Transcriptome analysis to decipher molecular mechanism in radiation-induced lung fibrosis in DNA-PKcs knockout mouse model
Ontology highlight
ABSTRACT: The potential mechanisms of DNA-PKcs and its related signaling pathways in radiation-induced pulmonary toxicity is unclear. The current study utilized genetic engineering DNA-PKcs knockout mouse model, to investigate the molecular mechanisms after dose-response exposure of of the fractionated low-LET photon and high-LET carbon-ion exposure to the whole thorax.
Project description:Normal lung tissue tolerance constitutes a limiting factor in delivering the required dose of radiotherapy to cure thoracic and chest wall malignancies. Patient genetic predisposition, the volume of irradiated lung and combination regimens consisting of concurrent chemotherapy are correlated with increased risk of radiation induced toxicity in lung. The main purpose of this study is to investigate dose-response regulations of mouse lung irradiation based on a comprehensive dose-escalation program, for a better understanding of molecular mechanism governing radiation induced lung fibrosis by high-LET carbon-ions versus conventional low-LET X-ray.
Project description:The time factor in the development of radiation induced lung fibrosis is important but not well characterized so far. This study was to investigate the time series of acute-, subacute-, early and late- timepoints after exposure with low-LET photons versus proton versus and high-LET carbon-ions. The role of CTGF inhibitor in modulating inflammation related signaling pathways were also studied at the acute timepoints. The potential mechanisms underlining the time-dependent progression of inflammatory and fibrotic response is to be illustrated in the present study.
Project description:Endostatin is a naturally occurring 183-amino acid proteolytic fragment of collagen XVIII that localizes in the basement membrane around blood vessels. The anti-tumor properties of this protein have been extensively described, demarcating endostatin as an endogenous inhibitor of angiogenesis. Further, it supresses many signaling cascades such as pro-inflammatory NF-κB, coagulation and adhesion cascades. Yamaguchi et al. reported that endostatin via its C-terminal domain (E4 peptide) has elicit anti-fibrosis effects. However, the zinc binding domain has been previously confined to the N terminus (endostatin mP1 peptide) and was critical to numerous functions of the molecule. The present study aimed to better understand the impact of oligomerization (Fc-Endostatin) as well as N- vs. C-terminal fragments of endostatin (mP1, CE4) on modulating radiation-induced lung fibrosis. Mice were treated with Fc-endostatin (Fc-Endo), N-terminus endostatin peptide (mP1) or C-terminus endostatin E4 peptide (CE4) combined with photon 20 Gy or carbon-ions 12.5 Gy whole thoracic irradiation
Project description:The environment outside the Earth’s protective magnetosphere is a much more threatening and complex space environment. The dominant causes for radiation exposure, solar particle events and galactic cosmic rays, contain high-energy protons. In space, astronauts need healthy and highly functioning cognitive abilities, of which the hippocampus plays a key role. Therefore, understanding the effects of 1H exposure on hippocampal-dependent cognition is vital for de-veloping mitigative strategies and protective countermeasures for future missions. To investi-gate these effects, we subjected 6-month-old female CD1 mice to 0.75 Gy fractionated 1H (250 MeV) whole-body irradiation at the NASA Space Radiation Laboratory. The cognitive perfor-mance of the mice was tested 3 months after irradiation using Y-maze and morris water maze tests. Both sham-irradiated and 1H-irradiated mice significantly preferred exploration of the novel arm compared to the familiar and start arms, indicating intact spatial and short-term memory. Both groups statistically spent more time in the target quadrant, indicating spatial memory retention. There were no significant differences in neurogenic and gliogenic cell counts after irradiation. In addition, proteomic analysis revealed no significant upregulation or down-regulation of proteins related to behavior, neurological disease, or neural morphology. Our data suggests 1H exposure does not impair hippocampal-dependent spatial or short-term memory in female mice.
Project description:The aim of our study is to investigate and compare the effects of carbon and photon irradiation on microvascular endothelial cells. Therefore we irradiated human pulmonary microvascular endothelial cells (HPMEC) with either 2Gy Carbon or 6Gy Photon (bioequivalent doses) and performed microarray analysis both 2 hours (short-term effect) and 6 days (long-term effects) after irradiation. All experiments were performed in 3 biological replicates.
Project description:We examined molecular responses using transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy, 1 GeV proton (1H) and 15 cGy, 1 GeV/nucleon (n) proton (56Fe) particles 1, 3, 7, 14 and 28 days after exposure. Unsupervised clustering analysis of gene expression segregated samples according to the radiation (IR) response, and time after exposure with 56Fe-IR showing the greatest level of gene modulation. 1H-IR exposures showed little differential transcript modulation. Network analysis categorized the major differentially expressed genes into cell cycle, oxidative responses and transcriptional regulation functional groups. Transcriptional networks identified key nodes regulating expression. Individual transcription factors were inferred to be active at 1, 3, 7, 14 and 28 days after exposure. Validation of the signal transduction network by protein analysis showed that particle IR clearly regulates a long lived signaling mechanism for p38 MAPK signaling and NFATc4 activation. Electrophoresis mobility shift assays supported the role of additional key transcription factors GATA-4, STAT-3 and NF-κB as regulators of the response at specific time points. These data suggest that the molecular response to 56Fe-IR is unique and shows long-lasting gene expression in cardiomyocytes, up to 28 days after exposure. Additionally, proteins involved in signal transduction and transcriptional activation via DNA binding play a role in the response to high charge (Z) and energy (E) particles (HZE). Our study may have implications for NASAâs efforts to develop heart disease risk estimates for astronauts safety via identification of specific HZE-IR molecular markers and for patients receiving conventional and particle radiotherapy. Transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy, 1 GeV proton (1H) and 15 cGy, 1 GeV/nucleon (n) proton (56Fe) particles 1, 3, 7, 14 and 28 days after exposure.
Project description:We examined molecular responses using transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy, 1 GeV proton (1H) and 15 cGy, 1 GeV/nucleon (n) iron (56Fe) particles 1, 3, 7, 14 and 28 days after exposure. Unsupervised clustering analysis of gene expression segregated samples according to the radiation (IR) response, and time after exposure with 56Fe-IR showing the greatest level of gene modulation. 1H-IR exposures showed little differential transcript modulation. Network analysis categorized the major differentially expressed genes into cell cycle, oxidative responses and transcriptional regulation functional groups. Transcriptional networks identified key nodes regulating expression. Individual transcription factors were inferred to be active at 1, 3, 7, 14 and 28 days after exposure. Validation of the signal transduction network by protein analysis showed that particle IR clearly regulates a long lived signaling mechanism for p38 MAPK signaling and NFATc4 activation. Electrophoresis mobility shift assays supported the role of additional key transcription factors GATA-4, STAT-3 and NF-κB as regulators of the response at specific time points. These data suggest that the molecular response to 56Fe-IR is unique and shows long-lasting gene expression in cardiomyocytes, up to 28 days after exposure. Additionally, proteins involved in signal transduction and transcriptional activation via DNA binding play a role in the response to high charge (Z) and energy (E) particles (HZE). Our study may have implications for NASAâ??s efforts to develop heart disease risk estimates for astronauts safety via identification of specific HZE-IR molecular markers and for patients receiving conventional and particle radiotherapy. Transcriptome profiling in isolated left ventricular murine cardiomyocytes to 90 cGy, 1 GeV proton (1H) and 15 cGy, 1 GeV/nucleon (n) iron (56Fe) particles 1, 3, 7, 14 and 28 days after exposure.
Project description:Carbon-ion irradiation is an emerging therapeutic option for several tumor entities including lung cancer. Well oxygenated tumor areas compared to a hypoxic environment favor therapeutic photon irradiation efficiency of solid tumors due to increased amounts of DNA damage. The resistance of hypoxic tumor areas towards photon irradiation is enhanced through increased HIF-1 signaling. Here, we compared the effects of oxygen and HIF 1 after photon and carbon-ion irradiation with biological equivalent doses in a human non-small lung cancer model. In hypoxia compared to normoxia, A549 and H1299 cells displayed improved survival after photon irradiation. Knockdown of HIF-1M-NM-1 combined with photon irradiation synergistically delayed tumor growth in vivo. Photon irradiation induced HIF-1M-NM-1 and several of its target genes such as PDK1, GLUT-1, LDHA, and VEGF with subsequent enhanced tumor angiogenesis in vivo, a signaling cascade that was not targeted by carbon-ion irradiation. We present evidence that photons but not carbon-ions induce HIF-1M-NM-1 via mTOR pathway. Importantly, after carbon-ion irradiation in vivo, we observed substantial downregulation of HIF-1M-NM-1 and a drastically delayed tumor growth indicating a considerable higher relative biological effectiveness (RBE) than anticipated from the cell survival data. In sum, our results demonstrate that carbon-ions mediate an improved therapeutic response of tumor treatment compared to photon irradiation that is independent of cell oxygenation and HIF-1 signaling. 16 independent cell cultures were used. Each culture was split into an irradiated and a control plate, yieldin a total of 16 paired samples. Paired samples were analysed in 16 two-color hybridizations. Factors time (after irradiation) with levels 1h and 4h and factor radiation quality with levels C12 and X-rays were analyzed. Each of the 2x2 combinations was analyzed in 4 independent experiments.
Project description:The aim of our study is to investigate the effects of carbon ion and photon irradiation on A549 tumor cells and analyse how these effects are altered by PML knockdown. Therefore we created PML knockdown A549 cells (shPML) and irradiated them with either 2Gy carbon ion or 6Gy Photon (bioequivalent doses). 4 days after irradiation microarray analysis was performed. All experiments were performed in 3 biological replicates and control groups were transduced with an empty vector.