RESUMO
Polysorbate 80 (PS80) is commonly used in pre-clinical formulations. The dose threshold for cardiovascular (CV) changes and hypersensitivity reaction in the dog was assessed and compared to other species. PS80 was administered by intravenous (IV) bolus (.5, 1 mg/kg), IV infusion (.3, .5, 1, 3 mg/kg), subcutaneous (SC) injection (5, 10, 15 mg/kg) and oral gavage (10 mg/kg) to dogs with CV monitoring. Monkeys and minipigs received PS80 by IV infusion at 3 mg/kg. Plasma histamine concentration was measured following PS80 IV infusion and with diphenhydramine pre-treatment in dogs only. In dogs, PS80 was not associated with CV changes at doses up to 15 mg/kg SC and 10 mg/kg oral, but decreased blood pressure and increased heart rate with IV bolus at ≥ .5 mg/kg and IV infusion at ≥ 1.0 mg/kg and decreased body temperature with IV infusion at 3 mg/kg was observed. Transient edema and erythema were noted with all administration routes, in all three species including doses that were devoid of CV effects. In monkeys and minipigs, PS80 did not induce CV, cutaneous or histamine concentration changes. These results suggest that mild, transient skin changes occur following PS80 administration at doses that are not associated with CV effects in the dogs. In dogs, the cardiovascular effect threshold was <.5 mg/kg for IV bolus, .3 mg/kg for IV infusion, 15 mg/kg for SC injection, and 10 mg/kg for oral administration. Monkey and minipig were refractory to PS80-induced histamine release at 3 mg/kg by IV infusion over 15 minutes.
Assuntos
Anafilaxia , Polissorbatos , Anafilaxia/induzido quimicamente , Animais , Cães , Histamina , Injeções Intravenosas , Polissorbatos/toxicidade , Suínos , Porco MiniaturaRESUMO
Intrathecal administration is an important route for drug delivery, and in pharmacology and toxicology studies, cerebrospinal fluid (CSF) collection and analysis is required for evaluating blood-brain barrier penetration and central nervous system exposure. The characteristics of CSF in commonly used nonrodent models are lacking. The purpose of this study is to evaluate and provide some insights into normal cellular and biochemical composition of CSF as well as diffusion potential following intrathecal injection across several nonrodent species. Cerebrospinal fluid samples were collected from the cerebellomedullary cistern of beagle dogs, cynomolgus monkeys, and Göttingen minipigs and analyzed for clinical chemistry and cytological evaluation. Diffusion into the intrathecal space following intrathecal injection was assessed following administration of a contrast agent using fluoroscopy. The predominant cell types identified in CSF samples were lymphocytes and monocytoid cells; however, lymphocytes were represented in a higher percentage in dogs and monkeys as opposed to monocytoid cells in minipigs. Clinical chemistry parameters in CSF revealed higher Cl- concentrations than plasma, but lower K+, Ca2+, phosphorus, glucose, creatinine, and total protein levels consistent across all 3 species. Diffusion rates following intrathecal injection of iodixanol showed some variability with dogs, showing the greatest diffusion distance; however, the longest diffusion time through the intervertebral space, followed by monkeys and minipigs. Minimal diffusion was observed in minipigs, which could have been attributed to anatomical spinal constraints that have been previously identified in this species.
Assuntos
Líquido Cefalorraquidiano/química , Animais , Contagem de Células , Líquido Cefalorraquidiano/citologia , Meios de Contraste/farmacocinética , Cães , Feminino , Injeções Espinhais , Vértebras Lombares , Macaca fascicularis , Masculino , Suínos , Porco Miniatura , Ácidos Tri-Iodobenzoicos/líquido cefalorraquidiano , Ácidos Tri-Iodobenzoicos/farmacocinéticaRESUMO
BACKGROUND: Although extensive studies have investigated radiation-induced injuries in particular gastrointestinal (GI) segments, a systematic comparison among the different segments on the basis of mode, magnitude and mechanism has not been reported. Here, a comparative study of segment-specific molecular and cellular responses was performed on jejunum, ileum and colon obtained at three time points (4, 7 and 12 days after irradiation) from non-human primate (Rhesus macaque) models exposed to 6.7 Gy or 7.4 Gy total body irradiation (TBI). RESULTS: Pathway analysis on the gene expression profiles identified radiation-induced time-, dose- and segment-dependent activation of tumor necrosis factor α (TNFα) cascade, tight junction, apoptosis, cell cycle control/DNA damage repair and coagulation system signaling. Activation of these signaling pathways suggests that colon sustained the severest mucosal barrier disruption and inflammation, and jejunum the greatest DNA damage, apoptosis and endothelial dysfunction. These more pronounced alterations correlate with the high incidence of macroscopic pathologies that are observed in the colon after TBI. Compared to colon and jejunum, ileum was resistant to radiation injury. In addition to the identification a marked increase of TNFα cascade, this study also identified radiation induced strikingly up-regulated tight junction gene CLDN2 (196-fold after 7.4-Gy TBI), matrix degradation genes such as MMP7 (increased 11- and 41-fold after 6.7-Gy and 7.4-Gy TBI), and anoikis mediated gene EDA2R that mediate mucosal shedding and barrier disruption. CONCLUSIONS: This is the first systematic comparative study of the molecular and cellular responses to radiation injury in jejunum, ileum and colon. The strongest activation of TNFα cascades and the striking up-regulation of its down-stream matrix-dissociated genes suggest that TNFα modulation could be a target for mitigating radiation-induced mucosal barrier disruption.
Assuntos
Colo/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos da radiação , Íleo/metabolismo , Jejuno/metabolismo , Transcriptoma , Irradiação Corporal Total , Animais , Anoikis/genética , Apoptose/genética , Ciclo Celular , Análise por Conglomerados , Colo/imunologia , Íleo/imunologia , Imunidade nas Mucosas/genética , Imunidade nas Mucosas/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos da radiação , Jejuno/imunologia , Macaca mulatta , Masculino , Doses de Radiação , Lesões Experimentais por Radiação , Receptores de Hidrocarboneto Arílico/metabolismo , Transdução de Sinais/efeitos da radiação , Junções Íntimas/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Cynomolgus monkeys are widely used as models of diseases and in pre-clinical studies to assess the impact of new pharmacotherapies on brain function and behaviour. However, the time course of electroencephalographic delta activity during sleep, which represents the main marker of sleep intensity associated with recovery during sleep, has never been described in this non-human primate. In this study, telemetry implants were used to record one spontaneous 24-h sleep-wake cycle in four freely-moving Cynomolgus monkeys, and to quantify the time course of electroencephalographic activity during sleep using spectral analysis. Animals presented a diurnal activity pattern interrupted by short naps. During the dark period, most of the time was spent in sleep with non-rapid eye movement sleep/rapid eye movement sleep alternations and sleep consolidation profiles intermediate between rodents and humans. Deep non-rapid eye movement sleep showed a typical predominance at the beginning of the night with decreased propensity in the course of the night, which was accompanied by a progressive increase in rapid eye movement sleep duration. Spectral profiles showed characteristic changes between vigilance states as reported in other mammalian species. Importantly, delta activity also followed the expected time course of variation, showing a build-up with wakefulness duration and dissipation across the night. Thus, Cynomolgus monkeys present typical characteristics of sleep architecture and spectral structure as those observed in other mammalian species including humans, validating the use of telemetry in this non-human primate model for translational sleep studies.
Assuntos
Macaca fascicularis/fisiologia , Sono/fisiologia , Telemetria , Animais , Atenção/fisiologia , Atenção/efeitos da radiação , Escuridão , Eletroencefalografia , Humanos , Luz , Masculino , Modelos Animais , Polissonografia , Sono/efeitos da radiação , Sono REM/fisiologia , Sono REM/efeitos da radiação , Fatores de Tempo , Vigília/fisiologia , Vigília/efeitos da radiaçãoRESUMO
Diabetic retinopathy (DR) is a major microvascular complication associated with type 1 and type 2 diabetes mellitus, which can lead to visual impairment and blindness. Current treatment strategies for DR are mostly limited to laser therapies, steroids, and anti-VEGF agents, which are often associated with unwanted side effects leading to further complications. Recent evidence suggests that kinins play a primary role in the development of DR through enhanced vascular permeability, leukocytes infiltration, and other inflammatory mechanisms. These deleterious effects are mediated by kinin B1 and B2 receptors, which are expressed in diabetic human and rodent retina. Importantly, kinin B1 receptor is virtually absent in sane tissue, yet it is induced and upregulated in diabetic retina. These peptides belong to the kallikrein-kinin system (KKS), which contains two separate and independent pathways of regulated serine proteases, namely plasma kallikrein (PK) and tissue kallikrein (TK) that are involved in the biosynthesis of bradykinin (BK) and kallidin (Lys-BK), respectively. Hence, ocular inhibition of kallikreins or antagonism of kinin receptors offers new therapeutic avenues in the treatment and management of DR. Herein, we present an overview of the principal features and known inflammatory mechanisms associated with DR along with the current therapeutic approaches and put special emphasis on the KKS as a new and promising therapeutic target due to its link with key pathways directly associated with the development of DR.
Assuntos
Retinopatia Diabética/enzimologia , Calicreínas/metabolismo , Cininas/metabolismo , Retina/enzimologia , Animais , Anti-Inflamatórios/uso terapêutico , Retinopatia Diabética/tratamento farmacológico , Retinopatia Diabética/fisiopatologia , Humanos , Retina/efeitos dos fármacos , Retina/fisiopatologia , Vasos Retinianos/efeitos dos fármacos , Vasos Retinianos/enzimologia , Vasos Retinianos/fisiopatologia , Transdução de Sinais , Visão OcularRESUMO
The vasoactive kinin B1 receptor (B1R) is overexpressed in the retina of diabetic rats in response to hyperglycemia and oxidative stress. The aim of the present study was to determine whether B1R could contribute to the early retinal blood flow changes occurring in diabetes. Male Wistar rats were rendered diabetic with a single i.p. injection of Streptozotocin (STZ) and studied 4 days or 6 weeks after diabetes induction. The presence of B1R in the retina was confirmed by Western blot. The impact of oral administration of the B1R selective antagonist SSR240612 (10mg/kg) was measured on alteration of retinal perfusion in awake diabetic rats by quantitative autoradiography. Data showed that B1R was upregulated in the STZ-diabetic retina at 4 days and 6 weeks. Retinal blood flow was not altered in 4-day diabetic rats compared with age-matched controls but was significantly decreased following SSR240612 treatment. In 6-week diabetic rats, retinal blood flow was markedly reduced compared to control rats and SSR240612 did not further decrease the blood flow. These results suggest that B1R is upregulated in STZ-diabetic retina and has a protective compensatory role on retinal microcirculation at 4 days but not at 6 weeks following diabetes induction.
Assuntos
Diabetes Mellitus Experimental/fisiopatologia , Retinopatia Diabética/fisiopatologia , Receptor B1 da Bradicinina/metabolismo , Fluxo Sanguíneo Regional/fisiologia , Vasos Retinianos/fisiologia , Animais , Autorradiografia , Velocidade do Fluxo Sanguíneo , Western Blotting , Antagonistas de Receptor B1 da Bradicinina , Circulação Cerebrovascular/fisiologia , Diabetes Mellitus Experimental/metabolismo , Retinopatia Diabética/metabolismo , Dioxóis/farmacologia , Angiofluoresceinografia , Masculino , Ratos , Ratos Wistar , Sulfonamidas/farmacologia , Regulação para CimaRESUMO
BACKGROUND: High dose ionizing radiation exposure is associated with myelo-depression leading to pancytopenia and the expected clinical manifestations of acute radiation syndrome (ARS). Herein, we evaluated the efficacy of sargramostim (Leukine®, yeast-derived rhu GM-CSF), with regimens delivered at 48, 72, 96, or 120 h after radiation exposure. METHODS: A randomized and blinded nonhuman primate (NHP) study was conducted to assess the effects of sargramostim treatment on ARS. NHPs were exposed to total body radiation (LD83/60 or lethal dose 83% by Day 60) and were randomized to groups receiving daily subcutaneous dosing of sargramostim starting from either 48, 72, 96, or 120 h post-irradiation. Additionally, separate groups receiving sargramostim treatment at 48 h post-irradiation also received prophylactic treatment with azithromycin. Sargramostim treatment of each animal continued until the preliminary absolute neutrophil count (ANC) returned to ≥1000/µL post-nadir for three consecutive days or the preliminary ANC exceeded 10,000/µL, which amounted to be an average of 15.95 days for all treatment groups. Prophylactic administration of enrofloxacin was included in the supportive care given to all animals in all groups. All animals were monitored for 60 days post-irradiation for mortality, hematological parameters, and sepsis. RESULTS: Delayed sargramostim treatment at 48 h post-irradiation significantly reduced mortality (p = .0032) and improved hematological parameters including neutrophil but also lymphocyte and platelet counts. Additional delays in sargramostim administration at 72, 96, and 120 h post-irradiation were also similarly effective at enhancing the recovery of lymphocyte, neutrophil, and platelet counts compared to control. Sargramostim treatment also improved the survival of the animals when administered at up to 96 h post-irradiation. While sargramostim treatment at 48 h significantly reduced mortality associated with sepsis (p ≤ .01), the additional prophylactic treatment with azithromycin did not have clinically significant effects. CONCLUSION: In a NHP ARS model, sargramostim administered starting at 48 h post-radiation was effective to improve survival, while beneficial hematological effects were observed with sargramostim initiated up to 120 h post exposure.
Assuntos
Síndrome Aguda da Radiação , Sepse , Animais , Síndrome Aguda da Radiação/tratamento farmacológico , Azitromicina/uso terapêutico , Fator Estimulador de Colônias de Granulócitos e Macrófagos/uso terapêutico , Macaca mulatta , Proteínas Recombinantes , Sepse/tratamento farmacológicoRESUMO
PURPOSE: Diabetic retinopathy is characterized by multiple microcirculatory dysfunctions and angiogenesis resulting from hyperglycemia, oxidative stress, and inflammation. In this study, the retina and retinal pigmented epithelium of non-insulin-dependent diabetic Goto-Kakizaki (GK) rats were examined to detect microvascular alterations, gliosis, macrophage infiltration, lipid deposits, and fibrosis. Emphasis was given to the distribution of kinin B1 receptor (B1R) and vascular endothelial growth factor (VEGF), two major factors in inflammation and angiogenesis. MATERIALS AND METHODS: 30-week-old male GK rats and age-matched Wistar rats were used. The retinal vascular bed was examined using ADPase staining. The level of lipid accumulation was graded using triglyceride staining with Oil red O. Macrophage and retinal microglia activation, as well as other markers, were revealed by immunohistochemistry and studied with confocal laser scanning microscopy. RESULTS: Abundant lipid deposits were observed in the Bruch's membrane of GK rats. Immunohistochemistry and quantitative analysis showed significantly higher B1R, VEGF, Iba1 (microglia), CD11 (macrophages), fibronectin, and collagen I labeling in the diabetic retina. B1R immunolabeling was detected in the vascular layers of the GK retina. A strong VEGF staining within different retinal cell processes was detected and a pattern of GFAP staining suggested strong Müller cells/astrocytes reactivity. Microgliosis was apparent in the GK retina. A greater tortuosity of the retinal microvessels (an index of endothelial dysfunction) and their increased number were also observed in GK retinas. CONCLUSIONS: Data suggest retinal vascular bed alterations in spontaneous type 2 diabetic retinas at 30 weeks. Lipid and collagen accumulation in the retina and choroid, in addition to retinal upregulation of VEGF and B1R, microgliosis, and Müller cell reactivity, may contribute to vascular alterations and inflammatory processes.
Assuntos
Diabetes Mellitus Tipo 2/patologia , Retinopatia Diabética/patologia , Vasos Retinianos/patologia , Retinite/patologia , Animais , Colágeno Tipo I/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Retinopatia Diabética/metabolismo , Modelos Animais de Doenças , Fibronectinas/metabolismo , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/patologia , Imuno-Histoquímica , Inflamação/metabolismo , Inflamação/patologia , Metabolismo dos Lipídeos , Macrófagos/patologia , Masculino , Microscopia Confocal , Ratos Mutantes , Ratos Wistar , Receptor B1 da Bradicinina/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/patologia , Vasos Retinianos/metabolismo , Retinite/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Purpose: Characterization of a novel partial-body irradiation (PBI) shielding strategy in nonhuman primates (NHP; rhesus macaques), aimed at protecting the oral cavity, with respect to various gastrointestinal acute radiation syndrome (GI-ARS) syndrome parameters as well as buccal ulceration development.Materials and methods: NHPs were irradiated using a Cobalt-60 gamma source, in a single uniform dose, ranging from 9-13 Gy and delivered at 0.60-0.80 Gy min-1. Animals were either partially shielded via oral cavity shielding (PBIOS) or underwent total-body irradiation (TBI).Results: Clinical manifestations of GI-ARS, and also radiation-induced hematology and clinical chemistry changes, following PBIOS were comparable to the PBI NHP GI-ARS model utilizing shielding of the distal pelvic limbs and were significantly milder than TBI at similar radiation doses. Nadir citrulline levels were comparable between PBIOS and TBI but signs of recovery appeared earlier in PBIOS-treated animals. The PBIOS model prevented oral mucositis, whereas the TBI model presented buccal ulcerations at all tested radiation dose levels.Conclusions: Taken together, these results suggest that the PBIOS model is a suitable alternative to traditional PBI. For GI-ARS investigations requiring orally administered medical countermeasures, PBIOS confers added value due to the prevention of oral mucositis over traditional PBI.
Assuntos
Boca/efeitos da radiação , Proteção Radiológica/métodos , Síndrome Aguda da Radiação/sangue , Síndrome Aguda da Radiação/etiologia , Síndrome Aguda da Radiação/patologia , Animais , Citrulina/sangue , Radioisótopos de Cobalto/efeitos adversos , Raios gama/efeitos adversos , Macaca mulatta , Masculino , Análise de Sobrevida , Úlcera/sangue , Úlcera/etiologia , Úlcera/patologiaRESUMO
Quantitative and regional measurement of retinal blood flow in rodents is of prime interest for the investigation of regulatory mechanisms of ocular circulation in physiological and pathological conditions. In this study, a quantitative autoradiographic method using N-isopropyl-p-(14)C-iodoamphetamine ([(14)C]-IMP), a diffusible radioactive tracer, was evaluated for its ability to detect changes in retinal blood perfusion during hypercapnia. Findings were compared to cerebral blood flow values measured simultaneously. Hypercapnia was induced in awaken Wistar rats by inhalation of 5% or 8% CO(2) in medical air for 5 min. [(14)C]-IMP (100 microCi/kg) was injected in the femoral vein over a 30 s period and the rats were sacrificed 2 min later. Blood flow was calculated from whole-mount retinae and 20 microm thick brain sections in discrete regions of interest by quantitative autoradiography or from digested samples of retina and brain by liquid scintillation counting. Retinal blood flow values measured with quantitative and regional autoradiography were higher in the central (108 +/- 20 ml/100 g/min) than in peripheral (84 +/- 15 ml/100 g/min) retina. These values were within the same range as cortical blood flow values (97 +/- 4 ml/100 g/min). The retinal blood flow values obtained on whole-mount retinae were validated by the sampling method. Hypercapnia significantly increased overall blood flow in the retina (24-53%) with a maximal augmentation in the peripheral region and in the brain (22-142%). The changes were stronger in the brain compared to retina (p = 0.016). These results demonstrate that retinal blood flow can be quantified using [(14)C]-IMP and compared with cerebral blood flow. This technique is a powerful tool to study how retinal blood flow is regulated in different regions of the rat retina.
Assuntos
Vasos Retinianos/fisiologia , Animais , Autorradiografia/métodos , Dióxido de Carbono/sangue , Radioisótopos de Carbono , Circulação Cerebrovascular/fisiologia , Hipercapnia/sangue , Hipercapnia/fisiopatologia , Iofetamina , Masculino , Microcirculação/fisiologia , Disco Óptico/irrigação sanguínea , Pressão Parcial , Ratos , Ratos Wistar , Vasos Retinianos/fisiopatologiaRESUMO
Seizures are amongst the most frequent neurological issues encountered in pre-clinical safety testing. The objective was to characterize EEG morphologies and premonitory signs in drug-induced seizures in preclinical species. A comparative (inter-species) retrospective analysis for drug-induced seizures recorded by video-telemetry was conducted in rats (nâ¯=â¯53), dogs (nâ¯=â¯195), and non-human primates (nâ¯=â¯234). The most frequent premonitory signs were, in rats, myoclonus (100%), tremors (93%), salivation (75%), partial ptosis (58%) and chewing/bruxism (58%); in dogs, tremors (77%), ataxia/uncoordination (60%), myoclonus (45%), salivation (43%), excessive licking (38%), high vocalization (38%) and decreased activity (34%); in non-human primates, tremors (79%), decreased activity (70%), myoclonus (57%), retching/emesis (37%), hunched posture (30%) and ataxia/uncoordination (27%). Seizure duration ranged from 3â¯s to 14â¯min with an average of 46⯱â¯21â¯s, comparable across species. At seizure onset, spike frequency averaged 9.4â¯Hz for the three species compared to 4.3â¯Hz at seizure end. Peak average amplitudes were attained at mid-seizure and amplitudes at seizure end decreased from peak but remained higher than onset amplitudes. Spike duration was inversely correlated with frequency and presented a crescendo pattern. Morphological characteristics can serve to refine automated EEG analysis. From a regulatory perspective, the most common paradigm is to use the most sensitive species in seizure liability studies but translational potential and clinical relevance may be under represented in the decision making process in some cases. EEG morphologies during drug-induced seizures presented remarkable similarities between species and tremors were identified as a predominant premonitory clinical sign in all species.
Assuntos
Convulsões/induzido quimicamente , Convulsões/fisiopatologia , Animais , Comportamento Animal/fisiologia , Cães , Avaliação Pré-Clínica de Medicamentos/métodos , Eletroencefalografia/métodos , Macaca fascicularis , Primatas , Ratos , Ratos Sprague-Dawley , Estudos Retrospectivos , Telemetria/métodosRESUMO
Seizures are amongst the most frequent neurological issues encountered in pre-clinical safety testing. The objective was to characterize EEG morphologies and premonitory signs in drug-induced seizures in preclinical species. A comparative (inter-species) retrospective analysis for drug-induced seizures recorded by video-telemetry was conducted in rats (nâ¯=â¯53), dogs (nâ¯=â¯195), and non-human primates (nâ¯=â¯234). The most frequent premonitory signs were, in rats, myoclonus (100%), tremors (93%), salivation (75%), partial ptosis (58%) and chewing/bruxism (58%); in dogs, tremors (77%), ataxia/uncoordination (60%), myoclonus (45%), salivation (43%), excessive licking (38%), high vocalization (38%) and decreased activity (34%); in non-human primates, tremors (79%), decreased activity (70%), myoclonus (57%), retching/emesis (37%), hunched posture (30%) and ataxia/uncoordination (27%). Seizure duration ranged from 3â¯s to 14â¯min with an average of 46⯱â¯21â¯s, comparable across species. At seizure onset, spike frequency averaged 9.4â¯Hz for the three species compared to 4.3â¯Hz at seizure end. Peak average amplitudes were attained at mid-seizure and amplitudes at seizure end decreased from peak but remained higher than onset amplitudes. Spike duration was inversely correlated with frequency and presented a crescendo pattern. Morphological characteristics can serve to refine automated EEG analysis. From a regulatory perspective, the most common paradigm is to use the most sensitive species in seizure liability studies but translational potential and clinical relevance may be under represented in the decision making process in some cases. EEG morphologies during drug-induced seizures presented remarkable similarities between species and tremors were identified as a predominant premonitory clinical sign in all species.
RESUMO
Spontaneous arrhythmia characterization in healthy rats can support interpretation when studying novel therapies. Male (nâ¯=â¯55) and female (nâ¯=â¯40) Sprague-Dawley rats with telemetry transmitters for a derivation II ECG. Arrhythmias were assessed from continuous ECG monitoring over a period of 24-48â¯h, and data analyzed using an automated detection algorithm with 100% manual over-read. While a total of 1825 spontaneous ventricular premature beats (VPB) were identified, only 7 rats (or 7.4%) did not present with any over the recording period. Spontaneous episode(s) of ventricular tachycardia (VT) were noted in males (27%) and females (3%). The incidence of VPB was significantly higher (pâ¯<â¯0.01) during the night time (7â¯pm-7â¯am) compared to daytime, while males presented with significantly (pâ¯<â¯0.001) more VPB than females. Most VPB were observed as single ectopic beats, followed by salvos (2 or 3 consecutive VPBs), and VT (i.e. 4 consecutive VPBs). Most VPBs were single premature ventricular contractions (PVCs) (57%), while the remaining were escape complexes (43%). Spontaneous premature junctional complexes (PJC) were also observed and were significantly more frequent during the night, and in males. Lastly, 596 episodes of spontaneous 2nd-degree atrioventricular (AV) block were identified and were significantly more frequent during the day time in males. Most 2nd-degree AV block episodes were Mobitz type I (57%), with a significantly (pâ¯<â¯0.05) higher incidence in males. This work emphasizes the importance of obtaining sufficient baseline data when undertaking arrhythmia analysis in safety study and provides a better understanding of both sex- and time- dependent effects of spontaneous arrhythmias in rats.
RESUMO
Acute radiation syndrome (ARS) is a complex multi-organ disease resulting from total body exposure to high doses of radiation. Individuals can be exposed to total body irradiation (TBI) in a number of ways, including terrorist radiological weapons or nuclear accidents. In order to determine whether an individual has been exposed to high doses of radiation and needs countermeasure treatment, robust biomarkers are needed to estimate radiation exposure from biospecimens such as blood or urine. In order to identity such candidate biomarkers of radiation exposure, high-resolution proteomics was used to analyze plasma from non-human primates following whole body irradiation (Co-60 at 6.7 Gy and 7.4 Gy) with a twelve day observation period. A total of 663 proteins were evaluated from the plasma proteome analysis. A panel of plasma proteins with characteristic time- and dose-dependent changes was identified. In addition to the plasma proteomics study reported here, we recently identified candidate biomarkers using urine from these same non-human primates. From the proteomic analysis of both plasma and urine, we identified ten overlapping proteins that significantly differentiate both time and dose variables. These shared plasma and urine proteins represent optimal candidate biomarkers of radiation exposure.
Assuntos
Proteínas Sanguíneas/análise , Proteoma/efeitos da radiação , Proteômica/métodos , Irradiação Corporal Total/métodos , Animais , Biomarcadores/sangue , Proteínas Sanguíneas/classificação , Análise por Conglomerados , Relação Dose-Resposta à Radiação , Eletroforese em Gel de Poliacrilamida , Raios gama , Macaca mulatta , Espectrometria de Massas/métodos , Proteoma/classificação , Transdução de Sinais/efeitos da radiação , Fatores de TempoRESUMO
Systemic C-fiber blockade (CFB) has been reported to inhibit induced swallowing in adult guinea pigs. The aim of this study was to investigate the effects of CFB on spontaneous, non-nutritive swallowing (NNS) frequency and NNS-respiration coordination in the neonatal period. Seven CFB lambs and seven control lambs aged 2+/-1 days were chronically instrumented for recording electroencephalogram, eye movements, diaphragm EMG, thyroarytenoid muscle EMG, nasal airflow and electrocardiogram. Polysomnographic recordings were performed in non-sedated lambs, using radiotelemetry transmission. CFB lambs spent more time in active sleep than controls (p=0.02). Frequency of non-nutritive swallowing was not different in CFB and control lambs, whatever the state of alertness. In addition, CFB did not disrupt the overall respiratory-swallowing coordination, inspiratory-related NNS being the most frequent and expiratory-related NNS the least in both CFB and control lambs. Further analyses revealed that CFB had no effect on baseline respiratory and heart rate, and apnea and sigh frequency, whatever the state of alertness. Our results suggest that, in the neonatal period, C-fibers are not involved in NNS frequency and have no influence on the overall respiratory-swallowing coordination.
Assuntos
Animais Recém-Nascidos/fisiologia , Deglutição/fisiologia , Bloqueio Nervoso/métodos , Fibras Nervosas Amielínicas/fisiologia , Animais , Capsaicina/farmacologia , Diafragma/fisiologia , Eletroencefalografia/métodos , Eletromiografia/métodos , Fibras Nervosas Amielínicas/efeitos dos fármacos , Polissonografia/métodos , Respiração , Ovinos , Sono/efeitos dos fármacos , Sono/fisiologiaRESUMO
Dysfunction of the intestinal epithelial barrier and leakage of luminal antigens and bacteria across the barrier have been linked to various human diseases. Intestinal permeability is regulated by intercellular structures, termed "tight junction" (Tj), which are disrupted after total-body irradiation (TBI). In this study, we investigated radiation-induced alterations in Tj-related proteins in the jejunum, ileum and colon of a non-human primate (NHP) model. NHPs were total-body irradiated with 6.7 and 7.4 Gy and intestines were procured at day 4, 7 and 12. Radiation exposure was found to induce significant increases in claudin-10 mRNA early (day 4) in all three gut segments and claudin-4 mRNA levels were repressed through day 12. TNF-alpha was highly induced in the jejunum and colon at early time points, but little induction was found in the ileum. Claudin-1 was induced only in the colon on day 4 postirradiation. Unlike the colon and jejunum, the ileum levels of claudin-7 were significantly downregulated through day 12 postirradiation. Western blot analysis revealed increased levels of claudin-2 on day 4 and of JAM-1 on day 7 postirradiation in all three gut segments. E-cadherin was downregulated on day 4 postirradiation in all segments, but remained reduced in the jejunum only until day 12. Taken together, these data suggest that exposure to radiation causes segment-specific alterations in the expression of Tj-related proteins. Interruption of Tjs may be a key factor contributing to injury to the intestinal mucosal barrier and increased intestinal permeability.
Assuntos
Absorção Intestinal/fisiologia , Absorção Intestinal/efeitos da radiação , Mucosa Intestinal/metabolismo , Intestinos/efeitos da radiação , Doses de Radiação , Proteínas de Junções Íntimas/metabolismo , Animais , Colo/metabolismo , Colo/efeitos da radiação , Relação Dose-Resposta à Radiação , Feminino , Regulação da Expressão Gênica/fisiologia , Regulação da Expressão Gênica/efeitos da radiação , Humanos , Íleo/metabolismo , Íleo/efeitos da radiação , Jejuno/metabolismo , Jejuno/efeitos da radiação , Macaca mulatta , Masculino , Especificidade de Órgãos/efeitos dos fármacos , Especificidade de Órgãos/fisiologia , Distribuição TecidualRESUMO
The molecular effects of total body gamma-irradiation exposure are of critical importance as large populations of people could be exposed either by terrorists, nuclear blast, or medical therapy. In this study, we aimed to identify changes in the urine proteome using a non-human primate model system, Rhesus macaque, in order to characterize effects of acute radiation syndrome following whole body irradiation (Co-60) at 6.7 Gy and 7.4 Gy with a twelve day observation period. The urine proteome is potentially a valuable and non-invasive diagnostic for radiation exposure. Using high-resolution mass spectrometry, we identified 2346 proteins in the urine proteome. We show proteins involved in disease, cell adhesion, and metabolic pathway were significantly changed upon exposure to differing levels and durations of radiation exposure. Cell damage increased at a faster rate at 7.4 Gy compared with 6.7 Gy exposures. We report sets of proteins that are putative biomarkers of time- and dose-dependent radiation exposure. The proteomic study presented here is a comprehensive analysis of the urine proteome following radiation exposure.
RESUMO
INTRODUCTION: Continuous video-electroencephalographic (EEG) monitoring remains the gold standard for seizure liability assessments in preclinical drug safety assessments. EEG monitored by telemetry was used to assess the behavioral and EEG effects of noribogaine hydrochloride (noribogaine) in cynomolgus monkeys. Noribogaine is an iboga alkaloid being studied for the treatment of opioid dependence. METHODS: Six cynomolgus monkeys (3 per gender) were instrumented with EEG telemetry transmitters. Noribogaine was administered to each monkey at both doses (i.e., 160 and 320mg/kg, PO) with an interval between dosing of at least 6days, and the resulting behavioral and EEG effects were evaluated. IV pentylenetetrazol (PTZ), served as a positive control for induced seizures. RESULTS: The administration of noribogaine at either of the doses evaluated was not associated with EEG evidence of seizure or with EEG signals known to be premonitory signs of increased seizure risk (e.g., sharp waves, unusual synchrony, shifts to high-frequency patterns). Noribogaine was associated with a mild reduction in activity levels, increased scratching, licking and chewing, and some degree of poor coordination and related clinical signs. A single monkey exhibited brief myoclonic movements that increased in frequency at the high dose, but which did not appear to generalize, cluster or to be linked with EEG abnormalities. Noribogaine was also associated with emesis and partial anorexia. In contrast, PTZ was associated with substantial pre-ictal EEG patterns including large amplitude, repetitive sharp waves leading to generalized seizures and to typical post-ictal EEG frequency attenuation. INTERPRETATION: EEG patterns were within normal limits following administration of noribogaine at doses up to 320mg/kg with concurrent clinical signs that correlated with plasma exposures and resolved by the end of the monitoring period. PTZ was invariably associated with EEG paroxysmal activity leading to ictal EEG. In the current study, a noribogaine dose of 320mg/kg was considered to be the EEG no observed adverse effect level (NOAEL) in conscious freely moving cynomolgus monkeys.
Assuntos
Eletroencefalografia/efeitos dos fármacos , Ibogaína/análogos & derivados , Síndromes Neurotóxicas/psicologia , Animais , Comportamento Animal/efeitos dos fármacos , Convulsivantes , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Feminino , Ibogaína/farmacocinética , Ibogaína/toxicidade , Macaca fascicularis , Masculino , Atividade Motora/efeitos dos fármacos , Pentilenotetrazol , Convulsões/induzido quimicamente , TelemetriaRESUMO
The non-human primate has been a useful model for studies of human acute radiation syndrome (ARS). However, to date structural changes in various parts of the intestine after total body irradiation (TBI) have not been systematically studied in this model. Here we report on our current study of TBI-induced intestinal structural injury in the non-human primate after doses typically associated with hematopoietic ARS. Twenty-four non-human primates were divided into three groups: sham-irradiated control group; and total body cobalt-60 (60Co) 6.7 Gy gamma-irradiated group; and total body 60Co 7.4 Gy gamma-irradiated group. After animals were euthanized at day 4, 7 and 12 postirradiation, sections of small intestine (duodenum, proximal jejunum, distal jejunum and ileum) were collected and fixed in 10% formalin. The intestinal mucosal surface length, villus height and crypt depths were assessed by computer-assisted image analysis. Plasma citrulline levels were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Total bone marrow cells were counted and hematopoietic stem/progenitor cells in bone marrow were analyzed by flow cytometer. Histopathologically, all segments exhibited conspicuous disappearance of plicae circulares and prominent atrophy of crypts and villi. Intestinal mucosal surface length was significantly decreased in all intestinal segments on day 4, 7 and 12 after irradiation (P < 0.02-P < 0.001). Villus height was significantly reduced in all segments on day 4 and 7 (P = 0.02-0.005), whereas it had recovered by day 12 (P > 0.05). Crypt depth was also significantly reduced in all segments on day 4, 7 and 12 after irradiation (P < 0.04-P < 0.001). Plasma citrulline levels were dramatically reduced after irradiation, consistent with intestinal mucosal injury. Both 6.7 and 7.4 Gy TBI reduced total number of bone marrow cells. And further analysis showed that the number and function of CD45(+)CD34(+) hematopoietic stem/progenitors in bone marrow decreased significantly. In summary, TBI in the hematopoietic ARS dose range induces substantial intestinal injury in all segments of the small bowel. These findings underscore the importance of maintaining the mucosal barrier that separates the gut microbiome from the body's interior after TBI.
Assuntos
Hematopoese/efeitos da radiação , Intestino Delgado/lesões , Intestino Delgado/efeitos da radiação , Irradiação Corporal Total/efeitos adversos , Animais , Citrulina/sangue , Relação Dose-Resposta à Radiação , Feminino , Macaca mulatta , Masculino , Fatores de TempoRESUMO
INTRODUCTION: Medication-induced sleep disturbances are a major concern in drug development as a multitude of prescription drugs alter sleep patterns, often negatively. Polysomnography is used in clinical diagnostics but is also applicable to animal models. Rodent sleep architecture (nocturnal) differs from larger diurnal mammals, including humans, increasing the translational potential of non-rodent species to the clinic. This study aimed to characterize the response to pharmacological agents known to affect sleep structure and EEG activity in a non-human primate (Macaca fascicularis) using telemetry-based polysomnography. METHODS: Animals were instrumented with telemetry transmitters for continuous electroencephalogram (EEG), electro-oculogram (EOG) and electromyogram (EMG) monitoring combined with video. EEG, EMG and EOG were monitored for 12 to 24h to establish baseline values, followed by administration of pharmacological agents (saline, d-amphetamine, diazepam or caffeine). RESULTS: Amphetamine (0.3 and 1mg/kg, by oral administration (PO)) significantly reduced total sleep time, including the duration of both non-rapid eye movement [NREM] sleep and REM sleep. It also decreased EEG activity in low frequencies (i.e., 4-6Hz) during wakefulness. Diazepam (2mg/kg, PO) did not significantly alter sleep duration, but importantly reduced EEG activity in low frequencies (approximately 2-12Hz) during wakefulness, NREM and REM sleep. Finally, caffeine (10 and 30mg/kg, PO) decreased both NREM and REM sleep duration. In addition, spectral analysis revealed important decreases in low frequency activity (i.e., 1-8Hz) during wakefulness with a parallel increase in high frequency activity (i.e., 20-50Hz) during NREM sleep. DISCUSSION: As these observations are similar to previously reported pharmacological effects in humans, results support that EEG, EOG and EMG monitoring by telemetry in Cynomolgus monkeys represents a useful non-clinical model to investigate and quantify drug-induced sleep disturbances.