Preclinical modeling of low energy X-rays radiological burn: Dosimetry study by monte carlo simulations and EPR spectroscopy

Interventional radiology has grown considerably over the last decades and become an essential tool for treatment or diagnosis. This technique is mostly beneficial and mastered but accidental overexposure can occur and lead to the appearance of deterministic effects. The lack of knowledge about the radiobiological consequences for the low-energy X-rays used for these practices makes the prognosis very uncertain for the different tissues. In order to improve the radiation protection of patients and better predict the risk of complications, we implemented a new preclinical mouse model to mimic radiological burn in interventional radiology and performed a complete characterization of the dose deposition. A new setup and collimator were designed to irradiate the hind legs of 15 mice at 30 Gy in air kerma at 80 kV. After irradiation, mice tibias were collected to evaluate bone dose by Electron Paramagnetic Resonance (EPR) spectroscopy measurements. Monte Carlo simulations with Geant4 were performed in simplified and voxelized phantoms to characterize the dose deposition in different tissues and evaluate the characteristics of secondary electrons (energy, path, momentum). 30 mice tibias were collected for EPR analysis. An average absorbed dose of 194.0 ± 27.0 Gy was measured in bone initially irradiated at 30 Gy in air kerma. A bone to air conversion factor of 6.5 ± 0.9 was determined. Inter sample and inter mice variability has been estimated to 13.9%. Monte Carlo simulations shown the heterogeneity of the dose deposition for these low X-rays energies and the dose enhancement in dense tissue. The specificities of the secondary electrons were studied and showed the influence of the tissue density on energies and paths. A good agreement between the experimental and calculated bone to air conversion factor was obtained. A new preclinical model allowing to perform radiological burn in interventional radiology-like conditions was implemented. For the development of new preclinical radiobiological model where the exact knowledge of the dose deposited in the different tissues is essential, the complementarity of Monte Carlo simulations and experimental measurements for the dosimetric characterization has proven to be a considerable asset

The HOXB4 Homeoprotein Promotes the Ex Vivo Enrichment of Functional Human Embryonic Stem Cell-Derived NK Cells

Human embryonic stem cells (hESCs) can be induced to differentiate into blood cells using either co-culture with stromal cells or following human embryoid bodies (hEBs) formation. It is now well established that the HOXB4 homeoprotein promotes the expansion of human adult hematopoietic stem cells (HSCs) but also myeloid and lymphoid progenitors. However, the role of HOXB4 in the development of hematopoietic cells from hESCs and particularly in the generation of hESC-derived NK-progenitor cells remains elusive. Based on the ability of HOXB4 to passively enter hematopoietic cells in a system that comprises a co-culture with the MS-5/SP-HOXB4 stromal cells, we provide evidence that HOXB4 delivery promotes the enrichment of hEB-derived precursors that could differentiate into fully mature and functional NK. These hEB-derived NK cells enriched by HOXB4 were characterized according to their CMH class I receptor expression, their cytotoxic arsenal, their expression of IFNγ and CD107a after stimulation and their lytic activity. Furthermore our study provides new insights into the gene expression profile of hEB-derived cells exposed to HOXB4 and shows the emergence of CD34+CD45RA+ precursors from hEBs indicating the lymphoid specification of hESC-derived hematopoietic precursors. Altogether, our results outline the effects of HOXB4 in combination with stromal cells in the development of NK cells from hESCs and suggest the potential use of HOXB4 protein for NK-cell enrichment from pluripotent stem cells

Une complication rare en implantologie : l’hématome du plancher buccal

L’hématome du plancher buccal est une complication rare pouvant mettre en jeu le pronostic vital. Sa survenue exceptionnelle explique sans doute la difficulté à établir un diagnostic positif. Il peut avoir diverses causes et se traduire dans les premiers stades de sa formation par une symptomatologie proche de celle de l’œdème de Quincke. Une femme de 77 ans a présenté un hématome du plancher buccal suite à la pose de deux implants symphysaires nécessitant une hospitalisation d’urgence

Extracellular vesicles derived from mesenchymal stromal cells mitigate intestinal toxicity in a mouse model of acute radiation syndrome

International audienceAbstract Background Human exposure to high doses of radiation resulting in acute radiation syndrome and death can rapidly escalate to a mass casualty catastrophe in the event of nuclear accidents or terrorism. The primary reason is that there is presently no effective treatment option, especially for radiation-induced gastrointestinal syndrome. This syndrome results from disruption of mucosal barrier integrity leading to severe dehydration, blood loss, and sepsis. In this study, we tested whether extracellular vesicles derived from mesenchymal stromal cells (MSC) could reduce radiation-related mucosal barrier damage and reduce radiation-induced animal mortality. Methods Human MSC-derived extracellular vesicles were intravenously administered to NUDE mice, 3, 24, and 48 h after lethal whole-body irradiation (10 Gy). Integrity of the small intestine epithelial barrier was assessed by morphologic analysis, immunostaining for tight junction protein (claudin-3), and in vivo permeability to 4 kDa FITC-labeled dextran. Renewal of the small intestinal epithelium was determined by quantifying epithelial cell apoptosis (TUNEL staining) and proliferation (Ki67 immunostaining). Statistical analyses were performed using one-way ANOVA followed by a Tukey test. Statistical analyses of mouse survival were performed using Kaplan-Meier and Cox methods. Results We demonstrated that MSC-derived extracellular vesicle treatment reduced by 85% the instantaneous mortality risk in mice subjected to 10 Gy whole-body irradiation and so increased their survival time. This effect could be attributed to the efficacy of MSC-derived extracellular vesicles in reducing mucosal barrier disruption. We showed that the MSC-derived extracellular vesicles improved the renewal of the small intestinal epithelium by stimulating proliferation and inhibiting apoptosis of the epithelial crypt cells. The MSC-derived extracellular vesicles also reduced radiation-induced mucosal permeability as evidenced by the preservation of claudin-3 immunostaining at the tight junctions of the epithelium. Conclusions MSC-derived extracellular vesicles promote epithelial repair and regeneration and preserve structural integrity of the intestinal epithelium in mice exposed to radiation-induced gastrointestinal toxicity. Our results suggest that the administration of MSC-derived extracellular vesicles could be an effective therapy for limiting acute radiation syndrome

Long-term effectiveness of local BM-MSCs for skeletal muscle regeneration: a proof of concept obtained on a pig model of severe radiation burn

BackgroundMedical management of the severe musculocutaneous radiation syndrome involves surgical intervention with debridement of necrotic tissue. Even when skin excision is replaced by specific plastic surgery, treatment of the muscle radiation injury nonetheless remains difficult, for it involves a massive muscle defect in an unpredictable environment, subject to inflammatory waves weeks to months after irradiation, which delay healing and predispose the patient to the development of fibrous scar tissue. In this study, we investigated the long-term effect of local injections of bone marrow-derived mesenchymal stromal cells (BM-MSCs), combined with plastic surgery, to treat muscle necrosis in a large animal model.MethodsThree months after irradiation to the rump, minipigs were treated by excision of necrotic muscle tissue, vascularized flap surgery, and four injections with or without local autologous BM-MSCs, performed weekly. The quality of the muscle wound healing was examined 1year post-surgery.ResultsThe skeletal muscle surgery without MSC treatment led to permanent deposition of collagen 1 and 3, decreased myofiber diameter, failed muscle fiber regeneration, a reduced number of capillaries, and the accumulation of high calcium and fat. In animals treated by surgery and MSC injections, these indicators were substantially better and demonstrated established regeneration. MSC therapy acts at several levels by stimulating growth factors such as VEGF, which is involved in angiogenesis and satellite cell pool maintenance, and creating a macrophage M1/M2 balance.ConclusionThus, cell therapy using BM-MSCs is an effective and safe way to improve recovery of irradiation-induced skeletal muscle damage without signs of long-term degeneration

Analysis of hematopoietic progenitor cells among hEB-derived cells.

<p>(A) Phenotypic analysis of hEB-derived cells. Cells derived from un-co-cultured hEBs or from hEB cells co-cultured with MS-5/SP-HOXB4 and MS-5/EGFP, were analysed by FACS according to CD34, CD43, CD45 and CD45RA antigen expression (one representative experiment out of three). (B) Percentages of cells co-expressing CD34 and CD43, CD45 and CD45RA among total nucleated cells collected at day 14 after EB cells co-cultured with MS-5/SP-HOXB4 or MS-5/EGFP or directly obtained from un-co-cultured hEBs (n = 3, *<i>p</i><0.05). (C) Relative expansion of total CD34⁺ cells. Total CD34⁺ cells were derived from hEB-derived cells co-culture with MS-5/SP-HOXB4 or MS-5/EGFP. Bar represents fold amplification relative to MS-5/EGFP control (designated as 100%) from three independent experiments.</p

NK-cell culture procedure of hESC-derived hematopoietic precursor cells.

<p>Schematic representation of the three steps of NK-cells differentiation from hESCs. hEBs derived from the H1 hESC cell line were dissociated then co-cultured with MS-5/SP-HOXB4 cells or MS-5/EGFP cells as control during 2 weeks. Then, the cells derived from the first step of co-culture were submitted to a second step of 3-week co-culture with unmodified MS-5 cells, in permissive conditions for NK-cell differentiation in presence of SCF, IL-2 and IL-15. NK-cell culture differentiation was conducted directly with un-co-cultured hEB-derived cells as control. (B) Analysis of the presence of the HOXB4 protein within hEB-derived cells co-cultured with either MS-5/SP-HOXB4 (dark line) or MS-5/EGFP (dotted line) stromal cell lines. Data are from one experiment out of two. Gray histogram corresponds to isotypic control. Abbreviations : cy, cytoplasmic.</p

Analysis of NK differentiation potential and NK progenitor cell expansion mediated by HOXB4.

<p>(A) Percentages of NK cells (CD56⁺CD3⁻CD19⁻) among nucleated cells collected at the end of 5 weeks of co-culture. Cells derived from the 2 weeks primary co-cultures with MS-5/SP-HOXB4 or MS-5/EGFP were then plated on unmodified MS-5 cells in conditions known to promote NK-cell differentiation and maintained during three weeks. At the end of the culture period, cells were analyzed by FACS for the expression of CD56 and CD3/CD19 markers. Un-co-cultured hEB cells were directly cultured under NK-cell differentiation conditions for 3 weeks (n = 5, *<i>p</i><0.05, **<i>p</i><0.01). (B) Fold increase of total NK cells. NK cells were derived from total cells isolated from the primary 2-week co-cultures of hEB-derived cells with either MS-5/SP-HOXB4 or MS-5/EGFP control and then cultured under NK-cell differentiation condition for three weeks. NK cells were then numbered. Bars represent fold amplifications relative to day-0 control (un-co-cultured hEBs) (designated as 100%) (n = 5, *<i>p</i><0,05).</p