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. 2020 Nov 5;39:234. doi: 10.1186/s13046-020-01742-4

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© The Author(s) 2020

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 6 — NOX4 is highly expressed in human GBC tissue/stroma, and is associated with poor prognosis. A (IHC, × 200; E = epithelium, S=Stroma). The expression (cytoplasmic and/or nuclear brown staining) of NOX4 in epithelium or stroma of GBC (n = 85) was significantly upregulated compared with gallbladder precancerous (adenomas, n = 10; epithelium: 5.471 ± 0.410 vs. 1.900 ± 0.348, P = 0.007, stroma: 5.965 ± 0.419 vs. 2.000 ± 0.394, P = 0.003) and benign lesions (cholecystitis, n = 10; epithelium: 5.471 ± 0.410 vs. 1.100 ± 0.379; stroma: 5.965 ± 0.419 vs. 1.700 ± 0.396; both P = 0.001), but no statistical difference between precancerous lesions and benign lesions (both P > 0.05). Magnified insets show representative NOX4 staining in stroma. B (CIF staining, × 200). The expression of NOX4 (green) co-localizes with α-SMA and FSP1 (red) positive stroma. Representative samples of NOX4, α-SMA and FSP1 are shown. Sections were counterstained with DAPI. Secondary antibody only controls are shown: anti-goat-alexa-488 for NOX4, anti-mouse-alexa-568 for α-SMA and anti-rabbit-alexa-568 for FSP1. Arrows and inset point to positive staining in fibroblastic cells. C. Kaplan-Meier analysis of GBC patients with high and low NOX4 expression in GBC cells/stroma. The survival time of GBC patients with high NOX4 expression in tumor cells or stroma was significantly shorter than that of GBC patients with low NOX4 expression (log-rank test, c1, GBCs; P = 0.026; c2, GCAFs, P = 0.020)