Bystander effectors of chondrosarcoma cells irradiated at different LET impair proliferation of chondrocytes

While the dose-response relationship of radiation-induced bystander effect (RIBE) is controversial at low and high linear energy transfer (LET), mechanisms and effectors of cell-to-cell communication stay unclear and highly dependent of cell type. In the present study, we investigated the capacity of chondrocytes in responding to bystander factors released by chondrosarcoma cells irradiated at different doses (0.05 to 8 Gy) with X-rays and C-ions. Following a medium transfer protocol, cell survival, proliferation and DNA damages were quantified in bystander chondrocytes. The bystander factors secreted by chondrosarcoma cells were characterized. A significant and major RIBE response was observed in chondrocyte cells (T/C-28a2) receiving conditioned medium from chondrosarcoma cells (SW1353) irradiated with 0.1 Gy of X-rays and 0.05 Gy of C-ions, resulting in cell survivals of 36% and 62%, respectively. Micronuclei induction in bystander cells was observed from the same low doses. The cell survival results obtained by clonogenic assays were confirmed using impedancemetry. The bystander activity was vanished after a heat treatment or a dilution of the conditioned media. The cytokines which are well known as bystander factors, TNF-alpha and IL-6, were increased as a function of doses and LET according to an ELISA multiplex analysis. Together, the results demonstrate that irradiated chondrosarcoma cells can communicate stress factors to non-irradiated chondrocytes, inducing a wide and specific bystander response related to both doses and LET

Altered Response to Total Body Irradiation of C57BL/6-Tg (CAG-EGFP) Mice

Application of green fluorescent protein (GFP) in a variety of biosystems as a unique bioindicator or biomarker has revolutionized biological research and made groundbreaking achievements, while increasing evidence has shown alterations in biological properties and physiological functions of the cells and animals overexpressing transgenic GFP. In this work, response to total body irradiation (TBI) was comparatively studied in GFP transgenic C57BL/6-Tg (CAG-EGFP) mice and C57BL/6 N wild type mice. It was demonstrated that GFP transgenic mice were more sensitive to radiation-induced bone marrow death, and no adaptive response could be induced. In the nucleated bone marrow cells of GFP transgenic mice exposed to a middle dose, there was a significant increase in both the percentage of cells expressing pro-apoptotic gene Bax and apoptotic cell death. While in wild type cells, lower expression of pro-apoptotic gene Bax and higher expression of anti-apoptotic gene Bcl-2, and significant lower induction of apoptosis were observed compared to GFP transgenic cells. Results suggest that presence of GFP could alter response to TBI at whole body, cellular and molecular levels in mice. These findings indicate that there could be a major influence on the interpretation of the results obtained in GFP transgenic mice

Reduced High-Dose Radiation-Induced Residual Genotoxic Damage by Induction of Radioadaptive Response and Prophylactic Mild Dietary Restriction in Mice

Radioadaptive response (RAR) describes a phenomenon in a variety of in vitro and in vivo systems that a low-dose of priming ionizing radiation (IR) reduces detrimental effects of a subsequent challenge IR at higher doses. Among in vivo investigations, studies using the mouse RAR model (Yonezawa Effect) showed that RAR could significantly extenuate high-dose IR-induced detrimental effects such as decrease of hematopoietic stem cells and progenitor cells, acute radiation hematopoietic syndrome, genotoxicity and genomic instability. Meanwhile, it has been demonstrated that diet intervention has a great impact on health, and dietary restriction shows beneficial effects on numerous diseases in animal models. In this work, by using the mouse RAR model and mild dietary restriction (MDR), we confirmed that combination of RAR and MDR could more efficiently reduce radiogenotoxic damage without significant change of the RAR phenotype. These findings suggested that MDR may share some common pathways with RAR to activate mechanisms consequently resulting in suppression of genotoxicity. As MDR could also increase resistance to chemotherapy and radiotherapy in normal cells, we propose that combination of MDR, RAR, and other cancer treatments (i.e., chemotherapy and radiotherapy) represent a potential strategy to increase the treatment efficacy and prevent IR risk in humans

High LET Radiation Overcomes in vitro Resistance to X-rays of Chondrosarcoma Cell Lines

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Characterization of homologous recombination induced by replication inhibition in mammalian cells

To analyze relationships between replication and homologous recombination in mammalian cells, we used replication inhibitors to treat mouse and hamster cell lines containing tandem repeat recombination substrates. In the first step, few double-strand breaks (DSBs) are produced, recombination is slightly increased, but cell lines defective in non-homologous end-joining (NHEJ) affected in ku86 (xrs6) or xrcc4 (XR-1) genes show enhanced sensitivity to replication inhibitors. In the second step, replication inhibition leads to coordinated kinetics of DSB accumulation, Rad51 foci formation and RAD51-dependent gene conversion stimulation. In xrs6 as well as XR-1 cell lines, Rad51 foci accumulate more rapidly compared with their respective controls. We propose that replication inhibition produces DSBs, which are first processed by the NHEJ; then, following DSB accumulation, RAD51 recombination can act

Central role of spatial ROS distribution at the nanometric scale in the molecular response to carbon ion irradiation

International audienceBackground: Hadrontherapy is an alternative to radiotherapy in the treatment of Head-and-Neck cancers (HNSCC), because of accurate ballistic and high biological efficiency, even in hypoxic tumor areas. These cancers are of poor prognosis because of a high risk of recurrence related to the presence of cancer stem cells (CSCs) located in hypoxic niches.Aim of this work was to determine the molecular specificities of the response to carbon-ion irradiation versus photons in HNSCC cancer cell lines and their CSCs’ subpopulation, under hypoxic and normoxic conditions.Methods : SQ20B, FaDu cells, and their CSCs were irradiated with photons or carbon-ion (290MeV/n,NIRS) in normoxia or hypoxia (1%O2). Cell survival curves, expression of HIF-1a reactive oxygen species (ROS) and Migration/Invasion processes were quantified.Results/Conclusions: For CSCs and non-CSCs, an oxygen-enhancement-ratio (OER) upper than 1.2 was measured in response to photons, associated with stabilization of HIF-1α. This stabilization, depending on the ROS production, appears earlier in CSCs. Inhibition of HIF-1α expression results in decreased survival in HNSCC-CSCs after both type of radiation under hypoxia associated with a significant increase in residual DNA-DSBs. Furthermore, a relationship is demonstrated between HIF-1α expression and the DSBs’ detection and repair by the Homologous-Recombination. Finally, the dense and homogeneous ROS production induced by photons, essential for HIF-1α stabilization, leads to the activation of the 3 major epithelio-mesenchymal transition (EMT) signaling pathways (STAT3,MEK/p38/JNK,Akt/mTOR). At the opposite, the ROS concentrated into the carbon ion tracks are insufficient to activate HIF-1α and the upstream EMT pathways.All these results, supported by Monte-Carlo simulations, converge towards the central role of spatial ROS distribution at the nanometric scale to explain the specificities of the molecular response to carbon ions. Their therapeutic advantage may result both from unrepaired complex-DNA lesions and the non-activation of ROS-dependent-signaling pathways involved in tumor cell defense.Supported by Labex PRIMES-ANR-11-LABX-0063;ANR-11-IDEX-0007;ITMO-Cancer-AVIESA

A PARADIGM TO EXPLAIN THE DIFFERENTIAL EFFECTS OF CARBON ION AND X-RAY IRRADIATION ON TUMOR CELLS

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Enhanced effects of chronic restraint-induced psychological stress on total body Fe-irradiation-induced hematopoietic toxicity in Trp53-heterozygous mice

Humans would be exposed to both psychological stress (PS) and radiation in some scenarios such as manned deep-space missions and radiotherapy. It is of great concern to verify possible enhanced deleterious effects from such concurrent exposure. Pioneer studies showed that chronic restraint-induced PS (CRIPS) could attenuate Trp53 functions and increase gamma-ray-induced carcinogenesis in Trp53-heterozygous mice while CRIPS did not significantly modify the effects on X-ray-induced hematopoietic toxicity in Trp53 wild-type mice. As high-linear energy transfer (LET) radiation is the most important component of space radiation in causing biological effects and is practically applied in hadrontherapy, we further investigated the effects of CRIPS on high-LET iron-particle radiation (Fe)-induced hematopoietic toxicity in Trp53-heterozygous mice. Results showed that CRIPS alone could hardly induce significant alteration in hematological parameters (peripheral hemogram and micronucleated erythrocytes in bone marrow) while Fe alone could lead to significant alteration. Meanwhile, concurrent exposure always caused elevated micronucleated incidence. The incidence was significantly higher in animals exposed to both CRIPS and 0.1 Gy Fe compared to that in animals exposed to 0.1 Gy Fe alone. These findings indicated that CRIPS could enhance the deleterious effects of high-LET radiation, particularly at a low dose, on the hematopoietic toxicity in Trp53-heterozygous mice.Keywords: chronic restraint-induced stress; total-body irradiation; iron-particle radiation; peripheral hemogram; bone marrow micronucleated erythrocytes; mouse restraint mode

ROS stress resets circadian clocks to coordinate pro-survival signals.

Dysfunction of circadian clocks exacerbates various diseases, in part likely due to impaired stress resistance. It is unclear how circadian clock system responds toward critical stresses, to evoke life-protective adaptation. We identified a reactive oxygen species (ROS), H2O2 -responsive circadian pathway in mammals. Near-lethal doses of ROS-induced critical oxidative stress (cOS) at the branch point of life and death resets circadian clocks, synergistically evoking protective responses for cell survival. The cOS-triggered clock resetting and pro-survival responses are mediated by transcription factor, central clock-regulatory BMAL1 and heat shock stress-responsive (HSR) HSF1. Casein kinase II (CK2) -mediated phosphorylation regulates dimerization and function of BMAL1 and HSF1 to control the cOS-evoked responses. The core cOS-responsive transcriptome includes CK2-regulated crosstalk between the circadian, HSR, NF-kappa-B-mediated anti-apoptotic, and Nrf2-mediated anti-oxidant pathways. This novel circadian-adaptive signaling system likely plays fundamental protective roles in various ROS-inducible disorders, diseases, and death