Heavy-ion radiation-induced colitis and colorectal carcinogenesis in Il10-/- mice display co-activation of β-catenin and NF-κB signaling

PLOS ONE RESEARCH ARTICLE Heavy-ion radiation-induced colitis and colorectal carcinogenesis in Il10-/- mice display co-activation of β-catenin and NF-κB signaling Shubhankar Suman1*, Bo-Hyun Moon1, Kamal Datta1,2, Bhaskar V. S. Kallakury3, Albert J. Fornace, Jr.1,2 1 Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC, United States of America, 2 Department of Biochemistry and Molecular & Cellular Biology, Georgetown University Medical Center, Washington, DC, United States of America, 3 Department of Pathology, Georgetown University Medical Center, Washington, DC, United States of America a1111111111 a1111111111 a1111111111 a1111111111 a1111111111 OPEN ACCESS Citation: Suman S, Moon B-H, Datta K, Kallakury BVS, Fornace AJ, Jr. (2022) Heavy-ion radiationinduced colitis and colorectal carcinogenesis in Il10-/- mice display co-activation of β-catenin and NF-κB signaling. PLoS ONE 17(12): e0279771. https://doi.org/10.1371/journal.pone.0279771 Editor: Hiroyasu Nakano, Toho University Graduate School of Medicine, JAPAN Received: May 20, 2022 Accepted: December 13, 2022 Published: December 30, 2022 Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. * Abstract Space radiation-induced gastrointestinal (GI) cancer risk models for future interplanetary astronauts are being developed that primarily rely on quantitative animal model studies to assess radiation-quality effects of heavy-ion space radiation exposure in relation to γ-rays. While current GI-cancer risk estimation efforts are focused on sporadic GI-cancer mouse models, emerging in-vivo data on heavy-ion radiation-induced long-term GI-inflammation are indicative of a higher but undetermined risk of GI-inflammation associated cancers, such as colitis-associated cancer (CAC). Therefore, we aimed to assess radiation quality effects on colonic inflammation, colon cancer incidence, and associated signaling events using an in-vivo CAC model i.e., Il10-/- mice. Male Il10-/- mice (8–10 weeks, n = 12/group) were irradiated with either sham, γ-rays or heavy-ions (28Si or 56Fe), and histopathological assessments for colitis and CAC were conducted at 2.5 months post-exposure. qPCR analysis for inflammation associated gene transcripts (Ptges and Tgfb1), and in-situ staining for markers of cell-proliferation (phospho-histone H3), oncogenesis (active-β-catenin, and cyclin D1), and inflammation (phospho-p65NF-κB, iNOS, and COX2) were performed. Significantly higher colitis and CAC frequency were noted after heavy-ion exposure, relative to γ and control mice. Higher CAC incidence after heavy-ion exposure was associated with greater activation of β-catenin and NF-κB signaling marked by induced expression of common downstream inflammatory (iNOS and COX2) and pro-proliferative (Cyclin D1) targets. In summary, IR-induced colitis and CAC incidence in Il10-/- mice depends on radiation quality and display co-activation of β-catenin and NF-κB signaling. Data Availability Statement: All data are included in the manuscript. Funding: This study is supported through NASA grant # NNX09AU95G and 80NSSC22K1279. The funding agency had no role in study planning, execution, data analysis, manuscript preparation, and decision to publish. Competing interests: The authors have declared that no competing interests exist. Introduction Epidemiological studies of A-bomb survivors have demonstrated a greater risk of gastrointestinal (GI) cancer development after low-linear energy transfer (LET) ionizing radiation (IR) such as, γ- or X-rays [1, 2]. Contrary to the established GI-cancer risk of low-LET IR, GI- PLOS ONE | https://doi.org/10.1371/journal.pone.0279771 December 30, 2022 1 / 15 PLOS ONE Abbreviations: IL-10, interleukin-10; CAC, Colitis associated cancer; CRC, colorectal cancer; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; COX2, cyclooxygenase-2; iNOS, inducible nitric oxide synthase; APC, adenomatous polyposis coli; LET, linear energy transfer; IR, ionizing radiation. Heavy-ion radiation-induced colitis and colorectal carcinogenesis cancer risk prediction for astronauts planning to travel to outer space has substantial uncertainty, mainly due to the lack of in-vivo data demonstrating radiation quality effects of the high-LET heavy-ion component of the space radiation [3–5]. Therefore, differential assessment of GI-cancer incidence in animal models exposed to low and high-LET radiation exposure is being conducted to understand radiation quality effects. Earlier differential assessments of GI-cancer incidence have been conducted using sporadic GI-cancer mouse models [6, 7]. Since colitis-associated cancer (CAC) constitutes a significant number of total CRC related mortality [8–10], and emerging data on heavy-ion IR-induced chronic GI inflammation [11, 12], altered microbiome [13, 14], and epithelial barrier function [15] suggest a greater but uncertain risk of colonic inflammation that might contribute to enhanced CAC incidence after heavy-ion exposure. Therefore, studies using inflammation-associated GI-cancer models are warranted to analyze the differential effect of low and high-LET radiation on CAC development. Heavy-ion induced increased levels of pro-inflammatory factors have been attributed to the higher GI cancer incidence [11, 15, 16]. Interestingly, accumulation of pro-inflammatory factors such as cyclooxygenase (COX-1 and 2) and TGFβ1 (transforming growth factor beta-1) in the GI-tract have been reported after heavy-ion exposure [11, 17, 18], as well as during colitis and CAC development [19, 20]. Additionally, COX2 and TGFβ1 are known to activate oncogenic β-catenin signaling and NF-κB signaling, respectively and have been implicated in colitis and CAC colitis development [21, 22]. Once activated, both β-catenin and NF-kB transactivate many common target genes such as Ptges2, Nos2 and Ccnd1 and respective protein levels [COX2, inducible-NOS (iNOS) and Cyclin D1] with established roles in the development of colitis and CAC [23–27]. Furthermore, COX2 is also known to participate in a positive feedback loop leading to a vicious cycle of continuous β-catenin activation and CAC development [28]. The Il10-/- mouse is a well-characterized mouse model to study the progression of CAC and recapitulate progressive colonic inflammation leading to CAC, as observed in humans [29– 33]. In this study, using Il10-/- mice, we demonstrate that exposure to heavy-ion radiation is associated with a higher incidence of colitis and CRC as well as co-activation of β-Catenin and NF-κB signaling. Additionally, this study also emphasizes that colitis and CAC incidence in Il10-/- mice are dependent on radiation quality which has implications (...truncated)