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PURPOSE: Sotrovimab (VIR-7831), a human IgG1κ monoclonal antibody (mAb), binds to a conserved epitope on the SARS-CoV-2 spike protein receptor binding domain (RBD). The Fc region of VIR-7831 contains an LS modification to promote neonatal Fc receptor (FcRn)-mediated recycling and extend its serum half-life. Here, we aimed to evaluate the impact of the LS modification on tissue biodistribution, by comparing VIR-7831 to its non-LS-modified equivalent, VIR-7831-WT, in cynomolgus monkeys. METHODS: 89Zr-based PET/CT imaging of VIR-7831 and VIR-7831-WT was performed up to 14 days post injection. All major organs were analyzed for absolute concentration as well as tissue:blood ratios, with the focus on the respiratory tract, and a physiologically based pharmacokinetics (PBPK) model was used to evaluate the tissue biodistribution kinetics. Radiomics features were also extracted from the PET images and SUV values. RESULTS: SUVmean uptake in the pulmonary bronchi for 89Zr-VIR-7831 was statistically higher than for 89Zr-VIR-7831-WT at days 6 (3.43 ± 0.55 and 2.59 ± 0.38, respectively) and 10 (2.66 ± 0.32 and 2.15 ± 0.18, respectively), while the reverse was observed in the liver at days 6 (5.14 ± 0.80 and 8.63 ± 0.89, respectively), 10 (4.52 ± 0.59 and 7.73 ± 0.66, respectively), and 14 (4.95 ± 0.65 and 7.94 ± 0.54, respectively). Though the calculated terminal half-life was 21.3 ± 3.0 days for VIR-7831 and 16.5 ± 1.1 days for VIR-7831-WT, no consistent differences were observed in the tissue:blood ratios between the antibodies except in the liver. While the lung:blood SUVmean uptake ratio for both mAbs was 0.25 on day 3, the PBPK model predicted the total lung tissue and the interstitial space to serum ratio to be 0.31 and 0.55, respectively. Radiomics analysis showed VIR-7831 had mean-centralized PET SUV distribution in the lung and liver, indicating more uniform uptake than VIR-7831-WT. CONCLUSION: The half-life extended VIR-7831 remained in circulation longer than VIR-7831-WT, consistent with enhanced FcRn binding, while the tissue:blood concentration ratios in most tissues for both drugs remained statistically indistinguishable throughout the course of the experiment. In the bronchiolar region, a higher concentration of 89Zr-VIR-7831 was detected. The data also allow unparalleled insight into tissue distribution and elimination kinetics of mAbs that can guide future biologic drug discovery efforts, while the residualizing nature of the 89Zr label sheds light on the sites of antibody catabolism.
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COVID-19 , SARS-CoV-2 , Animales , Recién Nacido , Humanos , Distribución Tisular , Macaca fascicularis/metabolismo , SARS-CoV-2/metabolismo , Tomografía Computarizada por Tomografía de Emisión de Positrones , Anticuerpos Monoclonales/metabolismo , CirconioRESUMEN
Liposomes are promising targeted drug delivery systems with the potential to improve the efficacy and safety profile of certain classes of drugs. Though attractive, there are unique analytical challenges associated with the development of liposomal drugs including human dose prediction given these are multi-component drug delivery systems. In this study, we developed a multimodal imaging approach to provide a comprehensive distribution assessment for an antibacterial drug, GSK2485680, delivered as a liposomal formulation (Lipo680) in a mouse thigh model of bacterial infection to support human dose prediction. Positron emission tomography (PET) imaging was used to track the in vivo biodistribution of Lipo680 over 48 h post-injection providing a clear assessment of the uptake in various tissues and, importantly, the selective accumulation at the site of infection. In addition, a pharmacokinetic model was created to evaluate the kinetics of Lipo680 in different tissues. Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) was then used to quantify the distribution of GSK2485680 and to qualitatively assess the distribution of a liposomal lipid throughout sections of infected and non-infected hindlimb tissues at high spatial resolution. Through the combination of both PET and MALDI IMS, we observed excellent correlation between the Lipo680-radionuclide signal detected by PET with the GSK2485680 and lipid component signals detected by MALDI IMS. This multimodal translational method can reduce drug attrition by generating comprehensive biodistribution profiles of drug delivery systems to provide mechanistic insight and elucidate safety concerns. Liposomal formulations have potential to deliver therapeutics across a broad array of different indications, and this work serves as a template to aid in delivering future liposomal drugs to the clinic.
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Enfermedades Transmisibles , Liposomas , Animales , Ratones , Humanos , Liposomas/química , Distribución Tisular , Antibacterianos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Tomografía de Emisión de Positrones , Imagen Multimodal , LípidosRESUMEN
Autosomal dominant polycystic kidney disease (ADPKD) is a leading monogenetic cause of end-stage renal disease with limited therapeutic repertoire. A targeted drug delivery strategy that directs a small molecule to renal niches around cysts could increase the safety margins of agents that slow the progression of ADPKD but are poorly tolerated due to extrarenal toxicity. Herein, we determined whether previously characterized lysine-based and glutamic acid-based megalin-binding peptides can achieve renal-specific localization in the juvenile cystic kidney (JCK) mouse model of polycystic kidney disease and whether the distribution is altered compared with control mice. We performed in vivo optical and magnetic resonance imaging studies using peptides conjugated to the VivoTag 680 dye and demonstrated that megalin-interacting peptides distributed almost exclusively to the kidney cortex in both normal and JCK mice. Confocal analysis demonstrated that the peptide-dye conjugate distribution overlapped with megalin-positive renal proximal tubules. However, in the JCK mouse, the epithelium of renal cysts did not retain expression of the proximal tubule markers aquaporin 1 and megalin, and therefore these cysts did not retain peptide-dye conjugates. Furthermore, human kidney tumor tissues were evaluated by immunohistochemistry and revealed significant megalin expression in tissues from patients with renal cell carcinoma, raising the possibility that these tumors could be treated using this drug delivery strategy. Taken together, our data suggest that linking a small-molecule drug to these carrier peptides could represent a promising opportunity to develop a new platform for renal enrichment and targeting in the treatment of ADPKD and certain renal carcinomas.
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Sistemas de Liberación de Medicamentos/métodos , Riñón/efectos de los fármacos , Péptidos/administración & dosificación , Enfermedades Renales Poliquísticas/tratamiento farmacológico , Animales , Acuaporina 1/metabolismo , Colorantes , Diseño de Fármacos , Epitelio/metabolismo , Ácido Glutámico/química , Humanos , Corteza Renal/diagnóstico por imagen , Corteza Renal/metabolismo , Neoplasias Renales/metabolismo , Túbulos Renales Proximales/metabolismo , Proteína 2 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Lisina/química , Imagen por Resonancia Magnética , Ratones , Péptidos/química , Péptidos/farmacocinética , Enfermedades Renales Poliquísticas/diagnóstico por imagen , Distribución TisularRESUMEN
Drug-induced liver injury (DILI) is a leading cause of acute liver failure and transplantation. DILI can be the result of impaired hepatobiliary transporters, with altered bile formation, flow, and subsequent cholestasis. We used gadoxetate dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), combined with pharmacokinetic modelling, to measure hepatobiliary transporter function in vivo in rats. The sensitivity and robustness of the method was tested by evaluating the effect of a clinical dose of the antibiotic rifampicin in four different preclinical imaging centers. The mean gadoxetate uptake rate constant for the vehicle groups at all centers was 39.3 +/- 3.4 s-1 (n = 23) and 11.7 +/- 1.3 s-1 (n = 20) for the rifampicin groups. The mean gadoxetate efflux rate constant for the vehicle groups was 1.53 +/- 0.08 s-1 (n = 23) and for the rifampicin treated groups was 0.94 +/- 0.08 s-1 (n = 20). Both the uptake and excretion transporters of gadoxetate were statistically significantly inhibited by the clinical dose of rifampicin at all centers and the size of this treatment group effect was consistent across the centers. Gadoxetate is a clinically approved MRI contrast agent, so this method is readily transferable to the clinic. CONCLUSION: Rate constants of gadoxetate uptake and excretion are sensitive and robust biomarkers to detect early changes in hepatobiliary transporter function in vivo in rats prior to established biomarkers of liver toxicity.
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Medios de Contraste , Gadolinio DTPA , Hígado , Imagen por Resonancia Magnética , Animales , Transporte Biológico Activo/efectos de los fármacos , Biomarcadores/metabolismo , Medios de Contraste/farmacocinética , Medios de Contraste/farmacología , Evaluación Preclínica de Medicamentos , Gadolinio DTPA/farmacocinética , Gadolinio DTPA/farmacología , Hígado/diagnóstico por imagen , Hígado/metabolismo , Masculino , Ratas , Ratas WistarRESUMEN
Long-Acting Parenterals (LAPs) have been used in the clinic to provide sustained therapeutic drug levels at a target site, and thereby reducing the frequency of dosing required. In an effort to understand the factors associated with long-acting cabotegravir (GSK1265744 LAP) pharmacokinetic variability, the current study was designed to investigate the temporal relationship between intramuscular (IM) or subcutaneous (SC) drug depot morphology and distribution kinetics with plasma pharmacokinetics. Therefore, a multi-modal molecular imaging (MRI & MALDI IMS) approach was employed to examine the temporal GSK1265744 LAP biodistribution in rat following either IM or SC administration. Serial MRI was performed immediately post drug administration, and then at day 1 (24h post), 2, 3, 4, 7, and 14. In a separate cohort of rats, an MRI contrast agent, Feraheme® (USPIO), was administered 2days post IM drug injection in order to investigate the potential involvement of macrophages trafficking to the GSK1265744 LAP and Vehicle depot sites. The GSK1265744 LAP depot volume increased rapidly by day 2 in the IM injected rats (~3-7 fold) compared with a ~1 fold increase in the SC injected rats. In addition, the USPIO contrast agent labeled macrophages were shown to be present in the depot region of the GSK1265744 LAP injected gastrocnemius while the Vehicle injected gastrocnemius appeared to show reduced uptake. Matrix-assisted laser desorption ionization (MALDI) imaging mass spectrometry (IMS) of muscle and abdominal tissue sections identified the drug content primarily within the depot. Co-registration of the GSK1265744 ion images with immunohistochemical images established that the drug was taken up by macrophages associated with the depot. Linear regression analysis demonstrated that the drug depot characteristics including volume, surface area, and perimeter assessed by MRI at day 2 correlated with early time point plasma drug concentrations. In summary, a multimodal molecular imaging approach was used to identify the drug depot location and volumetric/physiologic changes in both IM and SC locations following GSK1265744 LAP administration. The IM depot volume increased rapidly to a maximum volume at 2days post-GSK1265744 LAP administration, while the Vehicle depot did not suggesting that the active drug substance and/or related particle was a key driver for drug depot evolution. The depot expansion was associated with an increase in macrophage infiltration and edema in and around the depot region and was correlated to plasma drug concentration at early time points (0-4days). Consequently, molecular imaging approaches may be used in patients to help understand the biodistribution of GSK1265744 LAP and its associated pharmacokinetics.
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Piridonas/administración & dosificación , Piridonas/farmacocinética , Animales , Medios de Contraste/administración & dosificación , Preparaciones de Acción Retardada/administración & dosificación , Preparaciones de Acción Retardada/farmacocinética , Óxido Ferrosoférrico/administración & dosificación , Inyecciones Intramusculares , Inyecciones Subcutáneas , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Imagen por Resonancia Magnética , Masculino , Imagen Multimodal , Piridonas/sangre , Ratas Sprague-Dawley , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Distribución TisularRESUMEN
To determine if amiodarone induces hepatic phospholipidosis (PLD) sufficient to detect changes in hepatobiliary transporter function as assessed by gadoxetate dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI), rats were orally dosed with vehicle (1% methyl cellulose) or amiodarone (300 mg/kg/day) for 7 consecutive days. Gadoxetate DCE-MRI occurred at baseline, day 7, and following a 2-week washout of amiodarone. At day 7, the gadoxetate washout rate was significantly decreased compared to the vehicle group. Blood chemistry analysis revealed no significant changes in liver enzymes (alanine aminotransferase [ALT]/aspartate aminotransferase [AST]/alkaline phosphatase [ALP]), bilirubin, or bile acids between vehicle or amiodarone groups. Hepatic PLD was confirmed in all rats treated with amiodarone at day 7 by transmission electron microscopy. Following the 2-week washout, there was no ultrastructural evidence of hepatic PLD in rats and the gadoxetate washout rate returned to baseline levels. This is the first study to show the application of gadoxetate DCE-MRI to detect hepatobiliary functional changes associated with PLD and offer a potential new technique with clinical utility in patients suspected of having PLD. These results also suggest PLD itself has functional consequences on hepatobiliary function in the absence of biomarkers of toxicity, given the cause/effect relationship between PLD and function has not been fully established.
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Sistema Biliar/fisiopatología , Gadolinio DTPA/farmacocinética , Lipidosis/fisiopatología , Hígado/fisiopatología , Imagen por Resonancia Magnética/métodos , Fosfolípidos/metabolismo , Amiodarona/toxicidad , Animales , Sistema Biliar/metabolismo , Sistema Biliar/patología , Lipidosis/inducido químicamente , Lipidosis/metabolismo , Hígado/metabolismo , Hígado/patología , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND: Despite systemic exposure to risk factors, the circulatory system develops varying patterns of atherosclerosis for unclear reasons. In a porcine model, we investigated the relationship between site-specific lesion development and inflammatory pathways involved in the coronary arteries (CORs) and distal abdominal aortas (AAs). METHODS AND RESULTS: Diabetes mellitus (DM) and hypercholesterolemia (HC) were induced in 37 pigs with 3 healthy controls. Site-specific plaque development was studied by comparing plaque severity, macrophage infiltration, and inflammatory gene expression between CORs and AAs of 17 DM/HC pigs. To assess the role of lipoprotein-associated phospholipase A2 (Lp-PLA2) in plaque development, 20 DM/HC pigs were treated with the Lp-PLA2 inhibitor darapladib and compared with the 17 DM/HC untreated pigs. DM/HC caused site-specific differences in plaque severity. In the AAs, normalized plaque area was 4.4-fold higher (P<0.001) and there were more fibroatheromas (9 of the 17 animals had a fibroatheroma in the AA and not the COR, P=0.004), while normalized macrophage staining area was 1.5-fold higher (P=0.011) compared with CORs. DM/HC caused differential expression of 8 of 87 atherosclerotic genes studied, including 3 important in inflammation with higher expression in the CORs. Darapladib-induced attenuation of normalized plaque area was site-specific, as CORs responded 2.9-fold more than AAs (P=0.045). CONCLUSIONS: While plaque severity was worse in the AAs, inflammatory genes and inflammatory pathways that use Lp-PLA2 were more important in the CORs. Our results suggest fundamental differences in inflammation between vascular sites, an important finding for the development of novel anti-inflammatory therapeutics.
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1-Alquil-2-acetilglicerofosfocolina Esterasa/antagonistas & inhibidores , Aorta Abdominal/patología , Aterosclerosis/metabolismo , Vasos Coronarios/patología , Inflamación/metabolismo , Placa Aterosclerótica/patología , Animales , Aorta Abdominal/inmunología , Benzaldehídos/farmacología , Vasos Coronarios/inmunología , Diabetes Mellitus/inmunología , Diabetes Mellitus/patología , Modelos Animales de Enfermedad , Expresión Génica/efectos de los fármacos , Hipercolesterolemia/inmunología , Hipercolesterolemia/patología , Macrófagos/inmunología , Masculino , Oximas/farmacología , Inhibidores de Fosfolipasa A2/farmacología , Placa Aterosclerótica/metabolismo , PorcinosRESUMEN
Thymosin beta 4 (Tß4) was previously shown to reduce infarct size and improve contractile performance in chronic myocardial ischemic injury via two phases of action: an acute phase, just after injury, when Tß4 preserves ischemic myocardium via antiapoptotic or anti-inflammatory mechanisms; and a chronic phase, when Tß4 activates the growth of vascular or cardiac progenitor cells. In order to differentiate between the effects of Tß4 during the acute and during the chronic phases, and also in order to obtain detailed hemodynamic and biomarker data on the effects of Tß4 treatment suitable for use in clinical studies, we tested Tß4 in a rat model of chronic myocardial ischemia using two dosing regimens: short term dosing (Tß4 administered only during the first 3 days following injury), and long term dosing (Tß4 administered during the first 3 days following injury and also every third day until the end of the study). Tß4 administered throughout the study reduced infarct size and resulted in significant improvements in hemodynamic performance; however, chamber volumes and ejection fractions were not significantly improved. Tß4 administered only during the first 3 days following injury tended to reduce infarct size, chamber volumes and improve hemodynamic performance. Plasma biomarkers of myocyte injury were significantly reduced by Tß4 treatment during the acute injury period, and plasma ANP levels were significantly reduced in both dosing groups. Surprisingly, neither acute nor chronic Tß4 treatment significantly increased blood vessel density in peri-infarct regions. These results suggest the following: repeated dosing may be required to achieve clinically measureable improvements in cardiac function post-myocardial infarction (MI); improvement in cardiac function may be observed in the absence of a high degree of angiogenesis; and that plasma biomarkers of cardiac function and myocardial injury are sensitive pharmacodynamic biomarkers of the effects of Tß4.
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The loss of cardiac reserve is, in part, responsible for exercise intolerance in late-stage heart failure (HF). Exercise tolerance testing (ETT) has been performed in mouse models of HF; however, treadmill performance and at-rest cardiac indexes determined by magnetic resonance imaging (MRI) rarely correlate. The present study adopted a stress-MRI technique for comparison with ETT in HF models, using isoproterenol (ISO) to evoke cardiac reserve responses. Male C57BL/6J mice were randomly subjected to myocardial infarction (MI), transverse aortic constriction (TAC), or sham surgery under general anesthesia. Mice underwent serial ETT on a graded treadmill with follow-up ISO stress-MRI. TAC mice showed consistent exercise intolerance, with a 16.2% reduction in peak oxygen consumption vs. sham at 15-wk postsurgery (WPS). MI and sham mice had similar peak oxygen consumption from 7 WPS onward. Time to a respiratory exchange ratio of 1.0 correlated with ETT distance (r = 0.64; P < 0.001). The change in ejection fraction under ISO stress was reduced in HF mice at 4 WPS [10.1 ± 3.9% change (Δ) and 8.9 ± 3.5%Δ in MI and TAC, respectively, compared with 32.0 ± 3.5%Δ in sham; P < 0.001]. However, cardiac reserve differences between surgery groups were not observed at 16 WPS in terms of ejection fraction or cardiac output. In addition, ETT did not correlate with cardiac indexes under ISO stress. In conclusion, ISO stress was unable to reflect consistent differences in ETT between HF and healthy mice, suggesting cardiac-specific indexes are not the sole factors in defining exercise intolerance in mouse HF models.
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Insuficiencia Cardíaca/fisiopatología , Esfuerzo Físico/fisiología , Agonistas Adrenérgicos beta , Animales , Cardiotónicos , Modelos Animales de Enfermedad , Prueba de Esfuerzo , Tolerancia al Ejercicio/fisiología , Insuficiencia Cardíaca/etiología , Humanos , Isoproterenol , Imagen por Resonancia Magnética , Masculino , Ratones , Ratones Endogámicos C57BL , Infarto del Miocardio/complicaciones , Consumo de Oxígeno , Función Ventricular IzquierdaRESUMEN
The temporal evolution of heart failure and associated pulmonary congestion in rodent heart failure models has not yet been characterized simultaneously and noninvasively. In this study, MRI was used to assess the serial progression of left-ventricular dysfunction and lung congestion in mice following myocardial infarction (MI). Cardiac and lung (1) H MRI was performed at baseline and every 3 days up to 13 days postsurgery in sham and MI mice. Respiratory parameters and terminal lung mechanics were assessed followed by histological analysis. MRI revealed that the MI induced significant pulmonary congestion/edema as detected by increased MRI signal intensity and was associated with increased lung volume and reduced cardiac contractility. Pulmonary function was also depressed in MI-mice, reflected by a reduced tidal volume and a low minute ventilation rate. Additionally, MI significantly increased lung resistance, markedly reduced lung compliance and total lung capacity and significantly increased lung weights by 57%. Significant correlations were observed between the MRI measured lung congestion, lung volume, ejection fraction, and lung wet-weight parameters. This study demonstrates that MRI may be of significant value in evaluating therapies aimed at primary intervention for lung congestion and secondary prevention of unfavorable cardiac remodeling.
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Imagen por Resonancia Magnética/métodos , Edema Pulmonar/patología , Edema Pulmonar/fisiopatología , Disfunción Ventricular Izquierda/patología , Disfunción Ventricular Izquierda/fisiopatología , Remodelación Ventricular/fisiología , Animales , Pulmón/patología , Pulmón/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Edema Pulmonar/complicaciones , Disfunción Ventricular Izquierda/complicacionesRESUMEN
Mismatches between tissue perfusion-weighted imaging (PWI; an index of blood flow deficit) and cellular diffusion-weighted imaging (DWI; an index of tissue injury) provide information on potentially salvageable penumbra tissue in focal stroke and can identify "treatable" stroke patients. The present pre-clinical studies were conducted to: a.) Determine PWI (using perfusion delay) and DWI measurements in two experimental stroke models, b.) Utilize these measurements to characterize selective ET(A) receptor antagonism (i.e., determine efficacy, time-to-treatment and susceptibility to treatment in the different stroke models), and c.) Determine if increasing the reduced blood flow following a stroke is a mechanism of protection. Permanent middle cerebral artery occlusion (MCAO) or sham surgeries were produced in Sprague Dawley rats (SD; proximal MCAO; hypothesized to be a model of slowly evolving brain injury with a significant penumbra) and in spontaneously hypertensive rats (SHR; distal MCAO; hypothesized to be a model of rapidly evolving brain injury with little penumbra). Infusions of vehicle or SB 234551 (3, 10, or 30 microg/kg/min) were initiated at 0, 75, and/or 180 min post-surgery and maintained for the remainder of 24 h post-surgery. Hyper-intense areas of perfusion delay (PWI) in the forebrain were measured using Gadolinium (Gd) bolus contrast. DWI hyper-intense areas were also measured, and the degree of forebrain DWI-PWI mismatch was determined. Region specific analyses (ROI) were also conducted in the core ischemic and low perfusion/penumbra areas to provide indices of perfusion and changes in the degree of tissue perfusion due to SB 234551 treatment. At 24 h post-surgery, final infarct volume was measured by DWI and by staining forebrain slices. Following SD proximal MCAO, there was a significant mismatch in the ischemic forebrain PWI compared to DWI (PWI>DWI) at 60 min which was maintained up to 150 min (all p<0.05). By 24 h post-stroke, infarct volume was identical to the area of early perfusion deficit/PWI, suggesting a slow progression of infarct development that expanded into the significant, earlier cortical penumbra (i.e., model with salvageable tissue with potential for intervention). When SB 234551 was administered within the period of peak mismatch (i.e., at 75 min post-stroke), SB 234551 provided significant dose-related reductions in cortical (penumbral) progression to infarction (p<0.05). Cortical protection was related to an increased/normalization of the stroke-induced decrease in tissue perfusion in cortical penumbra areas (p<0.05). No SB 234551-induced changes in reduced tissue perfusion were observed in the striatum core ischemic area. Also, when SB-234551 was administered beyond the time of mismatch, no effect on cortical penumbra progression to infarct was observed. In comparison and strikingly different, following SHR distal MCAO there was no mismatch between PWI and DWI (PWI=DWI) as early as 60 min post-stroke, with this early change in SHR DWI being identical to the final infarct volume at 24 h, suggesting a rapidly occurring brain injury with little cortical penumbra (i.e., model with little salvageable tissue or potential for intervention). In distal MCAO, SB 234551 administered immediately at the time of stroke did not have any effect on infarct volume in SHR. These data demonstrate that selective blockade of ET(A) receptors is protective following proximal MCAO in SD (i.e. a model similar to "treatable" clinical patients). The protective mechanism appears to be due to enhanced collateral blood flow and salvage of penumbra. Therefore, the use of PWI-DWI mismatch signatures can identify treatable stroke models characterized by a salvageable penumbra and can define appropriate time to treatment protocols. In addition, tissue perfusion information obtained under these conditions might clarify mechanism of protection in the evaluation of protective compounds for focal stroke.
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Infarto Encefálico/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Imagen de Difusión por Resonancia Magnética/métodos , Dioxoles/farmacología , Antagonistas de los Receptores de la Endotelina A , Pirazoles/farmacología , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Encéfalo/patología , Encéfalo/fisiopatología , Infarto Encefálico/patología , Infarto Encefálico/fisiopatología , Arterias Cerebrales/efectos de los fármacos , Arterias Cerebrales/metabolismo , Arterias Cerebrales/fisiopatología , Circulación Cerebrovascular/efectos de los fármacos , Circulación Cerebrovascular/fisiología , Dioxoles/uso terapéutico , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Servicios Médicos de Urgencia/normas , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/metabolismo , Infarto de la Arteria Cerebral Media/fisiopatología , Masculino , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Pirazoles/uso terapéutico , Ratas , Ratas Endogámicas SHR , Ratas Sprague-Dawley , Receptor de Endotelina A/metabolismo , Accidente Cerebrovascular/patología , Accidente Cerebrovascular/fisiopatología , Factores de Tiempo , Resultado del TratamientoRESUMEN
BACKGROUND/AIMS: Isradipine, a calcium channel blocker, provides consistent protection of the brain from injury and reduces neurological deficits produced by ischemic stroke in hypertensive rats. In these experiments, isradipine was utilized to cross-validate both the serial MRI measurement of brain infarctions with histology measurements and to validate a series of simple neurological deficit tests in order to establish a more rapid, higher throughput approach to screening compounds for utility in stroke. METHODS: Spontaneously hypertensive rats were treated with vehicle, or 2.5 or 5.0 mg/kg isradipine and middle cerebral artery occlusion. T(2)-weighted MRI image analysis was compared to standard triphenyltetrazolium chloride-stained histological image analysis of brain sections to quantify isradipine neuroprotection 1, 3, and 30 days after middle cerebral artery occlusion (MCAO; stroke). In addition, serial evaluation (i.e. 1, 2, 5, 12, 20 and 30 days after MCAO) of four simple neurobehavioral tests were completed for each animal. Tests included assessment of hindlimb and forelimb function, and balance beam and proprioception performance. RESULTS: At 1, 3 and 30 days there was a significant positive correlation of the percent hemispheric infarct for T(2)-weighted MRI and histology (p < 0.05). Practically identical isradipine dose-response neuroprotection curves were observed for both measurement procedures. Isradipine produced a dose-related reduction in all neurological deficits scored by the four neurological deficit tests (p < 0.05). In addition, a significant time-related recovery from neurological deficits in vehicle-treated rats was observed (p < 0.05). The four different neurological deficit tests did provide unique time-related profiles of neurological recovery. CONCLUSIONS: The present study validates the use of serial MRI in experimental stroke and establishes several simple neurological tests that can be used to measure neurological/behavioral deficits associated with brain injury and brain recovery of function over time. Under these conditions, T(2)-weighted MRI and neurological testing required only about 10 min each per animal, thus providing rapid data collection and analysis and requiring reduced scientific personnel.
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Conducta Animal/efectos de los fármacos , Infarto Encefálico , Bloqueadores de los Canales de Calcio/uso terapéutico , Isradipino/uso terapéutico , Imagen por Resonancia Magnética , Animales , Infarto Encefálico/tratamiento farmacológico , Infarto Encefálico/patología , Infarto Encefálico/fisiopatología , Bloqueadores de los Canales de Calcio/administración & dosificación , Modelos Animales de Enfermedad , Isradipino/administración & dosificación , Masculino , Pruebas Neuropsicológicas , Valor Predictivo de las Pruebas , Ratas , Ratas Endogámicas SHR , Resultado del TratamientoRESUMEN
Previously, it was shown that selective deletion of peroxisome proliferator activated receptor delta (PPARdelta) in the heart resulted in a cardiac lipotoxicity, hypertrophy, and heart failure. The aim of the present study was to determine the effects of chronic and selective pharmacological activation of PPARdelta in a model of congestive heart failure. PPARdelta-specific agonist treatment (GW610742X at 30 and 100 mg/kg/day for 6-9 weeks) was initiated immediately postmyocardial infarction (MI) in Sprague-Dawley rats. Magnetic resonance imaging/spectroscopy was used to assess cardiac function and energetics. A 1-(13)C glucose clamp was performed to assess relative cardiac carbohydrate versus fat oxidation. Additionally, cardiac hemodynamics and reverse-transcription polymerase chain reaction gene expression analysis was performed. MI rats had significantly reduced left ventricle (LV) ejection fractions and whole heart phosphocreatine/adenosine triphosphate ratio compared with Sham animals (reduction of 43% and 14%, respectively). However, GW610742X treatment had no effect on either parameter. In contrast, the decrease in relative fat oxidation rate observed in both LV and right ventricle (RV) following MI (decrease of 58% and 54%, respectively) was normalized in a dose-dependent manner following treatment with GW610742X. These metabolic changes were associated with an increase in lipid transport/metabolism target gene expression (eg, CD36, CPT1, UCP3). Although there was no difference between groups in LV weight or infarct size measured upon necropsy, there was a dramatic reduction in RV hypertrophy and lung congestion (decrease of 22-48%, P<0.01) with treatment which was associated with a >7-fold decrease (P<0.05) in aterial natriuretic peptide gene expression in RV. Diuretic effects were not observed with GW610742X. In conclusion, chronic treatment with a selective PPARdelta agonist normalizes cardiac substrate metabolism and reduces RV hypertrophy and pulmonary congestion consistent with improvement in congestive heart failure.
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Insuficiencia Cardíaca/tratamiento farmacológico , Hipertrofia Ventricular Derecha/tratamiento farmacológico , PPAR delta/agonistas , Animales , Transporte Biológico , Diuresis/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Metabolismo Energético , Expresión Génica/efectos de los fármacos , Insuficiencia Cardíaca/etiología , Insuficiencia Cardíaca/fisiopatología , Hipertrofia Ventricular Derecha/etiología , Hipertrofia Ventricular Derecha/fisiopatología , Lípidos/sangre , Espectroscopía de Resonancia Magnética , Masculino , Infarto del Miocardio/complicaciones , Oxidación-Reducción , PPAR delta/metabolismo , Edema Pulmonar/tratamiento farmacológico , Edema Pulmonar/etiología , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Función Ventricular Izquierda/efectos de los fármacosRESUMEN
Peroxisome proliferator-activated receptor-delta (PPARdelta) activation results in upregulation of genes associated with skeletal muscle fatty acid oxidation and mitochondrial uncoupling. However, direct, noninvasive assessment of lipid metabolism and mitochondrial energy coupling in skeletal muscle following PPARdelta stimulation has not been examined. Therefore, in this study we examined the response of a selective PPARdelta agonist (GW610742X at 5 or 100 mg.kg(-1).day(-1) for 8 days) on skeletal-muscle lipid metabolism and mitochondrial coupling efficiency in rats by using in vivo magnetic resonance spectroscopy (MRS). There was a decrease in the intramyocellular lipid-to-total creatine ratio as assessed by in vivo (1)H-MRS in soleus and tibialis anterior muscles by day 7 (reduced by 49 and 46%, respectively; P < 0.01) at the high dose. Following the (1)H-MRS experiment (day 8), [1-(13)C]glucose was administered to conscious rats to assess metabolism in the soleus muscle. The relative fat-vs.-carbohydrate oxidation rate increased in a dose-dependent manner (increased by 52 and 93% in the 5 and 100 mg.kg(-1).day(-1) groups, respectively; P < 0.05). In separate experiments where mitochondrial coupling was assessed in vivo (day 7), (31)P-MRS was used to measure hindlimb ATP synthesis and (13)C-MRS was used to measure the hindlimb tricarboxylic acid cycle flux (V(tca)). There was no alteration, at either dose, in mitochondrial coupling efficiency measured as the ratio of unidirectional ATP synthesis flux to V(tca). Soleus muscle GLUT4 expression was decreased by twofold, whereas pyruvate dehydrogenase kinase 4, carnitine palmitoyl transferase 1a, and uncoupling protein 2 and 3 expression was increased by two- to threefold at the high dose (P < 0.05). In summary, these are the first noninvasive measurements illustrating a selective PPARdelta-mediated decrease in muscle lipid content that was consistent with a shift in metabolic substrate utilization from carbohydrate to lipid. However, the mitochondrial-energy coupling efficiency was not altered in the presence of increased uncoupling protein expression.
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Isoindoles/farmacología , Mitocondrias Musculares/efectos de los fármacos , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , PPAR delta/agonistas , Sulfonamidas/farmacología , Animales , Colesterol/sangre , Ciclo del Ácido Cítrico/efectos de los fármacos , Ácidos Grasos no Esterificados/sangre , Expresión Génica , Glucosa/metabolismo , Transportador de Glucosa de Tipo 4/metabolismo , Metabolismo de los Lípidos/efectos de los fármacos , Espectroscopía de Resonancia Magnética , Mitocondrias Musculares/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Oxidación-Reducción , PPAR delta/farmacología , ARN/química , ARN/genética , Ratas , Ratas Sprague-Dawley , Triglicéridos/sangreRESUMEN
OBJECTIVE: To non-invasively characterize ectopic uterine tissue (EUT) development in a modified autologous rat surgical model of endometriosis using magnetic resonance imaging (MRI). DESIGN: Investigational MRI study. SETTING: A pharmaceutical company. ANIMAL(S): Female Sprague Dawley rats. INTERVENTION(S): Uterine tissue was autotransplanted on the right peritoneal wall of rats. Rats were serially imaged after surgery and after endogenous hormone suppression, hormone supplementation, or ovariectomy. In addition, an MRI contrast agent was administered to examine EUT perfusion characteristics. MAIN OUTCOME MEASURE(S): Changes in transplanted EUT volume and perfusion were monitored using MRI. RESULT(S): The EUT growth could be readily monitored non-invasively by MRI. Although EUT growth was rapid during the initial 4 days after surgery, volume stabilized by the third week and maintained for at least 9 weeks after transplantation. The EUT volumes varied with the estrous cycle and were hormonally sensitive to ovariectomy, to Antide (GnRH antagonist), and to Antide followed by 17beta-E(2) supplementation. The use of an MRI contrast agent facilitated visualization of EUT wall perfusion. CONCLUSION(S): MRI allows for noninvasive, dynamic evaluation of transplanted EUT growth in the rat. This reproducible model will allow for performing quantifiable pharmacologic studies in pre-clinical drug discovery for therapies targeting endometriosis.
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Coristoma/diagnóstico , Imagen por Resonancia Magnética/métodos , Cavidad Peritoneal , Útero , Animales , Coristoma/fisiopatología , Endometriosis/diagnóstico , Endometriosis/fisiopatología , Ciclo Estral/fisiología , Femenino , Ratas , Ratas Sprague-DawleyRESUMEN
PURPOSE: To compare atherosclerotic plaque uptake of a first (ferumoxtran-10) and second generation (ferumoxytol) ultrasmall superparamagnetic iron oxide (USPIO) contrast agent with different pharmacokinetic/pharmacodynamic properties. MATERIALS AND METHODS: New Zealand White rabbits maintained on a high cholesterol/fat diet were subjected to balloon injury to the abdominal aorta. Ferumoxtran-10 or ferumoxytol (500 micromol/kg) was administered at 2, 4, and 8 weeks following injury. In vivo magnetic resonance imaging (MRI) was performed immediately prior to, immediately after, and 6 days post-contrast administration. Ex vivo MRI, histologic, and inductively coupled plasma-mass spectrometry (ICP-MS) iron analyses were performed on the excised vessels. RESULTS: The blood pool clearance of ferumoxytol (t(1/2) < or = 6 hours) was more rapid than that of ferumoxtran-10 (t(1/2) < or = 48 hours). Decreased in vivo MRI signal intensity in the abdominal aorta was observed at 2, 4, and 8 weeks following injury with ferumoxtran-10, but not with ferumoxytol. Consistent with these observations, ex vivo MRI signal intensity was decreased in the ferumoxtran-10 vessels, and to a lesser degree in the ferumoxytol vs. control vessels (- contrast agent). In contrast, in vitro macrophage phagocytosis of USPIO was four to six fold greater with ferumoxytol than with ferumoxtran-10. Additionally, the absolute iron content correlated with ex vivo MRI signal intensity in all vessels (r = -0.86, P < 0.0001). CONCLUSIONS: These data suggest that the exposure period of atherosclerotic plaque to USPIO rather than the kinetics of the USPIO uptake by plaque alone is a critical criterion for experimental design of in vivo studies.
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Arteriosclerosis/diagnóstico , Medios de Contraste/farmacocinética , Hierro/farmacocinética , Imagen por Resonancia Magnética , Óxidos/farmacocinética , Animales , Arteriosclerosis/metabolismo , Dextranos , Óxido Ferrosoférrico , Macrófagos/metabolismo , Macrófagos/patología , Nanopartículas de Magnetita , Masculino , ConejosRESUMEN
Owing to its signal-enhancing characteristics in viable well-perfused tissue, divalent manganese (Mn2+) has been used as a myocardial imaging contrast agent. Because Mn2+ can enter excitable cells through the voltage-gated L-type calcium channels, manganese-enhanced MRI (MEMRI) has been used to determine the viability and the inotropic state of the heart. In this study, we examined the correlation between left ventricular infarction zone as assessed by cardiac MEMRI and function in mice with permanent coronary artery occlusion. At an Mn2+ infusion dose of 1.72+/-0.47 nmol/min/g body weight, the steady-state signal intensity (SI) enhancement 20-26 min post-Mn2+ infusion of the normal septum and left-ventricular wall during diastole was 128.2+/-14.4 and 127.9+/-26.5%, respectively, whereas the infarction zone was 56.0+/-7.1%. During systole, the SI enhancement was 144.6+/-33.0, 116.0+/-18.7 and 48.3+/-20.0% for the normal septum, left-ventricular wall and infarction zone, respectively. A good correlation was obtained between the MEMRI determined infarction volume and conventional histological TTC staining (r = 0.9582, p<0.01). There was also a strong negative correlation between MEMRI determined infarction percentage (compared with whole left ventricle) and ejection fraction (r = -0.94, p<0.05). These data suggest that the Mn2+ concentration at steady state in the heart may reflect altered tissue viability in the infarcted tissue as well as surrounding region following myocardial infarction. In conclusion, in vivo cardiac MEMRI offers a manner in which functional, pathologic and viability data may be obtained simultaneously in myocardial tissue.
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Medios de Contraste , Aumento de la Imagen/métodos , Interpretación de Imagen Asistida por Computador/métodos , Imagen por Resonancia Magnética/métodos , Manganeso , Infarto del Miocardio/diagnóstico , Disfunción Ventricular Izquierda/diagnóstico , Animales , Ratones , Ratones Endogámicos ICR , Infarto del Miocardio/complicaciones , Infarto del Miocardio/metabolismo , Índice de Severidad de la Enfermedad , Supervivencia Tisular , Disfunción Ventricular Izquierda/etiología , Disfunción Ventricular Izquierda/metabolismoRESUMEN
Recent evidence suggests p38 mitogen-activated protein kinase (MAPK) signal transduction plays an important role in the pathogenesis of progressive renal disease. Using dynamic contrast enhanced magnetic resonance imaging (MRI), we evaluated chronic treatment with a p38 MAPK inhibitor, trans-1-(4-hydroxycyclohexyl)-4-(4-fluorophenyl-methoxypyridimidin-4-yl)imidazole (SB-239063), on renal function in a hypertension model of progressing renal dysfunction. Spontaneously hypertensive-stroke prone rats were placed on a high salt/fat diet (SFD) or maintained on normal chow diet (ND). SFD animals with albuminuria at 4 to 8 weeks (> or =10 mg/day inclusion criteria), were randomized into p38 MAPK inhibitor treatment (SB-239063, 1200 ppm in diet) or vehicle groups. The progression of blood pressure and albuminuria during the treatment period (approximately 6 weeks) was decreased by 12 and 60%, respectively, in the SFD + SB-239063 versus SFD control group. Renal perfusion and filtration were assessed by in vivo MRI at the end of the study. Relative cortical perfusion was increased in the SFD + SB-239063 group compared with the SFD control group as reflected by a 29% decrease in time to peak of contrast agent in the cortex. Additionally, the regional renal glomerular filtration rate index (Kcl) was increased by 39% in the SFD + SB-239063 versus SFD control group and was normalized to the ND control group. Greater functional heterogeneity was observed in the SFD control versus SFD + SB-239063 or ND control group. All alterations of renal function were supported by histopathological findings. In conclusion, chronic treatment with a p38 MAPK inhibitor, SB-239063, attenuates functional and structural renal degeneration in a hypertensive model of established renal dysfunction.
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Endotelio Vascular/patología , Inhibidores Enzimáticos/uso terapéutico , Hipertensión/tratamiento farmacológico , Hipertensión/patología , Imidazoles/farmacología , Riñón/fisiopatología , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Pirimidinas/farmacología , Albuminuria/orina , Animales , Medios de Contraste , Creatinina/orina , Grasas de la Dieta/efectos adversos , Gadolinio DTPA , Tasa de Filtración Glomerular/efectos de los fármacos , Técnicas In Vitro , Riñón/patología , Pruebas de Función Renal , Imagen por Resonancia Magnética , Ratas , Ratas Endogámicas SHR , Sodio en la Dieta/efectos adversos , Accidente Cerebrovascular/genética , Accidente Cerebrovascular/fisiopatología , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
Ischemic preconditioning (PC) is a phenomenon whereby a brief exposure to ischemia renders a tissue more tolerant to a subsequent sustained ischemic insult. Animals of the Spontaneously Hypertensive (SHR) and the Spontaneously Hypertensive Stroke-Prone (SHR-SP) rat strains produce cerebral infarcts that are larger and more reproducible in size than infarcts of normotensive rats. This study compared the effects of PC in SHR and SHR-SP rats, under the hypothesis that PC may not be as effective in the SHR-SP, a strain genetically predisposed to stroke. There were two groups per strain, with between eight and ten animals each. The Precondition group (PC) had a 10 min occlusion of the middle cerebral artery on day -1. On the same day the Sham group (Sham) received sham surgery. On day 0, both groups underwent permanent occlusion of the middle cerebral artery. The ischemic lesion was measured on day 1 using T(2)-weighted magnetic resonance imaging. Percent hemispheric infarct was significantly reduced in SHR PC vs. SHR Sham, SHR-SP PC vs. SHR-SP Sham, and SHR PC vs. SHR-SP PC. Thus, rats of the SHR-SP strain respond to PC less markedly than SHR animals. Both models may now be used to elucidate the mechanisms underlying PC.