RESUMEN
This study evaluates the extravasation pathways of circulating macromolecules in a rat glioma model (RG2) which was observed by both magnetic resonance imaging using ultrasmall superparamagnetic iron oxide and electron microscopy. Although magnetic resonance imaging signal enhancement was observed as soon as 10 min after injection (9.4% 2 h after injection), electron microscopy showed that endothelial cells were still tightly sealed. However, circulating immunoglobulin G and ultrasmall superparamagnetic iron oxide were found in large membrane compartments of endothelial cells, in the basal lamina (7.4 ± 1.2 gold particles/µm2 in the tumor versus 0.38 ± 0.17 in healthy tissue, p = 1.4.10-5) and between tumoral cells. Altogether, this strongly suggests an active transport mediated by macropinocytosis. To challenge this transport mechanism, additional rats were treated with amiloride, an inhibitor of macropinocytosis, leading to a reduction of membrane protrusions (66%) and of macropinosomes. Amiloride however also opened tumoral tight junctions allowing a larger extravasation of ultrasmall superparamagnetic iron oxide (magnetic resonance imaging signal enhancement of 35.7% 2 h after injection). Altogether, these results suggest that ultrasmall superparamagnetic iron oxide and immunoglobulin G in the RG2 glioma model follow an active extravasation pathway mediated by a macropinocytosis process. Amiloride also appears as a potential strategy to facilitate the extravasation of chemotherapeutic drugs in glioma.
Asunto(s)
Barrera Hematoencefálica/fisiología , Neoplasias Encefálicas/irrigación sanguínea , Permeabilidad Capilar/fisiología , Glioma/irrigación sanguínea , Imagen por Resonancia Magnética/métodos , Pinocitosis/fisiología , Uniones Estrechas/fisiología , Animales , Barrera Hematoencefálica/diagnóstico por imagen , Barrera Hematoencefálica/patología , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/fisiopatología , Línea Celular Tumoral , Óxido Ferrosoférrico , Glioma/diagnóstico por imagen , Glioma/patología , Glioma/fisiopatología , Nanopartículas de Magnetita , Trasplante de Neoplasias , Ratas Endogámicas F344RESUMEN
BACKGROUND: In cancer cells in vitro, the glycolytic pathway and the mitochondrial tricarboxylic acid (TCA) cycle are programmed to produce more precursor molecules, and relatively less ATP, than in differentiated cells. We address the questions of whether and where these changes occur in vivo in glioblastomas grown from C6 cells in rat brain. These gliomas show some spatial organization, notably in the upregulation of membrane proton transporters near the rim. RESULTS: We immunolabeled pairs of proteins (as well as DNA) on sections of rat brains containing gliomas, measured the profiles of fluorescence intensity on strips 200 µm wide and at least 3 mm long running perpendicular to the tumor rim, and expressed the intensity in the glioma relative to that outside. On averaged profiles, labeling of a marker of the glycolytic pathway, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), was, as expected, greater in the glioma. Over distances up to 2.5 mm into the glioma, expression of a marker of the TCA cycle, Tom20, a pre-protein receptor on the translocation complex of the mitochondrial outer membrane, was also upregulated. The ratio of upregulation of Tom20 to upregulation of GAPDH was, on average, slightly greater than one. Near the rim (0.4-0.8 mm), GAPDH was expressed less and there was a peak in the mean ratio of 1.16, SEM = 0.001, N = 16 pairs of profiles. An antibody to V-ATPase, which, by pumping protons into vacuoles contributes to cell growth, also indicated upregulation by about 40%. When compared directly with GAPDH, upregulation of V-ATPase was only 0.764, SD = 0.016 of GAPDH upregulation. CONCLUSIONS: Although there was considerable variation between individual measured profiles, on average, markers of the glycolytic pathway, of mitochondria, and of cell proliferation showed coherent upregulation in C6 gliomas. There is a zone, close to the rim, where mitochondrial presence is upregulated more than the glycolytic pathway, in agreement with earlier suggestions that lactate is taken up by cells near the rim.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Glioma/metabolismo , Glioma/patología , Glucólisis , Mitocondrias/metabolismo , Bombas de Protones/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular , Gliceraldehído-3-Fosfato Deshidrogenasas/metabolismo , Masculino , Proteínas de Transporte de Membrana , Proteínas del Complejo de Importación de Proteínas Precursoras Mitocondriales , Proteínas Mitocondriales/metabolismo , Ratas Wistar , Receptores de Superficie Celular , Receptores Citoplasmáticos y Nucleares/metabolismo , Regulación hacia Arriba , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
Imaging heterogeneous cancer lesions is a real challenge. For diagnosis, histology often remains the reference, but it is widely acknowledged that biopsies are not reliable. There is thus a strong interest in establishing a link between clinical in vivo imaging and the biologic properties of tissues. In this study, we propose to construct histology-resembling images based on tissue microvascularization, a magnetic resonance imaging (MRI) accessible source of contrast. To integrate the large amount of information collected with microvascular MRI, we combined a manual delineation of a spatial region of interest with an unsupervised, model-based cluster analysis (Mclust). This approach was applied to two rat models of glioma (C6 and F98). Six MRI parameters were mapped: apparent diffusion coefficient, vessel wall permeability, cerebral blood volume fraction, cerebral blood flow, tissular oxygen saturation, and cerebral metabolic rate of oxygen. Five clusters, defined by their MRI features, were found to correspond to specific histologic features, and revealed intratumoral spatial structures. These results suggest that the presence of a cluster within a tumor can be used to assess the presence of a tissue type. In addition, the cluster composition, i.e., a signature of the intratumoral structure, could be used to characterize tumor models as histology does.
Asunto(s)
Neoplasias Encefálicas/patología , Encéfalo/patología , Glioma/patología , Imagen por Resonancia Magnética/métodos , Microvasos/patología , Animales , Encéfalo/irrigación sanguínea , Neoplasias Encefálicas/irrigación sanguínea , Línea Celular Tumoral , Análisis por Conglomerados , Análisis Discriminante , Glioma/irrigación sanguínea , Procesamiento de Imagen Asistido por Computador , Masculino , Trasplante de Neoplasias , Ratas , Ratas Endogámicas F344 , Ratas WistarRESUMEN
The aim of this study was to compare magnetic resonance imaging (MRI) and histological estimates of the mean vessel diameter (mVD), the vessel density (Density), and the vessel size index (VSI) obtained in the same tumor-bearing animals. Twenty-seven rats bearing intracranial glioma (C6 or RG2) were imaged by MRI. Changes in transverse relaxations (ΔR 2* and R(2)) were induced by the injection of an iron-based contrast agent and were mapped using a multi gradient-echo spin-echo sequence. Then, brain vascular network was studied ex vivo by histology. Three regions of interest were drawn in apparently normal tissue (neocortex and striatum) and in the tumor. In vivo mVD(MRI), Density(MRI), and VSI(MRI) were measured; ex vivo, mVD(histo), Density(histo), and VSI(histo) were quantified on the same animals. MRI and histology measurements differed by -15 to 26%. A positive correlation was found between MRI and histology for mVD, Density, and VSI counterparts (R(2) = 0.62, 0.50, 0.73, respectively; P < 0.001 in all cases). This study indicates that MRI and histology yields well correlated the estimates of mVD, Density, and VSI. VSI is the closest MRI estimate to histology. As Density and mVD or VSI provide complementary information, it is worth computing them to characterize angiogenesis beyond blood volume fraction.
Asunto(s)
Neoplasias Encefálicas/irrigación sanguínea , Encéfalo/irrigación sanguínea , Glioma/irrigación sanguínea , Imagen por Resonancia Magnética , Animales , Medios de Contraste , Cuerpo Estriado/irrigación sanguínea , Técnicas Histológicas , Hierro/sangre , Masculino , Neocórtex/irrigación sanguínea , Trasplante de Neoplasias , Ratas , Ratas Endogámicas F344 , Ratas WistarRESUMEN
BACKGROUND: Glioma is the most aggressive tumor of the brain and the most efficient treatments are based on radiotherapy. However, tumors are often resistant to radiotherapy due to an enhanced DNA repair activity. Short and stabilized DNA molecules (Dbait) have recently been proposed as an efficient strategy to inhibit DNA repair in tumor. METHODOLOGY/PRINCIPAL FINDINGS: The distribution of three formulations of Dbait, (i) Dbait alone, (ii) Dbait associated with polyethylenimine, and (iii) Dbait linked with cholesterol (coDbait), was evaluated one day after intratumoral delivery in an RG2 rat glioma model. Dbait molecule distribution was assessed in the whole organ with 2D-FRI and in brain sections. CoDbait was chosen for further studies given its good retention in the brain, cellular localization, and efficacy in inducing the activation of DNA repair effectors. The radiosensitizing effect of coDbait was studied in four groups of rats bearing RG2-glioma: no treatment, radiotherapy only, coDbait alone, and CoDbait with radiotherapy. Treatment started 7 days after tumor inoculation and consisted of two series of treatment in two weeks: coDbait injection followed by a selective 6-Gy irradiation of the head. We evaluated the radiosensitizing effect using animal survival, tumor volume, cell proliferation, and vasculature characteristics with multiparametric MRI. CoDbait with radiotherapy improved the survival of rats bearing RG2-glioma by reducing tumor growth and cell proliferation without altering tumor vasculature. CONCLUSION/SIGNIFICANCE: coDbait is therefore a promising molecular therapy to sensitize glioma to radiotherapy.
Asunto(s)
Colesterol/metabolismo , ADN/metabolismo , ADN/farmacología , Glioblastoma/patología , Fármacos Sensibilizantes a Radiaciones/metabolismo , Fármacos Sensibilizantes a Radiaciones/farmacología , Animales , Transporte Biológico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Química Farmacéutica , ADN/efectos adversos , ADN/química , Roturas del ADN de Doble Cadena , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Glioblastoma/irrigación sanguínea , Glioblastoma/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/efectos de la radiación , Imagen por Resonancia Magnética , Masculino , Vaina de Mielina/efectos de los fármacos , Vaina de Mielina/metabolismo , Vaina de Mielina/efectos de la radiación , Neostriado/efectos de los fármacos , Neostriado/metabolismo , Neostriado/patología , Neostriado/efectos de la radiación , Neovascularización Patológica , Polietileneimina/química , Fármacos Sensibilizantes a Radiaciones/efectos adversos , Fármacos Sensibilizantes a Radiaciones/química , Ratas , Análisis de Supervivencia , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/efectos de la radiaciónRESUMEN
PURPOSE: The purpose of this study was to evaluate high-dose single fraction delivered with monochromatic X-rays minibeams for the radiotherapy of primary brain tumors in rats. METHODS AND MATERIALS: Two groups of healthy rats were irradiated with one anteroposterior minibeam incidence (four minibeams, 123 Gy prescribed dose at 1 cm depth in the brain) or two interleaved incidences (54 Gy prescribed dose in a 5 × 5 × 4.8 mm(3) volume centered in the right hemisphere), respectively. Magnetic resonance imaging (MRI) follow-up was performed over 1 year. T2-weighted (T2w) images, apparent diffusion coefficient (ADC), and blood vessel permeability maps were acquired. F98 tumor bearing rats were also irradiated with interleaved minibeams to achieve a homogeneous dose of 54 Gy delivered to an 8 × 8 × 7.8 mm(3) volume centered on the tumor. Anatomic and functional MRI follow-up was performed every 10 days after irradiation. T2w images, ADC, and perfusion maps were acquired. RESULTS: All healthy rats were euthanized 1 year after irradiation without any clinical alteration visible by simple examination. T2w and ADC measurements remain stable for the single incidence irradiation group. Localized Gd-DOTA permeability, however, was observed 9 months after irradiation for the interleaved incidences group. The survival time of irradiated glioma bearing rats was significantly longer than that of untreated animals (49 ± 12.5 days versus 23.3 ± 2 days, p < 0.001). The tumoral cerebral blood flow and blood volume tend to decrease after irradiation. CONCLUSIONS: This study demonstrates the sparing effect of minibeams on healthy tissue. The increased life span achieved for irradiated glioma bearing rats was similar to the one obtained with other radiotherapy techniques. This experimental tumor therapy study shows the feasibility of using X-ray minibeams with high doses in brain tumor radiotherapy.
Asunto(s)
Neoplasias Encefálicas/radioterapia , Irradiación Craneana/métodos , Glioma/radioterapia , Tratamientos Conservadores del Órgano/métodos , Animales , Volumen Sanguíneo/efectos de la radiación , Neoplasias Encefálicas/irrigación sanguínea , Neoplasias Encefálicas/mortalidad , Neoplasias Encefálicas/patología , Irradiación Craneana/instrumentación , Estudios de Factibilidad , Glioma/irrigación sanguínea , Glioma/mortalidad , Glioma/patología , Imagen por Resonancia Magnética , Masculino , Modelos Animales , Tratamientos Conservadores del Órgano/instrumentación , Órganos en Riesgo , Radioterapia/métodos , Dosificación Radioterapéutica , Ratas , Ratas Endogámicas F344 , Análisis de Supervivencia , Sincrotrones/instrumentaciónRESUMEN
Diffusion tensor imaging (DTI) was used to study traumatic brain injury. The impact-acceleration trauma model was used in rats. Here, in addition to diffusivities (mean, axial and radial), fractional anisotropy (FA) was used, in particular, as a parameter to characterize the cerebral tissue early after trauma. DTI was implemented at 7 T using fast spiral k-space sampling and the twice-refocused spin echo radiofrequency sequence for eddy current minimization. The method was carefully validated on different phantom measurements. DTI of a trauma group (n = 5), as well as a sham group (n = 5), was performed at different time points during 6 h following traumatic brain injury. Two cerebral regions, the cortex and corpus callosum, were analyzed carefully. A significant decrease in diffusivity in the trauma group versus the sham group was observed, suggesting the predominance of cellular edema in both cerebral regions. No significant FA change was detected in the cortex. In the corpus callosum of the trauma group, the FA indices were significantly lower. A net discontinuity in fiber reconstructions in the corpus callosum was observed by fiber tracking using DTI. Histological analysis using Hoechst, myelin basic protein and Bielschowsky staining showed fiber disorganization in the corpus callosum in the brains of the trauma group. On the basis of our histology results and the characteristics of the impact-acceleration model responsible for the presence of diffuse axonal injury, the detection of low FA caused by a drastic reduction in axial diffusivity and the presence of fiber disconnections of the DTI track in the corpus callosum were considered to be related to the presence of diffuse axonal injury.
Asunto(s)
Lesión Axonal Difusa/diagnóstico , Lesión Axonal Difusa/patología , Imagen de Difusión Tensora/métodos , Animales , Butadienos/química , Calibración , Cuerpo Calloso/patología , Difusión , Modelos Animales de Enfermedad , Elastómeros/química , Masculino , Ratas , Ratas Wistar , Marcadores de SpinRESUMEN
Vessel size index (VSI), a parameter related to the distribution of vessel diameters, may be estimated using two MRI approaches: (i) dynamic susceptibility contrast (DSC) MRI following the injection of a bolus of Gd-chelate. This technique is routinely applied in the clinic to assess intracranial tissue perfusion in patients; (ii) steady-state susceptibility contrast with USPIO contrast agents, which is considered here as the standard method. Such agents are not available for human yet and the steady-state approach is currently limited to animal studies. The aim is to compare VSI estimates obtained with these two approaches on rats bearing C6 glioma (n = 7). In a first session, VSI was estimated from two consecutive injections of Gd-Chelate (Gd(1) and Gd(2)). In a second session (4 hours later), VSI was estimated using USPIO. Our findings indicate that both approaches yield comparable VSI estimates both in contralateral (VSI{USPIO} = 7.5 ± 2.0 µm, VSI{Gd(1)} = 6.5 ± 0.7 µm) and in brain tumour tissues (VSI{USPIO} = 19.4 ± 7.1 µm, VSI{Gd(1)} = 16.6 ± 4.5 µm). We also observed that, in the presence of BBB leakage (as it occurs typically in brain tumours), applying a preload of Gd-chelate improves the VSI estimate with the DSC approach both in contralateral (VSI{Gd(2)} = 7.1 ± 0.4 µm) and in brain tumour tissues (VSI{Gd(2)} = 18.5 ± 4.3 µm) but is not mandatory. VSI estimates do not appear to be sensitive to T(1) changes related to Gd extravasation. These results suggest that robust VSI estimates may be obtained in patients at 3 T or higher magnetic fields with the DSC approach.
Asunto(s)
Vasos Sanguíneos/patología , Neoplasias Encefálicas/irrigación sanguínea , Medios de Contraste , Dextranos , Gadolinio , Glioma/irrigación sanguínea , Imagen por Resonancia Magnética/métodos , Nanopartículas de Magnetita , Animales , Línea Celular Tumoral , Difusión , Modelos Animales de Enfermedad , Humanos , Masculino , Trasplante de Neoplasias , Tamaño de los Órganos , Ratas , Ratas WistarRESUMEN
PURPOSE: To analyze the contribution of the transverse relaxation parameter (T2), macroscopic field inhomogeneities (B0), and blood volume fraction (BVf) to blood oxygen level-dependent (BOLD)-based magnetic resonance (MR) measurements of blood oxygen saturation (SO2) obtained in a brain tumor model. MATERIALS AND METHODS: This study was approved by the local committee for animal care and use. Experiments were performed in accordance with permit 380 820 from the French Ministry of Agriculture. The 9L gliosarcoma cells were implanted in the brain of eight rats. Fifteen days later, 4.7-T MR examinations were performed to estimate T2*, T2, BVf, and T2*ΔB0corrected in the tumor and contralateral regions. MR estimates of SO2 were derived by combining T2, BVf, and T2*ΔB0corrected according to a recently described quantitative BOLD approach. Scatterplots and linear regression analysis were used to identify correlation between parameters. Paired Student t tests were used to compare the tumor region with the contralateral region. RESULTS: No significant correlations were found between T2* and any parameter in either tumor tissue or healthy tissue. T2* in the tumor and T2* in the uninvolved contralateral brain were the same (36 msec±4 [standard deviation] vs 36 msec±5, respectively), which might suggest similar oxygenation. Adding T2 information (98 msec±7 vs 68 msec±2, respectively) alone yields results that suggest apparent hypo-oxygenation of the tumor, while incorporating BVf (5.3%±0.6 vs 2.6%±0.3, respectively) alone yields results that suggest apparent hyperoxygenation. MR estimates of SO2 obtained with a complete quantitative BOLD analysis, although not correlated with T2* values, suggest normal oxygenation (68%±3 vs 65%±4, respectively). MR estimates of SO2 obtained in the contralateral tissue agree with previously reported values. CONCLUSION: Additional measurements, such as BVf, T2, and B0, are needed to obtain reliable information on oxygenation with BOLD MR imaging. The proposed quantitative BOLD approach, which includes these measurements, appears to be a promising tool with which to map tumor oxygenation.
Asunto(s)
Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/metabolismo , Glioblastoma/diagnóstico , Glioblastoma/metabolismo , Imagen por Resonancia Magnética/métodos , Oximetría/métodos , Oxígeno/análisis , Animales , Línea Celular Tumoral , Masculino , Ratas , Ratas Endogámicas F344 , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
To assess angiogenesis noninvasively in a C6 rat brain tumor model, the rapid-steady-state-T(1) (RSST(1)) magnetic resonance imaging (MRI) method was used for microvascular blood volume fraction (BVf) quantification with a novel contrast agent gadolinium per (3,6 anhydro) α-cyclodextrin (Gd-ACX). In brain tissue contralateral to the tumor, equal BVfs were obtained with Gd-ACX and the clinically approved gadoterate meglumine (Gd-DOTA). Contrary to Gd-DOTA, which leaks out of the tumor vasculature, Gd-ACX was shown to remain vascular in the tumor tissue allowing quantification of the tumor BVf. We sought to confirm the obtained tumor BVf using an independent method: instead of using a 'standard' two-dimensional histologic method, we study here how vascular morphometry combined with a stereological technique can be used for three-dimensional assessment of the vascular volume fraction (V(V)). The V(V) is calculated from the vascular diameter and length density. First, the technique is evaluated on simulated data and the healthy rat brain vasculature and is then applied to the same C6 tumor vasculature previously quantified by RSST(1)-MRI with Gd-ACX. The mean perfused V(V) and the BVf obtained by MRI in tumor regions are practically equal and the technique confirms the spatial heterogeneity revealed by MRI.
Asunto(s)
Determinación del Volumen Sanguíneo/métodos , Volumen Sanguíneo/fisiología , Neoplasias Encefálicas/patología , Glioma/patología , Imagen por Resonancia Magnética/métodos , Microvasos/patología , Neovascularización Patológica/patología , Animales , Mapeo Encefálico , Neoplasias Encefálicas/irrigación sanguínea , Neoplasias Encefálicas/fisiopatología , Línea Celular Tumoral , Medios de Contraste , Glioma/irrigación sanguínea , Glioma/fisiopatología , Compuestos Heterocíclicos , Procesamiento de Imagen Asistido por Computador , Microscopía Fluorescente , Microvasos/fisiopatología , Modelos Anatómicos , Trasplante de Neoplasias , Neovascularización Patológica/diagnóstico , Neovascularización Patológica/fisiopatología , Compuestos Organometálicos , Ratas , Ratas WistarRESUMEN
Early imaging or blood biomarkers of tumor response is needed to customize anti-tumor therapy on an individual basis. This study evaluates the sensitivity and relevance of five potential MRI biomarkers. Sixty nude rats were implanted with human glioma cells (U-87 MG) and randomized into three groups: one group received an anti-angiogenic treatment (Sorafenib), a second a cytotoxic drug [1,3-bis(2-chloroethyl)-1-nitrosourea, BCNU (Carmustine)] and a third no treatment. The tumor volume, apparent diffusion coefficient (ADC) of water, blood volume fraction (BVf), microvessel diameter (vessel size index, VSI) and vessel wall integrity (contrast enhancement, CE) were monitored before and during treatment. Sorafenib reduced tumor CE as early as 1 day after treatment onset. By 4 days after treatment onset, tumor BVf was reduced and tumor VSI was increased. By 14 days after treatment onset, ADC was increased and the tumor growth rate was reduced. With BCNU, ADC was increased and the tumor growth rate was reduced 14 days after treatment onset. Thus, the estimated MRI parameters were sensitive to treatment at different times after treatment onset and in a treatment-dependent manner. This study suggests that multiparametric MR monitoring could allow the assessment of new anti-tumor drugs and the optimization of combined therapies.
Asunto(s)
Inhibidores de la Angiogénesis/uso terapéutico , Glioma/tratamiento farmacológico , Glioma/patología , Imagen por Resonancia Magnética/métodos , Inhibidores de la Angiogénesis/farmacología , Animales , Bencenosulfonatos/farmacología , Bencenosulfonatos/uso terapéutico , Volumen Sanguíneo/efectos de los fármacos , Carmustina/farmacología , Carmustina/uso terapéutico , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Glioma/irrigación sanguínea , Humanos , Masculino , Microvasos/efectos de los fármacos , Microvasos/patología , Modelos Biológicos , Niacinamida/análogos & derivados , Compuestos de Fenilurea , Piridinas/farmacología , Piridinas/uso terapéutico , Ratas , Ratas Desnudas , Sorafenib , Coloración y Etiquetado , Análisis de SupervivenciaRESUMEN
Tumors create a heterogeneous acidic microenvironment which assists their growth and which must be taken into account in the design of drugs and their delivery. In addition, the acidic extracellular pH (pHe) is itself exploited in several experimental techniques for drug delivery. The way the acidity is created is not clear. We report here the spatial organization of key proton-handling proteins in C6 gliomas in rat brain. The mean profiles across the tumor rim of the Na+/H+ exchanger NHE1, and the lactate-H+ cotransporter MCT1, both showed peaks. NHE1, which is important for extension and migration of cells in vitro, showed a peak 1.55 times higher than in extratumoural tissue at 0.33 mm from the edge. MCT1 had a broader peak, further into the tumor (maximum 1.76 fold at 1.0 mm from the edge). In contrast, MCT4 and the carbonic anhydrase CAIX, which are associated with hypoxia, were not significantly upregulated in the rim. The spatial distribution of MCT4 was highly correlated with that of CAIX, suggesting that their expression is regulated by the same factors. Since protons extruded by NHE1 diffuse away through extracellular clefts, NHE1 requires a continuous source of intracellular protons. From the stoichiometries of metabolic pathways that produce or consume H+, and the greater availability of glucose compared to oxygen in most parts of a tumor, we support the classic view that most of the net proton efflux from C6 gliomas originates in glycolytic formation of lactate and H+ inside the tumor, but add that some lactate is taken up into cells in the rim on MCT1, and some lactate diffuses away, leaving its associated protons available to re-enter cells for extrusion on NHE1. Therapeutic inhibition of NHE1, MCT1 or CAIX is predicted to affect different parts of a tumor.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Ácido Láctico/metabolismo , Transportadores de Ácidos Monocarboxílicos/metabolismo , Protones , Intercambiadores de Sodio-Hidrógeno/metabolismo , Animales , Transporte Biológico/fisiología , Neoplasias Encefálicas/patología , Células Cultivadas , Glioma/patología , Transporte Iónico/fisiología , Modelos Biológicos , Ratas , Distribución Tisular , Células Tumorales CultivadasRESUMEN
Blood oxygen saturation (SO(2)) is a promising parameter for the assessment of brain tissue viability in numerous pathologies. Quantitative blood oxygenation level-dependent (qBOLD)-like approaches allow the estimation of SO(2) by modelling the contribution of deoxyhaemoglobin to the MR signal decay. These methods require a high signal-to-noise ratio to obtain accurate maps through fitting procedures. In this article, we present a version of the qBOLD method at long TE taking into account separate estimates of T(2), total blood volume fraction (BV(f)) and magnetic field inhomogeneities. Our approach was applied to the brains of 13 healthy rats under normoxia, hyperoxia and hypoxia. MR estimates of local SO(2) (MR_LSO(2)) were compared with measurements obtained from blood gas analysis. A very good correlation (R(2) = 0.89) was found between brain MR_LSO(2) and sagittal sinus SO(2).
Asunto(s)
Encéfalo/metabolismo , Oxígeno/sangre , Animales , Volumen Sanguíneo , Encéfalo/anatomía & histología , Vena Femoral/metabolismo , Masculino , Ratas , Ratas WistarRESUMEN
Gliomas are the most common primary brain tumor affecting human adults and remain a therapeutic challenge because cells of origin are still unknown. Here, we investigated the cellular origin of low-grade gliomas in a rat model based on transplacental exposure to N-ethyl-N-nitrosourea (ENU). Longitudinal magnetic resonance imaging coupled to immunohistological and immunocytochemical analyses were used to further characterize low-grade rat gliomas at different stages of evolution. We showed that early low-grade gliomas have characteristics of oligodendroglioma-like tumors and exclusively contain NG2-expressing slow dividing precursor cells, which express early markers of oligodendroglial lineage. These tumor-derived precursors failed to fully differentiate into oligodendrocytes and exhibited multipotential abilities in vitro. Moreover, a few glioma NG2+ cells are resistant to radiotherapy and may be responsible for tumor recurrence, frequently observed in humans. Overall, these findings suggest that transformed multipotent NG2 glial precursor cell may be a potential cell of origin in the genesis of rat ENU-induced oligodendroglioma-like tumors. This work may open up new perspectives for understanding biology of human gliomas.
Asunto(s)
Antígenos/análisis , Neoplasias Encefálicas/inducido químicamente , Etilnitrosourea/toxicidad , Células Madre Neoplásicas/patología , Oligodendroglioma/inducido químicamente , Proteoglicanos/análisis , Animales , Neoplasias Encefálicas/patología , Diferenciación Celular , Línea Celular Tumoral , Células Madre Neoplásicas/química , Oligodendroglioma/patología , Ratas , Ratas Sprague-Dawley , Proteínas Activadoras de ras GTPasa/análisisRESUMEN
We found that recombinant human erythropoietin (rhEPO) reduced significantly the development of brain edema in a rat model of diffuse traumatic brain injury (TBI) (impact-acceleration model). In this study, we investigated the molecular and intracellular changes potentially involved in these immediate effects. Brain tissue nitric oxide (NO) synthesis, phosphorylation level of two protein kinases (extracellular-regulated kinase (ERK)-1/-2 and Akt), and brain water content were measured 1 (H1) and 2 h (H2) after insult. Posttraumatic administration of rhEPO (5,000 IU/kg body weight, intravenously, 30 mins after injury) reduced TBI-induced upregulation of ERK phosphorylation, although it increased Akt phosphorylation at H1. These early molecular changes were associated with a reduction in brain NO synthesis at H1 and with an attenuation of brain edema at H2. Intraventricular administration of the ERK-1/-2 inhibitor, U0126, or the Akt inhibitor, LY294002, before injury showed that ERK was required for brain edema formation, and that rhEPO-induced reduction of edema could involve the ERK pathway. These results were obtained in the absence of any evidence of blood-brain barrier damage on contrast-enhanced magnetic resonance images. The findings of our study indicate that the anti edematous effect of rhEPO could be mediated through an early inhibition of ERK phosphorylation after diffuse TBI.
Asunto(s)
Edema Encefálico/prevención & control , Lesiones Encefálicas/tratamiento farmacológico , Eritropoyetina/farmacología , Proteínas Quinasas/efectos de los fármacos , Animales , Barrera Hematoencefálica/patología , Western Blotting , Edema Encefálico/etiología , Lesiones Encefálicas/complicaciones , Lesiones Encefálicas/enzimología , Modelos Animales de Enfermedad , Humanos , Masculino , Fosforilación , Proteínas Quinasas/metabolismo , Ratas , Ratas Wistar , Proteínas RecombinantesRESUMEN
This study aimed at combining an iron-based, steady-state, vessel size index magnetic resonance imaging (VSI MRI) approach, and a gadolinium (Gd)-based, dynamic contrast-enhanced MRI approach (DCE MRI) to characterize tumoral microvasculature. Rats bearing an orthotopic glioma (C6, n=14 and RG2, n=6) underwent DCE MRI and combined VSI and DCE MRI 4 h later, at 2.35 T. Gd-DOTA (200 mumol of Gd per kg) and ultrasmall superparamagnetic iron oxide (USPIO) (200 micromol of iron per kg) were used for DCE and VSI MRI, respectively. C6 and RG2 gliomas were equally permeable to Gd-DOTA but presented different blood volume fractions and VSI, in good agreement with histologic data. The presence of USPIO yielded reduced K(trans) values. The K(trans) values obtained with Gd-DOTA in the absence and in the presence of USPIO were well correlated for the C6 glioma but not for the RG2 glioma. It was also observed that, within the time frame of DCE MRI, USPIO remained intravascular in the C6 glioma whereas it extravasated in the RG2 glioma. In conclusion, VSI and DCE MRI can be combined provided that USPIO does not extravasate with the time frame of the DCE MRI experiment. The mechanisms at the origin of USPIO extravasation remain to be elucidated.
Asunto(s)
Gadolinio/farmacocinética , Hierro/farmacocinética , Imagen por Resonancia Magnética/métodos , Neovascularización Patológica/diagnóstico , Animales , Neoplasias Encefálicas/irrigación sanguínea , Neoplasias Encefálicas/diagnóstico , Permeabilidad de la Membrana Celular , Medios de Contraste , Dextranos/farmacocinética , Óxido Ferrosoférrico/farmacocinética , Glioma/diagnóstico , Glioma/patología , Compuestos Heterocíclicos/farmacocinética , Nanopartículas de Magnetita , Masculino , Microvasos , Compuestos Organometálicos/farmacocinética , Ratas , Ratas WistarRESUMEN
The purpose of this study is to measure the effects of a tomographic synchrotron irradiation on healthy mouse brain. The cerebral cortexes of healthy nude mice were irradiated with a monochromatic synchrotron beam of 79 keV at a dose of 15 Gy in accordance with a protocol of photoactivation of cisplatin previously tested in our laboratory. Forty-eight hours, one week and one month after irradiation, the blood brain barrier (BBB) permeability was measured in the irradiated area with intravital multiphoton microscopy using fluorescent dyes with molecular weights of 4 and 70 kDa. Vascular parameters and gliosis were also assessed using quantitative immunohistochemistry. No extravasation of the fluorescent dyes was observed in the irradiated area at any measurement time (48 h, 1 week, 1 month). It appears that the BBB remains impermeable to molecules with a molecular weight of 4 kDa and above. The vascular density and vascular surface were unaffected by irradiation and no gliosis was induced. These findings suggest that a 15 Gy/79 keV synchrotron irradiation does not induce important damage on brain vasculature and tissue on the short term following irradiation.
Asunto(s)
Barrera Hematoencefálica/efectos de la radiación , Encéfalo/irrigación sanguínea , Encéfalo/efectos de la radiación , Animales , Membrana Basal/química , Encéfalo/patología , Colágeno Tipo IV/análisis , Femenino , Proteína Ácida Fibrilar de la Glía/análisis , Gliosis/patología , Inmunohistoquímica , Ratones , Ratones Desnudos , Dosificación Radioterapéutica , SincrotronesRESUMEN
Assessment of angiogenesis may help to determine tumor grade and therapy follow-up. In vivo imaging methods for non-invasively monitoring microvasculature evolution are therefore of major interest for tumor management. MRI evaluation of blood volume fraction (BVf) and vessel size index (VSI) was applied to assess the evolution of tumor microvasculature in two rat models of glioma (C6 and RG2). The results show that repeated MRI of BVf and VSI - which involves repeated injection of an iron-based MR contrast agent - does not affect either the physiological status of the animals or the accuracy of the MR estimates of the microvascular parameters. The MR measurements were found to correlate well with those obtained from histology. They indicate that microvascular evolution differs significantly between the two glioma models, in good agreement with expression of angiogenic factors (vascular endothelial growth factor, angiopoietin-2) and with activities of matrix metalloproteinases, also assessed in this study. These MRI methods thus provide considerable potential for assessing the response of gliomas to anti-angiogenic and anti-vascular agents, in preclinical studies as well as in the clinic. Furthermore, as differences between the fate of tumor microvasculature may underlie differences in therapeutic response, there is a need for preclinical study of several tumor models.
Asunto(s)
Proteínas Angiogénicas/análisis , Neoplasias Encefálicas/fisiopatología , Glioma/fisiopatología , Imagen por Resonancia Magnética/métodos , Microvasos/patología , Microvasos/fisiopatología , Neovascularización Patológica/fisiopatología , Inductores de la Angiogénesis/análisis , Animales , Biomarcadores de Tumor/análisis , Volumen Sanguíneo , Neoplasias Encefálicas/diagnóstico , Línea Celular Tumoral , Glioma/diagnóstico , Masculino , Proteínas de Neoplasias/análisis , Neovascularización Patológica/diagnóstico , Tamaño de los Órganos , Ratas , Ratas Endogámicas F344 , Ratas Wistar , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
The aim of this work focuses on the description of the short-term response of a 9L brain tumor model and its vasculature to microbeam radiation therapy (MRT) using magnetic resonance imaging (MRI). Rat 9L gliosarcomas implanted in nude mice brains were irradiated by MRT 13 days after tumor inoculation using two orthogonal arrays of equally spaced 28 planar microbeams (25 microm width, 211 microm spacing and dose 500 Gy). At 1, 7 and 14 days after MRT, apparent diffusion coefficient, blood volume and vessel size index were mapped by MRI. Mean survival time after tumor inoculation increased significantly between MRT-treated and untreated groups (23 and 28 days respectively, log-rank test, p < 0.0001). A significant increase of apparent diffusion coefficient was observed 24 h after MRT in irradiated tumors versus non-irradiated ones. In the untreated group, both tumor size and vessel size index increased significantly (from 7.6 +/- 2.2 to 19.2 +/- 4.0 mm(2) and +23%, respectively) between the 14th and the 21st day after tumor cell inoculation. During the same period, in the MRT-treated group, no difference in tumor size was observed. The vessel size index measured in the MRT-treated group increased significantly (+26%) between 14 and 28 days of tumor growth. We did not observe the significant difference in blood volume between the MRT-treated and untreated groups. MRT slows 9L tumor growth in a mouse brain but MRI results suggest that the increase in survival time after our MRT approach may be rather due to a cytoreduction than to early direct effects of ionizing radiation on tumor vessels. These results suggest that MRT parameters need to be optimized to further damage tumor vessels.
Asunto(s)
Neoplasias Encefálicas/radioterapia , Gliosarcoma/radioterapia , Neovascularización Patológica/radioterapia , Radioterapia/métodos , Sincrotrones , Animales , Neoplasias Encefálicas/patología , Gliosarcoma/patología , Imagen por Resonancia Magnética/métodos , Masculino , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Neovascularización Patológica/patología , Radioterapia/instrumentación , Dosificación Radioterapéutica , Tasa de Supervivencia , Factores de Tiempo , Resultado del TratamientoRESUMEN
Cerebral edema is one of the main acute complications arising after irradiation of brain tumors. Microbeam radiation therapy (MRT), an innovative experimental radiotherapy technique using spatially fractionated synchrotron x-rays, has been shown to spare radiosensitive tissues such as mammal brains. The aim of this study was to determine if cerebral edema occurs after MRT using diffusion-weighted MRI and microgravimetry. Prone Swiss nude mice's heads were positioned horizontally in the synchrotron x-ray beam and the upper part of the left hemisphere was irradiated in the antero-posterior direction by an array of 18 planar microbeams (25 mm wide, on-center spacing 211 mm, height 4 mm, entrance dose 312 Gy or 1000 Gy). An apparent diffusion coefficient (ADC) was measured at 7 T 1, 7, 14, 21 and 28 days after irradiation. Eventually, the cerebral water content (CWC) was determined by microgravimetry. The ADC and CWC in the irradiated (312 Gy or 1000 Gy) and in the contralateral non-irradiated hemispheres were not significantly different at all measurement times, with two exceptions: (1) a 9% ADC decrease (p < 0.05) was observed in the irradiated cortex 1 day after exposure to 312 Gy, (2) a 0.7% increase (p < 0.05) in the CWC was measured in the irradiated hemispheres 1 day after exposure to 1000 Gy. The results demonstrate the presence of a minor and transient cellular edema (ADC decrease) at 1 day after a 312 Gy exposure, without a significant CWC increase. One day after a 1000 Gy exposure, the CWC increased, while the ADC remained unchanged and may reflect the simultaneous presence of cellular and vasogenic edema. Both types of edema disappear within a week after microbeam exposure which may confirm the normal tissue sparing effect of MRT.