RESUMEN
In a large retrospective study, we assessed the putative use of circulating microvesicles (MVs), as innovative biomarkers of radiation toxicity in a cohort of 208 patients with prostate adenocarcinoma overexposed to radiation. The level of platelet (P)-, monocyte (M)- and endothelial (E)-derived MVs were assessed by flow cytometry. Rectal bleeding toxicity scores were collected at the time of blood sampling and during the routine follow-up and were tested for association with MVs using a multivariate logistic regression. MVs dosimetric correlation was investigated using dose volume histograms information available for a subset of 36 patients. The number of PMVs was significantly increased in patients with highest toxicity grades compared to lower grades. Risk prediction analysis revealed that increased numbers of PMVs, and an increased amount of MMVs relative to EMVs, were associated with worst rectal bleeding grade compared to the time of blood sampling. Moreover, a significant correlation was found between PMV and MMV numbers, with the range of doses up to the median exposure (40 Gy) of bladder/rectum and anterior rectal wall, respectively. MVs could be considered as new biomarkers to improve the identification of patients with high toxicity grade and may be instrumental for the prognosis of radiation therapy complications.
Asunto(s)
Gastritis , Proctitis , Neoplasias de la Próstata , Traumatismos por Radiación , Recto , Humanos , Masculino , Proctitis/etiología , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/radioterapia , Traumatismos por Radiación/patología , Dosificación Radioterapéutica , Recto/patología , Recto/efectos de la radiación , Estudios RetrospectivosRESUMEN
BACKGROUND: The intestine is particularly sensitive to moderate-high radiation dose and the development of gastrointestinal syndrome (GIS) leads to the rapid loss of intestinal mucosal integrity, resulting in bacterial infiltration, sepsis that comprise patient survival. There is an urgent need for effective and rapid therapeutic countermeasures. The stromal vascular fraction (SVF) derived from adipose tissue is an easily accessible source of cells with angiogenic, anti-inflammatory and regenerative properties. We studied the therapeutic impact of SVF and its action on the intestinal stem cell compartment. METHODS: Mice exposed to the abdominal radiation (18 Gy) received a single intravenous injection of stromal vascular fraction (SVF) (2.5 × 106 cells), obtained by enzymatic digestion of inguinal fat tissue, on the day of irradiation. Mortality was evaluated as well as intestinal regeneration by histological analyses and absorption function. RESULTS: The SVF treatment limited the weight loss of the mice and inhibited the intestinal permeability and mortality after abdominal irradiation. Histological analyses showed that SVF treatment stimulated the regeneration of the epithelium by promoting numerous enlarged hyperproliferative zones. SVF restored CD24+/lysozyme- and Paneth cell populations in the ISC compartment with the presence of Paneth Ki67+ cells. SVF has an anti-inflammatory effect by repressing pro-inflammatory cytokines, increasing M2 macrophages in the ileum and anti-inflammatory monocyte subtypes CD11b+Ly6clowCX3CR1high in the spleen. CONCLUSIONS: Through the pleiotropic effects that contribute to limiting radiation-induced lethality, SVF opens up attractive prospects for the treatment of emergency GIS.
Asunto(s)
Traumatismos por Radiación , Células del Estroma , Tejido Adiposo , Animales , Humanos , Macrófagos , Ratones , Células MadreRESUMEN
Radiation therapy is crucial in the therapeutic arsenal to cure cancers; however, non-neoplastic tissues around an abdominopelvic tumor can be damaged by ionizing radiation. In particular, the radio-induced death of highly proliferative stem/progenitor cells of the colonic mucosa could induce severe ulcers. The importance of sequelae for patients with gastrointestinal complications after radiotherapy and the absence of satisfactory management has opened the field to the testing of innovative treatments. The aim of this study was to use adult epithelial cells from the colon, to reduce colonic injuries in an animal model reproducing radiation damage observed in patients. We demonstrated that transplanted in vitro-amplified epithelial cells from colonic organoids (ECO) of C57/Bl6 mice expressing green fluorescent protein implant, proliferate, and differentiate in irradiated mucosa and reduce ulcer size. To improve the therapeutic benefit of ECO-based treatment with clinical translatability, we performed co-injection of ECO with mesenchymal stromal cells (MSCs), cells involved in niche function and widely used in clinical trials. We observed in vivo an improvement of the therapeutic benefit and in vitro analysis highlighted that co-culture of MSCs with ECO increases the number, proliferation, and size of colonic organoids. We also demonstrated, using gene expression analysis and siRNA inhibition, the involvement of bone morphogenetic protein antagonists in MSC-induced organoid formation. This study provides evidence of the potential of ECO to limit late radiation effects on the colon and opens perspectives on combined strategies to improve their amplification abilities and therapeutic effects.
Asunto(s)
Proteínas Morfogenéticas Óseas/antagonistas & inhibidores , Colon/crecimiento & desarrollo , Células Madre Mesenquimatosas/metabolismo , Organoides/crecimiento & desarrollo , Traumatismos por Radiación/terapia , Animales , Proteínas Morfogenéticas Óseas/metabolismo , Diferenciación Celular/efectos de la radiación , Proliferación Celular/efectos de la radiación , Colon/efectos de la radiación , Células Epiteliales/metabolismo , Células Epiteliales/efectos de la radiación , Proteínas Fluorescentes Verdes/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Membrana Mucosa/patología , Membrana Mucosa/efectos de la radiación , Radiación Ionizante , Factores de TiempoRESUMEN
Skin lesions caused by accidental exposure to radiation or by radiotherapy are a major clinical challenge. We evaluated the effect of bone marrow mononuclear cells (BMMNC) on collagen remodeling and vascular function in radiation-induced skin lesions in the acute and late phases in mice. We studied the effect of BMMNC transplantation in a mouse model of cutaneous radiation injury combining local skin gamma-irradiation and biopsy punch wound. Mice were first irradiated, punched and then BMMNC were intramuscularly administered. Seven days after injury, BMMNC promoted wound healing by (i) increasing re-epithelialization, tissue collagen density and mRNA levels of collagens 1A1, 1A2, and 3A1, and (ii) inhibiting the radiation-induced vascular activation and limiting interactions between leukocytes and the vascular endothelium compared with control. Importantly, BMMNC did not amplify the inflammatory response despite the infiltration of neutrophils and macrophages associated with the expression of IL-6 and MCP-1 mRNAs in the tissue. Remarkably, the beneficial effects of BMMNC therapy on matrix remodeling were maintained for 2 months. Furthermore, BMMNC injection restored vascular function in skin tissue by increasing vascular density and vascular permeability. This therapeutic strategy based on BMMNC injection protects against radiation-induced skin lesions by preventing vascular dysfunction and unfavorable remodeling in the acute and late phases.
Asunto(s)
Trasplante de Médula Ósea/métodos , Cicatrización de Heridas/fisiología , Animales , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Colágeno/metabolismo , Modelos Animales de Enfermedad , Inmunohistoquímica , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Piel/citología , Piel/patologíaRESUMEN
Cutaneous radiation syndrome has severe long-term health consequences. Because it causes an unpredictable course of inflammatory waves, conventional surgical treatment is ineffective and often leads to a fibronecrotic process. Data about the long-term stability of healed wounds, with neither inflammation nor resumption of fibrosis, are lacking. In this study, we investigated the effect of injections of local autologous bone marrow-derived mesenchymal stromal cells (BM-MSCs), combined with plastic surgery for skin necrosis, in a large-animal model. Three months after irradiation overexposure to the rump, minipigs were divided into three groups: one group treated by simple excision of the necrotic tissue, the second by vascularized-flap surgery, and the third by vascularized-flap surgery and local autologous BM-MSC injections. Three additional injections of the BM-MSCs were performed weekly for 3 weeks. The quality of cutaneous wound healing was examined 1 year post-treatment. The necrotic tissue excision induced a pathologic scar characterized by myofibroblasts, excessive collagen-1 deposits, and inadequate vascular density. The vascularized-flap surgery alone was accompanied by inadequate production of extracellular matrix (ECM) proteins (decorin, fibronectin); the low col1/col3 ratio, associated with persistent inflammatory nodules, and the loss of vascularization both attested to continued immaturity of the ECM. BM-MSC therapy combined with vascularized-flap surgery provided mature wound healing characterized by a col1/col3 ratio and decorin and fibronectin expression that were all similar to that of nonirradiated skin, with no inflammation, and vascular stability. In this preclinical model, vascularized flap surgery successfully and lastingly remodeled irradiated skin only when combined with BM-MSC therapy. Stem Cells Translational Medicine 2018:569-582.
Asunto(s)
Trasplante de Células Madre Mesenquimatosas , Traumatismos por Radiación/terapia , Piel/patología , Animales , Células de la Médula Ósea/citología , Tratamiento Basado en Trasplante de Células y Tejidos , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/metabolismo , Proteínas del Choque Térmico HSP47/genética , Proteínas del Choque Térmico HSP47/metabolismo , Metaloproteinasa 2 de la Matriz/genética , Metaloproteinasa 2 de la Matriz/metabolismo , Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Necrosis , Radiación Ionizante , Porcinos , Trasplante Autólogo , Cicatrización de HeridasRESUMEN
Endothelial progenitor cell (EPC) transplantation has beneficial effects for therapeutic neovascularization. We therefore assessed the effect of a therapeutic strategy based on EPC administration in the healing of radiation-induced damage. To improve cell therapy for clinical use, we used pretreatment with ephrin B2-Fc (Eph-B2-Fc) and/or coadministration with smooth muscle progenitor cells. At day 3, EPCs promoted dermal wound healing in both nonirradiated and irradiated mice by 1.2- and 1.15-fold, respectively, compared with animals injected with phosphate-buffered saline. In addition, EPCs also improved skin-blood perfusion and capillary density in both irradiated and nonirradiated mice compared with PBS-injected animals. We also demonstrated that activation with Eph-B2-Fc increased wound closure by 1.6-fold compared with unstimulated EPCs in nonirradiated mice. Interestingly, the beneficial effect of Eph-B2-Fc was abolished in irradiated animals. In addition, we found that Eph-B2-Fc stimulation did not improve EPC-induced vascular permeability or adhesiveness compared to unstimulated EPCs. We hypothesized that this effect was due to high oxidative stress during irradiation, leading to inhibition of EPCs' beneficial effect on vascular function. In this line, we demonstrated that, in irradiated conditions, N-acetyl-l-cysteine treatment restored the beneficial effect of EPC stimulation with Eph-B2-Fc in the wound healing process. In conclusion, stimulation by Eph-B2-Fc improved the beneficial effect of EPCs in physiological conditions and irradiated conditions only in association with antioxidant treatment. Additionally, cotherapy was beneficial in pathological conditions.
Asunto(s)
Células Progenitoras Endoteliales/metabolismo , Miocitos del Músculo Liso/metabolismo , Cicatrización de Heridas/efectos de los fármacos , Anciano , Animales , Efrina-B2 , Femenino , Humanos , Inmunohistoquímica , Masculino , Ratones , Persona de Mediana EdadRESUMEN
Skin exposure to ionizing radiation affects the normal wound healing process. We investigated the beneficial effects of a pharmacological treatment with geranylgeranylacetone (GGA) on keratinocytes using in vitro scratch wound injury assay in nonirradiated and irradiated conditions. Irradiation affected the wound closure of keratinocytes 24 h after scratch injury, whereas re-epithelialization was markedly accelerated after GGA treatment when compared to nontreated keratinocytes. We demonstrated that GGA treatment increased migration of human epidermal keratinocytes and this migratory property was not related to RhoA signaling. Interestingly, Western blot analysis revealed that GGA treatment down-regulated caspase 3 active form expression and up-regulated the activated phenotype by inducing both keratin 6 (K6) expression and interleukin-1ß (IL-1ß) release without modification of the differentiate phenotype. Finally, the proteomic profiling was performed on keratinocytes, showing that global protein changes occurred after irradiation of keratinocytes treated or untreated with GGA.
Asunto(s)
Diterpenos/farmacología , Queratinocitos/efectos de los fármacos , Queratinocitos/efectos de la radiación , Protectores contra Radiación/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/efectos de la radiación , Movimiento Celular/efectos de los fármacos , Movimiento Celular/efectos de la radiación , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/efectos de la radiación , Humanos , Queratinocitos/citología , Queratinocitos/metabolismo , Proteómica , Factores de Tiempo , Proteína de Unión al GTP rhoA/metabolismoRESUMEN
We assessed the role of plasminogen activator inhibitor-1 (PAI-1) and matrix metalloproteinase 9 (MMP9) in wound healing process and in the bone marrow mononuclear cells (BMMNC)-related effects on physiological and pathological wound healing. A full thickness excision wound was created by removal of the skin on the midback of irradiated and nonirradiated animals. Angiogenesis and re-epithelialization were markedly increased in PAI-1-/- mice compared to wild-type (WT) animals. We revealed high MMP activity in tissue of PAI-1-/- animals. Of interest, the wound healing process was reduced in PAI-1-/-:MMP9-/- animals compared to PAI-1-/- mice, suggesting a key role of MMP9 in beneficial effect of PAI-1 deficiency on wound closure. To unravel the role of PAI-1 in BMMNC relative effects, mice were treated with or without local injection of BMMNC isolated from WT, PAI-1-/-, and PAI-1-/-: MMP9-/- animals for 14 days (10(6) cells, n = 6 per group). In WT nonirradiated mice, transplantation of BMMNC isolated from PAI-1-/- animals enhanced wound formation when compared with WT BMMNC. BMMNC differentiation into cells with endothelial phenotype was enhanced by PAI-1 deficiency. These effects were abrogated in PAI-1-/-:MMP9-/- and MMP9-/- BMMNC. In addition, using chimeric mice, we demonstrated that PAI-1 deficiency environment increased the BMMNC-GFP recruitment to the wound site, whereas this effect was abrogated when using PAI-1-/-:MMP9-/- BMMNC. PAI-1 deficiency, at least through MMP9 upregulation, enhanced wound healing and BMMNC therapeutic potential in irradiated and nonirradiated animals.
Asunto(s)
Células de la Médula Ósea/citología , Células de la Médula Ósea/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , Inhibidor 1 de Activador Plasminogénico/metabolismo , Cicatrización de Heridas/fisiología , Animales , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Masculino , Metaloproteinasa 9 de la Matriz/genética , Ratones , Ratones Noqueados , Inhibidor 1 de Activador Plasminogénico/genética , Cicatrización de Heridas/genética , Cicatrización de Heridas/efectos de la radiaciónRESUMEN
About half of people with cancer are treated with radiation therapy; however, normal tissue toxicity still remains a dose-limiting factor for this treatment. The skin response to ionizing radiation may involve multiple inflammatory outbreaks. The endothelium is known to play a critical role in radiation-induced vascular injury. Furthermore, endothelial dysfunction reflects a decreased availability of nitric oxide. Statins have been reported to preserve endothelial function through their antioxidant and anti-inflammatory activities. In this study, wild type and endothelial nitric oxide synthase (eNOS)(-/-) mice were subjected to dorsal skin irradiation and treated with pravastatin for 28 days. We demonstrated that pravastatin has a therapeutic effect on skin lesions and abolishes radiation-induced vascular functional activation by decreasing interactions between leukocytes and endothelium. Pravastatin limits the radiation-induced increase of blood CCL2 and CXCL1 production expression of inflammatory adhesion molecules such as E-selectin and intercellular adhesion molecule-1, and inflammatory cell migration in tissues. Pravastatin limits the in vivo and in vitro radiation-induced downregulation of eNOS. Moreover, pravastatin has no effect in eNOS(-/-) mice, demonstrating that eNOS plays a key role in the beneficial effect of pravastatin in radiation-induced skin lesions. In conclusion, pravastatin may be a good therapeutic approach to prevent or reduce radiation-induced skin damage.
Asunto(s)
Vasos Sanguíneos/fisiopatología , Endotelio Vascular/fisiopatología , Inhibidores de Hidroximetilglutaril-CoA Reductasas/uso terapéutico , Pravastatina/uso terapéutico , Radiodermatitis/prevención & control , Radioterapia/efectos adversos , Piel/irrigación sanguínea , Animales , Vasos Sanguíneos/efectos de los fármacos , Vasos Sanguíneos/efectos de la radiación , Comunicación Celular/efectos de la radiación , Quimiocina CCL2/metabolismo , Quimiocina CXCL1/metabolismo , Modelos Animales de Enfermedad , Selectina E/metabolismo , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/efectos de la radiación , Molécula 1 de Adhesión Intercelular/metabolismo , Leucocitos/efectos de la radiación , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo III/genética , Óxido Nítrico Sintasa de Tipo III/metabolismo , Pravastatina/farmacología , Radiodermatitis/metabolismo , Radiodermatitis/patologíaRESUMEN
Endothelial dysfunction has been implicated in the pathogenesis of atherosclerosis, fibrosis and vascular occlusion after radiation therapy. Statins have been reported to improve endothelial function; however, this beneficial effect on endothelial cells has never been investigated after irradiation. Therefore, using human microvascular endothelial cells from lung that had been irradiated with 5 or 10 Gy, we assessed the effect of pravastatin on endothelial activation by ELISA, cell-ELISA and electrophoretic mobility shift assay and increased blood-endothelial cell interactions by a flow adhesion assay. Pravastatin inhibited the overproduction of monocyte chemoattractant protein 1, IL6 and IL8 and the enhanced expression of intercellular adhesion molecule 1 but had no effect on platelet-endothelial cell adhesion molecule 1 expression. Moreover, pravastatin down-regulated the radiation-induced activation of the transcription factor activator protein 1 but not of nuclear factor-kappaB. Finally, an inhibition by pravastatin of increased adhesion of leukocytes and platelets to irradiated endothelial cells was observed. The effect of pravastatin was maintained up to 14 days after irradiation and was reversed by mevalonate. Pravastatin exerts persistent anti-inflammatory and anti-thrombotic effects on irradiated endothelial cells. Statins may be considered in therapeutic strategies for the management of patients treated with radiation therapy.
Asunto(s)
Células Endoteliales/efectos de la radiación , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Pravastatina/farmacología , Radioterapia/efectos adversos , Trombosis/prevención & control , Arteriosclerosis/tratamiento farmacológico , Arteriosclerosis/etiología , Células Cultivadas , Quimiocina CCL2/biosíntesis , Relación Dosis-Respuesta a Droga , Células Endoteliales/efectos de los fármacos , Células Endoteliales/patología , Humanos , Molécula 1 de Adhesión Intercelular/análisis , Interleucina-6/biosíntesis , Interleucina-8/biosíntesis , Ácido Mevalónico/farmacología , FN-kappa B/antagonistas & inhibidoresRESUMEN
Recovery from hematopoietic aplasia is a predominant factor in the survival of total-body-irradiated mice within 30 days after exposure. However, other radiation-induced pathophysiological events have been shown to play a role, among which an inflammatory reaction must be considered. In the present study, we evaluated the therapeutic potential of a hematopoietic growth factor (thrombopoietin, Tpo) and pleiotropic cytokines (Il4 or Il11), used alone or in combination, on the survival of mice, hematopoietic reconstitution, inflammatory reaction and vascular changes. All treatments including Tpo induced a higher level of survival than did treatment with a placebo, with combinations being the most efficient. The increased survival could not be explained solely by an improved hematopoietic recovery. Treatments with Tpo also reduced the level of the chemokine KC in plasma and the level of expression of mRNA for inflammatory and coagulation proteins in the lungs of irradiated mice. In addition, radiation- induced vascular hyperpermeability was reduced with the use of Tpo. In summary, our results show that Tpo may improve survival by limiting vascular leakage, which in turn could limit inflammatory reactions and the ensuing tissue damage.
Asunto(s)
Interleucina-11/uso terapéutico , Interleucina-4/uso terapéutico , Traumatismos por Radiación/tratamiento farmacológico , Traumatismos por Radiación/prevención & control , Trombopoyetina/uso terapéutico , Enfermedades Vasculares/tratamiento farmacológico , Enfermedades Vasculares/prevención & control , Enfermedad Aguda , Animales , Quimiocina CXCL1 , Quimiocinas , Quimiocinas CXC , Citocinas/sangre , Citocinas/uso terapéutico , Combinación de Medicamentos , Sinergismo Farmacológico , Masculino , Ratones , Ratones Endogámicos C57BL , Tasa de Supervivencia , Resultado del Tratamiento , Enfermedades Vasculares/sangre , Enfermedades Vasculares/patología , Irradiación Corporal TotalRESUMEN
Inflammatory reaction is a classical feature of radiation exposure, and pneumonitis is a dose-limiting complication in the handling of hematological disorders treated with total-body irradiation. In the present study, we first evaluated the inflammatory response in C57BL6/J mice exposed to lethal doses of gamma rays treated with antibiotics or not. Both interleukin 6 and KC (also known as Gro1) were increased in the plasma 10 to 18 days after radiation exposure, independent of bacterial infection, whereas fibrinogen release was linked to a bacterial infection. Furthermore, both Il6 and KC were increased in the lungs of irradiated mice. Our second objective was to characterize the endothelial cell changes in the lungs of total-body-irradiated mice. For this purpose, a quantitative RT-PCR was used to determine the expression of genes involved in inflammatory and coagulation processes. We found that the adhesion molecules P-selectin and platelet endothelial cell adhesion molecule 1 were up-regulated, whereas E-selectin remained unchanged. Tissue factor expression was up-regulated as well, and thrombomodulin gene expression was down-regulated. The investigation by immunohistochemistry of adhesion molecules confirmed the increase in the basal expression of both P-selectin and platelet endothelial cell adhesion molecule 1 on pulmonary endothelial cells. All together, our results suggest the involvement of endothelial cells in the development of radiation-induced inflammatory and thrombotic processes.
Asunto(s)
Coagulación Sanguínea/efectos de la radiación , Células Endoteliales/efectos de la radiación , Inflamación/etiología , Pulmón/efectos de la radiación , Irradiación Corporal Total , Animales , Antibacterianos/farmacología , Quimiocina CXCL1 , Quimiocinas CXC/sangre , Células Endoteliales/fisiología , Fibrinógeno/análisis , Mediadores de Inflamación/metabolismo , Péptidos y Proteínas de Señalización Intercelular/sangre , Interleucina-6/sangre , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Thrombopoietin is the major regulator of platelet production and a stimulator of multilineage hematopoietic recovery following irradiation. The efficacy of three different schedules of thrombopoietin administration was tested on blood cell counts, hematopoietic bone marrow progenitors, and 30-day animal survival in C57BL6/J mice receiving a total body irradiation, with doses ranging from 7 to 10 Gy. A single dose of murine thrombopoietin was injected 2 h before, 2 h after, or 24 h after irradiation. Thrombopoietin promoted multilineage hematopoietic recovery in comparison to placebo up to 9 Gy at the level of both blood cells and bone marrow progenitors, whatever the schedule of administration. The injection of thrombopoietin 2 h before or 2 h after irradiation equally led to the best results concerning hematopoietic recovery. On the other hand, thrombopoietin administration promoted 30-day survival up to 9 Gy with the highest efficacy obtained when thrombopoietin was injected either 2 h before or 2 h after irradiation. However, when its injection was delayed at 24 h, thrombopoietin had almost no effect on survival of 9 Gy irradiated mice. Altogether, our results show that the time schedule for thrombopoietin injection is of critical importance and when thrombopoietin is administered before or shortly after irradiation, it efficiently promotes mice survival to supra-lethal irradiation (up to 9 Gy) in relation with hematopoietic recovery.