Analysis of changes to lncRNAs and their target mRNAs in murine jejunum after radiation treatment.
ABSTRACT: LncRNAs have been reported to play an important role in various diseases. However, their role in the radiation-induced intestinal injury is unknown. The goal of the present study was to analyse the potential mechanistic role of lncRNAs in the radiation-induced intestinal injury. Mice were divided into two groups: Control (non-irradiated) and irradiated. Irradiated mice were administered 14 Gy of abdominal irradiation (ABI) and were assessed 3.5 days after irradiation. Changes to the jejuna of ABI mice were analysed using RNA-Seq for alterations to both lncRNA and mRNA. These results were validated using qRT-PCR. LncRNAs targets were predicted based on analysis of lncRNAs-miRNAs-mRNAs interaction. 29 007 lncRNAs and 17 142 mRNAs were detected in the two groups. At 3.5 days post-irradiation, 91 lncRNAs and 57 lncRNAs were significantly up- and downregulated respectively. Similarly, 752 mRNAs and 400 mRNAs were significantly up- and downregulated respectively. qRT-PCR was used to verify the altered expression of four lncRNAs (ENSMUST00000173070, AK157361, AK083183, AK038898) and four mRNAs (Mboat1, Nek10, Ccl24, Cyp2c55). Gene ontology and KEGG pathway analyses indicated the predicted genes were mainly involved in the VEGF signalling pathway. This study reveals that the expression of lncRNAs was altered in the jejuna of mice post-irradiation. Moreover, it provides a resource for the study of lncRNAs in the radiation-induced intestinal injury.
Project description:Radiation-induced gastric injury is a serious concern that may limit the duration and the delivered dose of radiation. However, the genome-wide molecular changes in stomach upon ionizing radiation have not been reported. In this study, mouse stomach was irradiated with 6 or 12 Gy X-ray irradiation and we found that radiation resulted in the atrophy of gastric mucosa and abnormal morphology of chief and parietal cells. Radiation-induced gastric injury was accompanied by an increase in the serum levels of pepsinogen A and pepsinogen C but not gastrin-17. The expression profiles of messenger RNA (mRNA) and long noncoding RNA (lncRNA) in normal and irradiated gastric tissues were measured by microarray analysis. Results revealed 17 upregulated and 10 downregulated mRNAs were consistent in 6 and 12 Gy irradiated gastric tissues, including D site-binding protein (Dbp) and fibrinogen-like protein 1 (Fgl1). Thirteen upregulated and 96 downregulated lncRNAs were commonly changed in 6 and 12 Gy irradiated gastric tissues. The dysregulated mRNAs were implicated in multiple pathways and showed coexpression with lncRNAs. To identify motifs for transcription factors and coactivators in the proximal promoter regions of the dysregulated RNAs, the bioinformatic tool Biopython was used. A variety of common motifs that are associated with transcription factors were identified, including ZNF263, LMX1B, and Dlx1. Our findings illustrate the molecular changes during radiation-induced gastric injury and the potential transcription factors driving this alteration.
Project description:PURPOSE:To evaluate the acute changes in leukocyte populations after focal irradiation and to assess the role of interleukin 6 (IL-6) in acute and late radiation injury. METHODS AND MATERIALS:Mice were surgically implanted with a radiopaque marker on the surface of the small intestine. Mice were then imaged with cone beam computed tomography to locate the marker and irradiated with 18 Gy of 5 × 5 mm collimated x-rays onto the marked intestine using the Small Animal Radiation Research Platform. Intestinal sections and blood were harvested 1, 3.5, 7, and 14 days and 2 months postirradiation (post-IR) for histology and complete blood count, respectively. Immune cell populations were assessed by immunofluorescence in the acute phase. Collagen deposition was assessed 2 months post-IR. IL-6<sup>-/-</sup> intestinal sections were assessed post-IR for morphology, EdU, Ki67, and TUNEL in comparison to IL-6<sup>+/+</sup> mice. Furthermore, a set of IL-6<sup>+/+</sup> mice were treated with anti-IL-6R to assess the role of IL-6 in late intestinal injury. RESULTS:Intestinal radiation damage peaked 14 days post-IR, and fibrosis had developed by 60 days post-IR. There was a marked infiltration of immune cells into the irradiated intestine, with increased neutrophils, macrophages, B-cells, and CD4<sup>+</sup> T cells maintained from 3.5 to 14 days post-IR. CD8<sup>+</sup> T cells were decreased from days 7 to 14 post-IR. Systemically, leukocytes were increased in the peripheral blood 14 days post-IR with anemia being maintained from 14 days to 2 months. IL-6 was significantly increased in the serum post-IR. IL-6<sup>-/-</sup> mice demonstrated worsened intestinal injury acutely post-IR. Moreover, anti-IL-6R-treated mice presented with worsened intestinal fibrosis 2 months post-IR. CONCLUSIONS:Focal irradiation of the intestine produced a significant increase in immune cells in the irradiated area and systemic inflammation and anemia. Blockade of IL-6 signaling was found to exacerbate acute intestinal injury and late intestinal injury after focal irradiation.
Project description:The role of bone marrow (BM) and BM-derived cells in radiation-induced acute gastrointestinal (GI) syndrome is controversial. Here we use bone marrow transplantation (BMT), total body irradiation (TBI) and abdominal irradiation (ABI) models to demonstrate a very limited, if any, role of BM-derived cells in acute GI injury and recovery. Compared with WT BM recipients, mice receiving BM from radiation-resistant PUMA KO mice show no protection from crypt and villus injury or recovery after 15 or 12?Gy TBI, but have a significant survival benefit at 12?Gy TBI. PUMA KO BM significantly protects donor-derived pan-intestinal haematopoietic (CD45+) and endothelial (CD105+) cells after IR. We further show that PUMA KO BM fails to enhance animal survival or crypt regeneration in radiosensitive p21 KO-recipient mice. These findings clearly separate the effects of radiation on the intestinal epithelium from those on the BM and endothelial cells in dose-dependent acute radiation toxicity.
Project description:<h4>Background</h4>Nuclear accidents and terrorism presents a serious threat for mass casualty. While bone-marrow transplantation might mitigate hematopoietic syndrome, currently there are no approved medical countermeasures to alleviate radiation-induced gastrointestinal syndrome (RIGS), resulting from direct cytocidal effects on intestinal stem cells (ISC) and crypt stromal cells. We examined whether bone marrow-derived adherent stromal cell transplantation (BMSCT) could restitute irradiated intestinal stem cells niche and mitigate radiation-induced gastrointestinal syndrome.<h4>Methodology/principal findings</h4>Autologous bone marrow was cultured in mesenchymal basal medium and adherent cells were harvested for transplantation to C57Bl6 mice, 24 and 72 hours after lethal whole body irradiation (10.4 Gy) or abdominal irradiation (16-20 Gy) in a single fraction. Mesenchymal, endothelial and myeloid population were characterized by flow cytometry. Intestinal crypt regeneration and absorptive function was assessed by histopathology and xylose absorption assay, respectively. In contrast to 100% mortality in irradiated controls, BMSCT mitigated RIGS and rescued mice from radiation lethality after 18 Gy of abdominal irradiation or 10.4 Gy whole body irradiation with 100% survival (p<0.0007 and p<0.0009 respectively) beyond 25 days. Transplantation of enriched myeloid and non-myeloid fractions failed to improve survival. BMASCT induced ISC regeneration, restitution of the ISC niche and xylose absorption. Serum levels of intestinal radioprotective factors, such as, R-Spondin1, KGF, PDGF and FGF2, and anti-inflammatory cytokines were elevated, while inflammatory cytokines were down regulated.<h4>Conclusion/significance</h4>Mitigation of lethal intestinal injury, following high doses of irradiation, can be achieved by intravenous transplantation of marrow-derived stromal cells, including mesenchymal, endothelial and macrophage cell population. BMASCT increases blood levels of intestinal growth factors and induces regeneration of the irradiated host ISC niche, thus providing a platform to discover potential radiation mitigators and protectors for acute radiation syndromes and chemo-radiation therapy of abdominal malignancies.
Project description:Background:Radiation-induced intestinal injury is one of the side effects in patients receiving radiotherapy. The aim of the present study was to investigate the protective effect of XH-103 on radiation-induced small intestinal injury and to explore its mechanism. Methods:C57BL/6N mice were irradiated and treated with XH-103. Firstly, the survival rate of mice exposed to 9.0?Gy and 11.0?Gy total body irradiation (TBI) was examined. Subsequently, at 3.5?d after IR, the small intestinal morphological changes were examined by HE. The numbers of crypt cells, the villus height, the expression of Ki67 and Lgr5, and the apoptotic cells in the intestinal crypts were examined by immunohistochemistry. Furthermore, the expression of p53 and Bax was analyzed by WB. Results:Compared to the irradiation group, XH-103 improved the mice survival rate, protected the intestinal morphology of mice, decreased the apoptotic rate of intestinal crypt cells, maintained cell regeneration, and promoted crypt proliferation and differentiation. XH-103 also reduced the expression of p53 and Bax in the small intestine compared to the IR group. Conclusion:These data demonstrate that XH-103 can prevent radiation-induced intestinal injury, which is beneficial for the protection of radiation injuries.
Project description:Radiation therapy is crucial for curative treatment of lung cancer, which frequently leads to lung injury. Long non-coding RNAs (lncRNAs) are a group of RNAs longer than 200 nucleotides and lack protein-coding capacity. Increasing evidences demonstrate the important roles of lncRNAs in biological processes. However, the mechanism underlying the association of ionizing radiation with alterations in mRNA and lncRNA expression and lung injury remains unclear. In our study, the male Sprague-Dawley (SD) rats were exposed to a dose of 18?Gy of 6 MV X-ray and the transcriptome spectrum was studied. To identify the differentially expressed mRNAs and lncRNAs induced by X-ray, the RNA sequencing data of lung tissues from irradiated and normal rats for 4, 8, and 16 weeks were analyzed, using |log2_ratio|???1 and q???0.05 as thresholds for significantly differential expression. The number of differentially expressed mRNAs was 1097 (686 up- and 411 down-) for 4-week radiotherapy group, 3006 (1935 up- and 1071 down-) for 8-week group and 1838 (1178 up- and 660 down-) for 16-week group. There were 606 (279 up- and 327 down-) differentially expressed lncRNAs in 4-week group, 1715 (831 up- and 884 down-) in 8-week group and 1043 (656 up- and 387 down-) in 16-week group. The differentially expressed mRNAs were mainly involved in cell cycle regulation and Fc receptor pathway, while the lncRNA target genes were significantly enriched in cellular stress response and regulation of cell migration. Moreover, compared with the control group, the irradiated group presented higher tissue specificity of lncRNAs. Radiation-induced lung injury, especially the dynamic network of lncRNAs and mRNAs, is worthy of study. Investigation on the regulatory details of related pathways is significant for the prevention of radiation-related lung injury, as well as the improvement of radiation therapy.
Project description:Radiation enteritis is an old but emerging question induced by the application of radiation. However, no effective drugs for radiation enteritis in clinic. In this study, we found that thymoquinone (TQ) could mitigate intestinal damages induced by irradiation. After exposure to irradiation, TQ-treated improved the irradiated mice survival rate, ameliorated intestinal injury and increased the numbers of intestinal crypts. Furthermore, Lgr5+ ISCs and their daughter cells, including Vil1+ enterocytes, Ki67+ cells and lysozyme+ Paneth cells, were all significantly increased with TQ treatment. In addition, P53, ?H2AX, caspase8, caspase9 and caspase3 expression were all reduced by TQ. Our data showed that TQ modulated DNA damages and decreased the apoptosis in the small intestine. TQ might be used for radiation enteritis treatment.
Project description:Radiation-induced intestinal toxicity is common among cancer patients after radiotherapy. Endothelial cell dysfunction is believed to be a critical contributor to radiation tissue injury in the intestine. Geranylgeranylacetone (GGA) has been used to treat peptic ulcers and gastritis. However, the protective capacity of GGA against radiation-induced intestinal injury has not been addressed. Therefore, we investigated whether GGA affects intestinal damage in mice and vascular endothelial cell damage in vitro. GGA treatment significantly ameliorated intestinal injury, as evident by intestinal crypt survival, villi length and the subsequently prolonged survival time of irradiated mice. In addition, intestinal microvessels were also significantly preserved in GGA-treated mice. To clarify the effect of GGA on endothelial cell survival, we examined endothelial function by evaluating cell proliferation, tube formation, wound healing, invasion and migration in the presence or absence of GGA after irradiation. Our findings showed that GGA plays a role in maintaining vascular cell function; however, it does not protect against radiation-induced vascular cell death. GGA promoted endothelial function during radiation injury by preventing the loss of VEGF/VEGFR1/eNOS signaling and by down-regulating TNF? expression in endothelial cells. This finding indicates the potential impact of GGA as a therapeutic agent in mitigating radiation-induced intestinal damage.
Project description:Radiation-induced lung injury (RILI) can be challenging for thoracic radiotherapy, thus investigating its mechanisms of related pathophysiological process is needed. Long noncoding RNAs (lncRNAs) was found to participate in normal tissue damage induced by ionizing irradiation. Here, we first profiled the dysregulation of lncRNAs, microRNAs (miRNAs), and messenger RNAs (mRNAs) of RILI in mice model receiving 12 Gy thoracic irradiation. The lung tissue was collected 48 hours after irradiation, after which an RNA library was built by RNA sequencing. Compared with the control group, 461 mRNAs and 401 lncRNAs were significantly upregulated, while 936 mRNAs and 501 lncRNAs were significantly downregulated. Then we predicted target miRNAs of the dysregulated lncRNAs and the target mRNAs of these miRNAs. Next, functional annotations of these target mRNAs were performed. Results showed some pathways apparently dysregulated, such as Th1 and Th2 cell differentiation, Th17 cell differentiation, and hematopoietic cell lineage. Through this study, we also highlighted that T helpers could be vital in RILI through lncRNA-miRNA-mRNA network, therefore causing fibrosis, indicating that RNA dysregulation in early stage of RILI may cause severe late complications. Thus, research on the target mechanism and early intervention of lncRNAs with associated competing endogenous RNA network will benefit the treatment of RILI.
Project description:Radiation therapy is a cornerstone of modern management methods for malignancies but is accompanied by diverse side effects. In the present study, we showed that food additives such as polysorbate 80 (P80) exacerbate irradiation-induced gastrointestinal (GI) tract toxicity. A 16S ribosomal RNA high-throughput sequencing analysis indicated that P80 consumption altered the abundance and composition of the gut microbiota, leading to severe radiation-induced GI tract injury. Mice harboring fecal microbes from P80-treated mice were highly susceptible to irradiation, and antibiotics-challenged mice also represented more sensitive to radiation following P80 treatment. Importantly, butyrate, a major metabolite of enteric microbial fermentation of dietary fibers, exhibited beneficial effects against P80 consumption-aggravated intestinal toxicity via the activation of G-protein-coupled receptors (GPCRs) and maintenance of the intestinal bacterial composition in irradiated animals. Moreover, butyrate had broad therapeutic effects on common radiation-induced injury. Collectively, our findings demonstrate that P80 are potential risk factors for cancer patients during radiotherapy and indicate that butyrate might be employed as a therapeutic option to mitigate the complications associated with radiotherapy.