Cancer Prevention (WAOCP), APOCP's West Asia Chapter. Asian Pacific Journal of Cancer Prevention 1513-7368 24 9 2023 09 01 Identification of AOC3 and LRRC17 as Colonic Fibroblast Activation Markers and Their Potential Roles in Colorectal Cancer Progression 3099 3107 90794 10.31557/APJCP.2023.24.9.3099 EN Sahira Syamimi Ahmad Zawawi Human Genome Centre, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. 0000-0002-4963-8721 Nur Azlien Shahira Mohd Azram Faculty of Applied Science, Universiti Teknologi MARA, Jengka 26400, Malaysia. Sarina Sulong Human Genome Centre, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. 0000-0001-5369-2707 Andee Dzulkarnaen Zakaria Department of Surgery, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. Yeong Yeh Lee Department of Internal Medicine, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. 0000-0002-6486-7717 Nur Asyilla Che Jalil Department of Pathology, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. 0000-0001-5871-2295 Marahaini Musa Human Genome Centre, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian 16150, Malaysia. 0000-0002-0165-5741 Journal Article 2023 03 20 Background: Accumulation of cancer-associated fibroblasts (CAFs) in the tumor stroma is linked to poor prognosis in colorectal cancer (CRC). CAF-cancer cell interplay, facilitated by secretomes including transforming growth factor-beta 1 (TGF-β1), supports fibroblast activation, drives colorectal carcinogenesis, and contributes to CRC aggressive phenotypes. Although widely used, traditional CAF biomarkers are found to have heterogeneous and non-specific expression. Amine oxidase copper containing 3 (AOC3) and leucine-rich repeat-containing 17 (LRRC17) have been reported to be emerging markers of myofibroblasts. Aim: Our objective was to investigate the potential of AOC3 and LRRC17 as biomarkers for fibroblast activation thus predicting their roles in CRC progression. Methods: Immunofluorescence (IF) staining of AOC3 and LRRC17 was performed on myofibroblast line (CCD-112CoN), primary fibroblasts from colorectal tumor (CAFs), and adjacent normal tissue (normal fibroblasts-NFs). SW620 (epithelial CRC cell line) was used as a control.  Conventional CAF biomarker (alpha-smooth muscle actin - α-SMA) was included in the IF analysis. Fluorescence intensity was compared between groups using ImageJ software. Proliferation and contractility of treated cells were assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and collagen gel contraction assays, respectively. Fibroblast contraction under TGF-β1 treatment was compared to those treated with complete medium (addition of 10% serum) and serum free (SF) medium. Results: Positive AOC3, LRRC17, and α-SMA expression were observed in colonic fibroblasts, more prominent in CAFs, whereas negative staining was found in SW620. Significant downregulation of AOC3, and upregulations in LRRC17 and α-SMA expression was found in TGF-β1-treated fibroblasts compared to SF medium treatment (p-value<0.05). All fibroblasts exhibited higher proliferation in complete medium and under treatment with conditioned medium from SW620 than SF medium. Significant contraction of NFs was recorded in complete medium and TGF-β1 (p-value<0.01). Conclusion: Our results demonstrate AOC3 and LRRC17 as the potential markers of CAF activation which promote CRC progression.  https://journal.waocp.org/article_90794_2e6eff64bd7a42ff5f29656850dbf00d.pdf