Effect of Caesium-137 chronic low dose exposure on neovascularization

International audienceBackgroundNeovascularization is crucial for tissue irrigation adaptation in response to pathological conditions such as ischemia. This process consists of the formation of new blood vessels either by angiogenesis or by vasculogenesis. Recent studies reported that acute exposure to low doses of ionising radiation, induces an increase of angiogenesis and vasculogenesis (Ministro A, 2016; Lerman O, 2017). Furthermore, nitric oxide is stimulated in these conditions. However, effects of chronic low dose radioelements, particularly Caesium-(Cs)137, one of the most released in the environment; on neovascularization has so far not been investigated. AimWe investigated the effect of chronic low dose contamination with Cs on NO-dependent neovascularization. MethodsC57BL/6 mice received 20KB/L, 100KB/L Cs or vehicle for 6 months in drinking water. 3 other groups received the same treatment, with addition of 500µg/mL L-NAME(LN), a NO synthase blocker. We evaluated angiogenesis with aortic rings sprouting . Moreover unilateral hindlimb femoral artery ligation was performed induce surgical ischemia. Cutaneous blood flow of the ischemic and non-ischemic limb was measured with Laser Doppler Imaging and capillary density was assessed by immunohistochemistry on hindlimb muscles. Vasculogenesis was assessed by measuring the capacity of bone marrow cells differentiation into EPCs and their ability to form tubular structures in vivo in matrigel after subcutaneous injection of SDF-1 in control or mice treated with CsLN. Results/conclusionsOur preliminary results indicated a dose-dependent increase of ischemic/non-ischemic blood flow ratio in Cs-treated mice, as compared to controls. An increased blood flow ratio was also observed in Cs20KB/L -treated as compared to control/LN-treated group. A reduction of blood flow was observed in Cs100KB/L - vs Cs100KB/LN-treated group. These results suggest a dose-dependent stimulation of post-ischemic neovascularization after Cs contamination. Reduction of stimulation in Cs100KB/LN -treated group may be due to LN

Multimodal analysis of small size data to understand the effects of low doses of ionizing radiation on atherosclerosis

National audienceBackground: Atherosclerosis is a hardening and narrowing of arteries. It can put blood flow at risk as arteries become blocked. Mechanistic understanding of the effects of low-dose ionizing radiation (LDIR) on atherosclerosis remains incomplete. The experimental studies have shown a protective effect of LDIR on atherosclerosis in rodent models 1, 2. How early responses in different cell types that are known to be involved in atherosclerosis contribute to this effect is not clear. Challenges in understanding biological mechanisms of LDIR include small size of experimental animal groups, low level changes and data heterogeneity and multimodality. Advanced methods of multimodal data integration, including machine learning algorithms, have proved useful in various biomedical applications but have not been used much in the studies of health effects of LDIR. In this study, we report results of applying a statistical and classical machine learning methods to reveal causal mechanistic links between biological responses of different modalities observed in ApoE-/- mice exposed to low to moderate doses of gamma-radiation. Revealing complex correlations and causal links related to health conditions, such as atherosclerosis, can help advance the concept adverse outcome pathway (AOP).Analysis: Fourteen to 16 weeks old ApoE-/- mice were exposed to a single dose of 0, 0.05, 0.5, or 1 Gy from 137Cs at a dose rate of 10.35 mGy.min-1. Samples for biological assay were collected at 24 hours, 10 days and 100 days post-irradiation. Bone marrow cells from mice sacrificed at 24 hours or 10 days post-exposure were used to grow bone marrow derived macrophages for polarization into M1 or M2 phenotypes. Cells were treated with Il-4 or INFγ for M2 or M1 polarization, respectively. Polarization was validated by measuring the mRNA levels of polarization markers using RT-qPCR. Also, cell culture supernatants of polarized macrophages were harvested and cytokine expression were determined using ELISA. In this study, we focused on a single time point of 24 after irradiation with three modalities of data. Data from flow cytometry, gene expression and ELISA assay originating from the same animal were gathered. To carry out an integral analysis this complex and heterogeneous data obtained at different levels of biological organization, we employed a two-step analysis. In the first step, we used univariate and descriptive statistics analysis (ANOVA, PCA, PLS, RGCCA) and compared it with four classical machine learning methods (SVM, random forest, tree decision and Gradient boosting). In the second step, an integration of the obtained correlations into a biologically relevant mechanistic model that can be used as a basis for an AOP to atherosclerosis after exposure to LDIR was carried out. Results: Our preliminary results confirm the difficulty of visualizing the effects of LDIR. The results of univariate ANOVA analysis, although could test important comparisons, were limited in its ability to reveal a comprehensive integral insight when multiple variables were used. Using PCA allowed to reveal certain hidden trends in the assembly of variables, which was an improvement when compared to ANOVA. PLS and RGCCA were both more efficient in identifying those variables that are responsive to different treatment options and mediate biological phenotypic changes. Despite the low classification rate, machine learning methods allowed to identify those features/variables that contribute to differences between groups. These different analyses helped to reconstruct in more detail the mechanistic and/or causal links between various molecular endpoints measured under various treatment (radiation dose) and phenotypic (M0, M1 or M2 macrophages) conditions. Overall, our results highlight the need for comprehensive data analyses of LDIR studies and provide an example of such analyses using a multitude of statistical methods to understand mechanisms of LDIR effects on atherosclerosis. [1] Ebrahimian TG et al. “Chronic Exposure to External Low-Dose Gamma Radiation Induces an Increase in Anti-inflammatory and Anti-oxidative Parameters Resulting in Atherosclerotic Plaque Size Reduction in ApoE-/- Mice.” Radiat Res. 2018[2] Rey et al. “Exposure to low to moderate doses of ionizing radiation induces a reduction of pro-inflammatory Ly6Chigh monocytes and a U-curved response of T cells in ApoE -/- mice.” Dose-Response.202

Exposure to low to moderate doses of ionizing radiation induces a reduction of pro-inflammatory Ly6Chigh monocytes and a U-curved response of T cells in ApoE -/- mice

International audienceLow dose ionizing radiation (LDIR) is known to have a protective effect on atherosclerosis in rodent studies, but how it impacts different cells types involved in lesion formation remains incompletely understood. We investigated the immunomodulatory response of different doses and dose-rates of irradiation in ApoE mice. Mice were exposed to external γ rays at very low (1.4 mGy.h-1) or low (50 mGy.h-1) dose-rates, with cumulative doses spanning 50 to 1000 mGy. Flow cytometry of circulating cells revealed a significant decrease in pro-inflammatory Ly6CHi monocytes at all cumulative doses at low dose-rate, but more disparate effects at very low dose-rate with reductions in Ly6CHi cells at doses of 50, 100 and 750 mGy only. In contrast, Ly6CLo monocytes were not affected by LDIR. Similarly, proportions of CD4+ T cell subsets in the spleen did not differ between irradiated mice and non-irradiated controls, whether assessing CD25+FoxP3+ regulatory or CD69+ activated lymphocytes. In the aorta, gene expression of cytokines such as IL-1 and TGF-ß and adhesion molecules such as E-Selectin, ICAM-1, and VCAM-1 were reduced at the intermediate dose of 200 mGy. These results suggest that LDIR may reduce atherosclerotic plaque formation by selectively reducing blood pro-inflammatory monocytes and by impairing adhesion molecule expression and inflammatory processes in the vessel wall. In contrast, splenic T lymphocytes were not affected by LDIR. Furthermore, some responses to irradiation were nonlinear; reductions in aortic gene expression were significant at intermediate doses, but not at either highest or lowest doses. This work furthers our understanding of the impact of LDIR with different dose-rates on immune system response in the context of atherosclerosis

Effects of low dose radiation on atherosclerosis in APOE(-/-) mice: study of short term effects on macrophage polarization

International audienceAthersoclerosis is a chronic inflammatory disease of medium and large arteries that can lead to myocardial infarction or stroke. Mechanistic understanding of the effects of low-dose ionizing radiation (LDIR) on atherosclerosis remains incomplete. The experimental studies have shown a protective effect of LDIR on atherosclerosis in rodent models. However early responses of LDIR in different cell types that are known to be involved in atherosclerosis is not clear. The objective is to understand biological mechanisms of LDIR include on animal groups with multimodal approach. In this study, we report results of applying the foldchange, usually considered a relevant criterion for stating difference and similarity between measurements and a multilevel multivariate approach. Revealing complex correlations and causal links related to health conditions, such as atherosclerosis, can help advance the concept adverse outcome pathway (AOP)

Chronic Exposure to External Low-Dose Gamma Radiation Induces an Increase in Anti-inflammatory and Anti-oxidative Parameters Resulting in Atherosclerotic Plaque Size Reduction in ApoE–/– Mice

International audiencePopulations living in radiation-contaminated territories,such as Chernobyl and Fukushima, are chronically exposedto external gamma radiation and internal radionuclidecontamination due to the large amount of137Cs released inthe environment. The effect of chronic low-dose exposure onthe development of cardiovascular diseases remains unclear.Previously reported studies have shown that low-doseradiation exposure could lead to discrepancies accordingto dose rate. In this study, we examined the effect of verylow-dose and dose-rate chronic external exposure onatherosclerosis development. ApoE–/– mice were chronicallyirradiated with a gamma source for 8 months at twodifferent dose rates, 12 and 28 lGy/h, equivalent to doserates measured in contaminated territories, with a cumulativedose of 67 and 157 mGy, respectively. We evaluatedplaque size and phenotype, inflammatory profile andoxidative stress status. The results of this study showed adecrease in plaque sizes and an increase in collagen contentin ApoE–/– mice exposed to 28 lGy/h for 8 months comparedto nonexposed animals. The plaque phenotype was associatedwith an increase in anti-inflammatory and antioxidativegene expression. These results suggest that chroniclow-dose gamma irradiation induces an upregulation oforganism defenses leading to a decrease in inflammationand plaque size. To our knowledge, this is the first study todescribe the possible effect of chronic external very low-doseionizing radiation exposure for 8 months. This work couldhelp to identify the potential existence of a dose threshold,below that which harmful effects are not exhibited andbeneficial effects are potentially observed. Furthermore,these findings permit consideration of the importance ofdose rate in radiation protection

Etude de l’implication du stress dans la réponse aux faibles doses de rayonnements ionisants sur le système cardio et cérébrovasculaire. : Projet SIROCCO

International audienceContexteLes études épidémiologiques mettent en évidence un lien entre l’exposition aux rayonnements ionisants à des doses modérées (>500mGy) et le développement de pathologies cardiovasculaires.Les études expérimentales ne montrent pas de lien entre exposition chronique à très faibles doses (150mGy-300mGy) de rayonnements ionisants et le développement de l’athérosclérose.Le stress est un des premiers facteurs de risque cardiovasculaire. Il a été montré qu’un stress psychosocial est associé à une augmentation de l’oxydation des lipides mais également des cytokines pro-inflammatoires induisant une aggravation des maladies cardiovasculaires.Aucune étude n’a cependant mis en évidence la réponse d’une co-exposition associant rayonnements ionisants et co-facteur de risque tel que le stress sur les pathologies du coeur et des vaisseaux.MéthodePar une approche in vivo nous allons exposer des souris ApoE-/- aux rayonnements ionisants gamma à faibles doses (dose cumulée de 70mGy et 800mGy) chronique (6μGy/h) et exposer dans le même temps au stress. A l’issue de cette co-exposition l’objectif sera d’évaluer : Les paramètres fonctionnels et physiopathologiques du coeur et des vaisseaux Le profil moléculaire des vaisseaux, du coeur et du sang (approches métabolomiques) Les conséquences sur la vascularisation cérébrale (densité vasculaire et l’inflammation cérébrale)RésultatsCe projet est dans une phase de validation du modèle de stress que nous avons initié par l’ajout de litière de rat. A cette étape de l’étude, les résultats ne mettent pas en évidence de perte de poids mais les mesures de pression artérielle et le dosage d’hormones de stress doivent venir compléter l’étude.ConclusionSIROCCO dispose d’atouts majeurs pour progresser vers l’importance des facteurs risques tels que l’hypercholestérolémie et le stress dans la réponse des faibles doses sur des pathologies chroniques comme les pathologies cardiovasculaires. SIROCCO va également permettre par une approche moléculaire à grande échelle de comprendre les mécanismes à court terme et de faire un lien avec d’éventuelles apparitions de troubles fonctionnels à long terme aussi bien chez les animaux mâles que chez les femelles. Ces études vont permettre en complémentarité avec les études épidémiologiques de mieux comprendre les mécanismes impliqués après exposition à faibles doses dans des contextes micro-environnementaux à risque.Mots-clés (5 maximum)Co-expositions, facteurs de risques, rayonnements ionisants, stress, maladies cardiovasculaire

Etude de l’implication du stress dans la réponse aux faibles doses de rayonnements ionisants sur le système cardio et cérébrovasculaire. : Projet SIROCCO

International audienceContexteLes études épidémiologiques mettent en évidence un lien entre l’exposition aux rayonnements ionisants à des doses modérées (>500mGy) et le développement de pathologies cardiovasculaires.Les études expérimentales ne montrent pas de lien entre exposition chronique à très faibles doses (150mGy-300mGy) de rayonnements ionisants et le développement de l’athérosclérose.Le stress est un des premiers facteurs de risque cardiovasculaire. Il a été montré qu’un stress psychosocial est associé à une augmentation de l’oxydation des lipides mais également des cytokines pro-inflammatoires induisant une aggravation des maladies cardiovasculaires.Aucune étude n’a cependant mis en évidence la réponse d’une co-exposition associant rayonnements ionisants et co-facteur de risque tel que le stress sur les pathologies du coeur et des vaisseaux.MéthodePar une approche in vivo nous allons exposer des souris ApoE-/- aux rayonnements ionisants gamma à faibles doses (dose cumulée de 70mGy et 800mGy) chronique (6μGy/h) et exposer dans le même temps au stress. A l’issue de cette co-exposition l’objectif sera d’évaluer : Les paramètres fonctionnels et physiopathologiques du coeur et des vaisseaux Le profil moléculaire des vaisseaux, du coeur et du sang (approches métabolomiques) Les conséquences sur la vascularisation cérébrale (densité vasculaire et l’inflammation cérébrale)RésultatsCe projet est dans une phase de validation du modèle de stress que nous avons initié par l’ajout de litière de rat. A cette étape de l’étude, les résultats ne mettent pas en évidence de perte de poids mais les mesures de pression artérielle et le dosage d’hormones de stress doivent venir compléter l’étude.ConclusionSIROCCO dispose d’atouts majeurs pour progresser vers l’importance des facteurs risques tels que l’hypercholestérolémie et le stress dans la réponse des faibles doses sur des pathologies chroniques comme les pathologies cardiovasculaires. SIROCCO va également permettre par une approche moléculaire à grande échelle de comprendre les mécanismes à court terme et de faire un lien avec d’éventuelles apparitions de troubles fonctionnels à long terme aussi bien chez les animaux mâles que chez les femelles. Ces études vont permettre en complémentarité avec les études épidémiologiques de mieux comprendre les mécanismes impliqués après exposition à faibles doses dans des contextes micro-environnementaux à risque.Mots-clés (5 maximum)Co-expositions, facteurs de risques, rayonnements ionisants, stress, maladies cardiovasculaire

Effect of repetitive potassium iodide on thyroid and cardiovascular functions in elderly rats

International audienceBackground: To date, paediatric thyroid cancer has been the most severe health consequence of the Chernobylaccident, caused by radioactive iodine (131I) aerosol’s dispersion. WHO recommends a single dose of potassiumiodide (KI) to reduce this risk. Following the Fukushima accident, it became obvious that repetitive doses of KImay be necessary due to multiple exposures to 131I. Knowledge about the effects of repeated ITB (Iodine ThyroidBlocking) is scarce and controversial. KI may affect the thyroid hormones synthesis; which is crucial for thecardiovascular function. Furthermore, myocardial and vascular endothelial tissues are sensitizes to subtlechanges at the concentration of circulating pituitary and/or thyroid hormones.Objective: In this preclinical study, we aimed to assess the effects of repeated ITB in elderly male rats.Methods: Twelve months old male Wistar rats were subjected to either KI or saline solution for eight days. Analyses were performed 24 h and 30 days after the treatment discontinuation.Findings: We reported a significant increase (18%) in some urinary parameters related to renal function, a subtledecrease of plasma TSH level, a significant increase (379%) in renin and a significant decrease (50%) in aldosterone upon KI administration. At the molecular level, the expression of thyroid and cardiovascular genes wassignificantly affected by the treatment. However, in our experimental settlement, animal heart rate was notsignificantly affected thirty days after KI discontinuation. ECG patterns did not change after administration of KI,and arrhythmia was not observed in these conditions despite the PR-intervals decreased significantly. Cardiovascular physiology was preserved.Conclusion: Our results indicate that repeated ITB in elderly rats is characterized by molecular modifications ofcardiovascular key actors, particularly the Renin-angiotensin-aldosterone axis with a preserved physiologicalhomeostasis. This new scientific evidence may be useful for the maturation of ITB guidelines especially forelderly sub-population

Effects of repeated potassium iodide administration on genes involved in synthesis and secretion of thyroid hormone in adult male rat

International audienceBackgroundA single dose of potassium iodide (KI) is recommended to reduce the risk of thyroid cancer during nuclear accidents. However in case of prolonged radioiodine exposure, more than one dose of KI may be necessary. This work aims to evaluate the potential toxic effect of repeated administration of KI.MethodsAdult Wistar rats received an optimal dose of KI 1 mg/kg over a period of 1, 4 or 8 days.Resultshormonal status (TSH, FT4) of treated rats was unaffected. Contrariwise, a sequential Wolff-Chaikoff effect was observed, resulting in a prompt decrease of NIS and MCT8 mRNA expression (−58% and −26% respectively), followed by a delayed decrease of TPO mRNA expression (−33%) in conjunction with a stimulation of PDS mRNA expression (+62%).Conclusionwe show for the first time that repeated administration of KI at 1 mg/kg/24h doesn't cause modification of thyroid hormones level, but leads to a reversible modification of the expression of genes involved in the synthesis and secretion of thyroid hormones

Studying biomarkers reflecting the adverse outcome pathways from exposures to diseases using molecular epidemiology: example of the molecular epidemiology protocol developed as part of the Concerted Uranium Research in Europe (CURE) project

International audienceResults produced during recent years have shown that epidemiology still has a strong potential to quantify radiation-related health effects after low dose exposures, even below 100 mGy. However below certain dose levels, which may differ according to the disease considered and population characteristics (e.g.: age at exposure), direct observations of dose-risk relationships by classical epidemiological studies are not possible. Indirect evidence, including subtle changes in various physiological functions and biomarkers (sub-clinical/ pre-pathological) reflecting key events in response to low or even very low doses will need to be determined in order to draw inferences on risk after exposures to very low doses. Such inference will be possible for instance, if very large cohorts, - which can even be non-radiation cohorts - quantify the relationships between (combinations of) these biomarkers or changes in physiological function and disease risks. Such inferences are in line with the so-called “meet-in-the-middle” approach. Following this philosophy, as part of the CURE project which aimed to prepare protocols to study the health effects of chronic uranium exposures at low dose, a rational approach was employed to select biomarkers of exposures and subclinical biomarkers of effects in uranium target organs (kidney, lung, bone, brain) and in systems in which potential effects could be suspected (e.g.: vascular system). The use of non-targeted techniques (omics) is also essential to generate new hypotheses of induced signal cascades resulting finally in disease pathways and disease onset (Adverse Outcome Pathways, AOP). A full molecular epidemiology protocol was then developed, including a questionnaire and other means of data collection to measure potential confounding factors. Such an approach will be useful in order 1) to determine the changes in biomarkers which may be attributed to uranium itself and 2) to better appreciate the relative influence of uranium versus other stressors on these biomarker changes, as part of AOP leading to chronic diseases. 3) These yet hypothetical diseases and their development can subsequently be studied in classical epidemiological approaches (at higher doses) and through well-justified radiobiological experiments. Finally, indirect evidence gained from molecular epidemiology studies will prove useful to quantify risks after very low dose exposures, in addition to experimental studies and classical epidemiology