RESUMEN
Inhaled nitric oxide (iNO) is a therapy used in neonates with pulmonary hypertension. Some evidence of its neuroprotective properties has been reported in both mature and immature brains subjected to injury. NO is a key mediator of the VEGF pathway, and angiogenesis may be involved in the reduced vulnerability to injury of white matter and the cortex conferred by iNO. Here, we report the effect of iNO on angiogenesis in the developing brain and its potential effectors. We found that iNO promotes angiogenesis in the developing white matter and cortex during a critical window in P14 rat pups. This shift in the developmental program of brain angiogenesis was not related to a regulation of NO synthases by exogenous NO exposure, nor the VEGF pathway or other angiogenic factors. The effects of iNO on brain angiogenesis were found to be mimicked by circulating nitrate/nitrite, suggesting that these carriers may play a role in transporting NO to the brain. Finally, our data show that the soluble guanylate cyclase/cGMP signaling pathway is likely to be involved in the pro-angiogenetic effect of iNO through thrombospondin-1, a glycoprotein of the extracellular matrix, inhibiting soluble guanylate cyclase through CD42 and CD36. In conclusion, this study provides new insights into the biological basis of the effect of iNO in the developing brain.
Asunto(s)
Óxido Nítrico , Roedores , Animales , Ratas , Óxido Nítrico/metabolismo , Animales Recién Nacidos , Roedores/metabolismo , Guanilil Ciclasa Soluble/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Encéfalo/metabolismo , Administración por InhalaciónRESUMEN
Intrauterine growth restriction (IUGR) increases the risk of bronchopulmonary dysplasia (BPD), one of the major complications of prematurity. Antenatal low-protein diet (LPD) exposure in rats induces IUGR and mimics BPD-related alveolarization disorders. Peroxisome proliferator-activated receptor-γ (PPARγ) plays a key role in normal lung development and was found deregulated following LPD exposure. The objective of this article was to investigate the effects of nebulized curcumin, a natural PPARγ agonist, to prevent IUGR-related abnormal lung development. We studied rat pups antenatally exposed to an LPD or control diet (CTL) and treated with nebulized curcumin (50 mg/kg) or vehicle from postnatal (P) days 1 to 5. The primary readouts were lung morphometric analyses at P21. Immunohistochemistry (P21) and microarrays (P6 and P11) were compared within animals exposed to LPD versus controls, with and without curcumin treatment. Quantitative morphometric analyses revealed that LPD induced abnormal alveolarization as evidenced by a significant increase in mean linear intercept (MLI) observed in P21 LPD-exposed animals. Early curcumin treatment prevented this effect, and two-way ANOVA analysis demonstrated significant interaction between diet and curcumin both for MLI [F(1,39) = 12.67, P = 0.001] and radial alveolar count at P21 [F(1,40) = 6.065, P = 0.0182]. Immunohistochemistry for fatty acid binding protein 4 (FABP4), a major regulator of PPARγ pathway, showed a decreased FABP4+ alveolar cell density in LPD-exposed animals treated by curcumin. Transcriptomic analysis showed that early curcumin significantly prevented the activation of profibrotic pathways observed at P11 in LPD-exposed animals. Nebulized curcumin appears to be a promising strategy to prevent alveolarization disorders in IUGR rat pups, targeting pathways involved in lung development.
Asunto(s)
Displasia Broncopulmonar/prevención & control , Curcumina/farmacología , Dieta con Restricción de Proteínas/efectos adversos , Alveolos Pulmonares/metabolismo , Animales , Animales Recién Nacidos , Displasia Broncopulmonar/etiología , Displasia Broncopulmonar/metabolismo , Displasia Broncopulmonar/patología , Femenino , Retardo del Crecimiento Fetal/tratamiento farmacológico , Retardo del Crecimiento Fetal/etiología , Retardo del Crecimiento Fetal/metabolismo , Retardo del Crecimiento Fetal/patología , Masculino , Nebulizadores y Vaporizadores , PPAR gamma/agonistas , PPAR gamma/metabolismo , Alveolos Pulmonares/patología , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND: Perinatal inflammation is a key factor of brain vulnerability in neonates born preterm or with intra-uterine growth restriction (IUGR), two leading conditions associated with brain injury and responsible for neurocognitive and behavioral disorders. Systemic inflammation is recognized to activate microglia, known to be the critical modulators of brain vulnerability. Although some evidence supports a role for metabotropic glutamate receptor 3 (mGlu3 receptor) in modulation of neuroinflammation, its functions are still unknown in the developing microglia. METHODS: We used a double-hit rat model of perinatal brain injury induced by a gestational low-protein diet combined with interleukin-1ß injections (LPD/IL-1ß), mimicking both IUGR and prematurity-related inflammation. The effect of LPD/IL-1ß on mGlu3 receptor expression and the effect of mGlu3 receptor modulation on microglial reactivity were investigated using a combination of pharmacological, histological, and molecular and genetic approaches. RESULTS: Exposure to LPD/IL-1ß significantly downregulated Grm3 gene expression in the developing microglia. Both transcriptomic analyses and pharmacological modulation of mGlu3 receptor demonstrated its central role in the control of inflammation in resting and activated microglia. Microglia reactivity to inflammatory challenge induced by LPD/IL-1ß exposure was reduced by an mGlu3 receptor agonist. Conversely, both specific pharmacological blockade, siRNA knock-down, and genetic knock-out of mGlu3 receptors mimicked the pro-inflammatory phenotype observed in microglial cells exposed to LPD/IL-1ß. CONCLUSIONS: Overall, these data show that Grm3 plays a central role in the regulation of microglial reactivity in the immature brain. Selective pharmacological activation of mGlu3 receptors may prevent inflammatory-induced perinatal brain injury.
Asunto(s)
Microglía/metabolismo , Receptores de Glutamato Metabotrópico/metabolismo , Animales , Animales Recién Nacidos , Lesiones Encefálicas/metabolismo , Lesiones Encefálicas/patología , Femenino , Perfilación de la Expresión Génica/métodos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/patología , Embarazo , Ratas , Ratas Sprague-Dawley , Receptores de Glutamato Metabotrópico/antagonistas & inhibidoresRESUMEN
The poly(ADP-ribose) polymerase (PARP) inhibitor PJ34 has been reported to improve endothelial dysfunction in the peripheral system. We addressed the role of PJ34 on the vascular tone and vasoreactivity during development in the mouse brain. Blood flows were measured in the basilar trunk using ultrasonography. Cerebral vasoreactivity or vasodilation reserve was estimated as a percentage increase in mean blood flow velocities (mBFV) recorded under normoxia-hypercapnia in control and after PJ34 administration. Non-selective and selective eNOS and nNOS inhibitors were used to evaluate the role of NO-pathway into the hemodynamic effects of PJ34. PJ34 increased mBFVs from 15.8 ± 1.6 to 19.1 ± 1.9 cm/s (p = 0.0043) in neonatal, from 14.6 ± 1.4 to 16.1 ± 0.9 cm/s (p = 0.0049) in adult, and from 15.7 ± 1.7 to 17.5 ± 2.0 cm/s (p = 0.0024) in aged mice 48 h after administration. These PJ34 values were similar to those measured in age-matched control mice under normoxia-hypercapnia. This recruitment was mediated through the activation of constitutive NO synthases in both the neonatal (38.2 ± 6.7 nmol/min/mg protein) and adult (31.5 ± 4.4 nmol/min/mg protein) brain, as compared to age-matched control brain (6.9 ± 0.4 and 6.3 ± 0.7 nmol/min/mg protein), respectively. In addition, quite selective eNOS inhibitor was able to inhibit the recruitment. PJ34 by itself is able to increase cerebral blood flow through the NO-pathway activation at least over 48 h after a single administration.
Asunto(s)
Óxido Nítrico/metabolismo , Fenantrenos/metabolismo , Fenantrenos/farmacología , Factores de Edad , Animales , Animales Recién Nacidos/metabolismo , Encéfalo/embriología , Encéfalo/metabolismo , Circulación Cerebrovascular/efectos de los fármacos , Circulación Cerebrovascular/fisiología , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Óxido Nítrico Sintasa/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Vasodilatadores/metabolismo , Vasodilatadores/farmacologíaRESUMEN
Prematurity and fetal growth restriction (FGR) are frequent conditions associated with adverse neurocognitive outcomes. We have previously identified early deregulation of genes controlling neuroinflammation as a putative mechanism linking FGR and abnormal trajectory of the developing brain. While the oxytocin system was also found to be impaired following adverse perinatal events, its role in the modulation of neuroinflammation in the developing brain is still unknown. We used a double-hit rat model of perinatal brain injury induced by gestational low protein diet (LPD) and potentiated by postnatal injections of subliminal doses of interleukin-1ß (IL1ß) and a zebrafish model of neuroinflammation. Effects of the treatment with carbetocin, a selective, long lasting, and brain diffusible oxytocin receptor agonist, have been assessed using a combination of histological, molecular, and functional tools in vivo and in vitro. In the double-hit model, white matter inflammation, deficient myelination, and behavioral deficits have been observed and the oxytocin system was impaired. Early postnatal supplementation with carbetocin alleviated microglial activation at both transcriptional and cellular levels and provided long-term neuroprotection. The central anti-inflammatory effects of carbetocin have been shown in vivo in rat pups and in a zebrafish model of early-life neuroinflammation and reproduced in vitro on stimulated sorted primary microglial cell cultures from rats subjected to LPD. Carbetocin treatment was associated with beneficial effects on myelination, long-term intrinsic brain connectivity and behavior. Targeting oxytocin signaling in the developing brain may be an effective approach to prevent neuroinflammation - induced brain damage of perinatal origin.
Asunto(s)
Lesiones Encefálicas/tratamiento farmacológico , Encéfalo/patología , Microglía/efectos de los fármacos , Receptores de Oxitocina/metabolismo , Animales , Animales Modificados Genéticamente , Animales Recién Nacidos , Lesiones Encefálicas/inducido químicamente , Lesiones Encefálicas/patología , Células Cultivadas , Biología Computacional , Dieta con Restricción de Proteínas/efectos adversos , Modelos Animales de Enfermedad , Femenino , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Interleucina-1beta , Lipopolisacáridos/toxicidad , Oxitócicos/uso terapéutico , Oxitocina/análogos & derivados , Oxitocina/uso terapéutico , Fragmentos de Péptidos , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/fisiopatología , ARN Mensajero/metabolismo , Pez CebraRESUMEN
Neonatal acute ischemic stroke is a cause of neonatal brain injury that occurs more frequently in males, resulting in associated neurobehavioral disorders. The bases for these sex differences are poorly understood but might include the number, morphology and activation of microglia in the developing brain when subjected to stroke. Interestingly, poly (ADP-ribose) polymerase (PARP) inhibition preferentially protects males against neonatal ischemia. This study aims to examine the effects of PJ34, a PARP inhibitor, on microglial phenotypes at 3 and 8â¯days and on neurobehavioral disorders in adulthood for both male and female P9 mice subjected to permanent middle cerebral artery occlusion (pMCAo). PJ34 significantly reduced the lesion size by 78% and reduced the density of CX3CR1gfp-labeled microglial cells by 46% when examined 3â¯days after pMCAo in male but not in female mice. Eight days after pMCAo, the number of Iba1+/Cox-2+ cells did not differ between male and female mice in the cortical peri-infarct region. In the amygdala, Iba1+/Cox-2+ (M1-like) cell numbers were significantly decreased in PJ34-treated males but not in females. Conversely, Iba1+/Arg-1+ (M2-like) and Arg-1+/Cox-2+ (Mtransitional) cell numbers were significantly increased in PJ34-treated females. Regarding neurobehavioral disorders during adulthood, pMCAo induced a motor coordination deficit and a spatial learning deficit in female mice only. PJ34 prevented MBP fibers, motor coordination and learning disorders during adulthood in female mice. Our data show significant sex differences in the effects of PARP inhibition on microglia phenotypes following neonatal ischemia, associated with improved behavior and myelination during adulthood in females only. Our findings suggest that modulating microglial phenotypes may play key roles in behavior disorders and white matter injury following neonatal stroke.
Asunto(s)
Isquemia Encefálica/patología , Microglía/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/metabolismo , Animales , Animales Recién Nacidos , Lesiones Encefálicas/complicaciones , Isquemia Encefálica/metabolismo , Modelos Animales de Enfermedad , Femenino , Infarto de la Arteria Cerebral Media/fisiopatología , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Neuronas/efectos de los fármacos , Fenantrenos/metabolismo , Fenantrenos/farmacología , Fenotipo , Poli(ADP-Ribosa) Polimerasa-1/antagonistas & inhibidores , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Factores Sexuales , Accidente Cerebrovascular/patologíaRESUMEN
While arterial reflow after a stroke represents an important challenge for better outcomes, it is also very important that sudden recanalization does not produce local oxidative and nitrogen species, deleterious for the brain and more particularly the immature brain. Our objective was to determine whether a supply in prostaglandin (Pg) E1 (Alprostadil), via its action on arterial pressure, might progressively improve cerebral reperfusion in a neonatal stroke model. Arterial blood flow was measured using ultrasonography. Rate-limiting and Pg terminal synthesizing enzymes were evaluated using reverse-transcriptase polymerase chain reaction. Our data suggests that a supply in PgE1 might delay and improve the ipsilateral reperfusion by decreasing thromboxane A synthase-1 gene, the density of reactive astrocytes and lesion volume.
Asunto(s)
Alprostadil/uso terapéutico , Circulación Colateral/efectos de los fármacos , Accidente Cerebrovascular/tratamiento farmacológico , Accidente Cerebrovascular/fisiopatología , Alprostadil/farmacología , Animales , Animales Recién Nacidos , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/enzimología , Isquemia Encefálica/genética , Isquemia Encefálica/fisiopatología , Circulación Cerebrovascular/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Masculino , Ratas Wistar , Accidente Cerebrovascular/complicaciones , Accidente Cerebrovascular/enzimología , Tromboxano-A Sintasa/genética , Tromboxano-A Sintasa/metabolismoRESUMEN
The cognitive and behavioural deficits caused by traumatic brain injury (TBI) to the immature brain are more severe and persistent than TBI in the mature brain. Understanding this developmental sensitivity is critical as children under four years of age sustain TBI more frequently than any other age group. Microglia (MG), resident immune cells of the brain that mediate neuroinflammation, are activated following TBI in the immature brain. However, the type and temporal profile of this activation and the consequences of altering it are still largely unknown. In a mouse model of closed head weight drop paediatric brain trauma, we characterized i) the temporal course of total cortical neuroinflammation and the phenotype of ex vivo isolated CD11B-positive microglia/macrophage (MG/MΦ) using a battery of 32 markers, and ii) neuropathological outcome 1 and 5days post-injury. We also assessed the effects of targeting MG/MΦ activation directly, using minocycline a prototypical microglial activation antagonist, on these processes and outcome. TBI induced a moderate increase in both pro- and anti-inflammatory cytokines/chemokines in the ipsilateral hemisphere. Isolated cortical MG/MΦ expressed increased levels of markers of endogenous reparatory/regenerative and immunomodulatory phenotypes compared with shams. Blocking MG/MΦ activation with minocycline at the time of injury and 1 and 2days post-injury had only transient protective effects, reducing ventricular dilatation and cell death 1day post-injury but having no effect on injury severity at 5days. This study demonstrates that, unlike in adults, the role of MG/MΦ in injury mechanisms following TBI in the immature brain may not be negative. An improved understanding of MG/MΦ function in paediatric TBI could support translational efforts to design therapeutic interventions.
Asunto(s)
Lesiones Traumáticas del Encéfalo/metabolismo , Activación de Macrófagos/fisiología , Microglía/metabolismo , Animales , Encéfalo/metabolismo , Lesiones Encefálicas/inmunología , Lesiones Encefálicas/metabolismo , Lesiones Traumáticas del Encéfalo/inmunología , Quimiocinas/inmunología , Quimiocinas/metabolismo , Citocinas/inmunología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Activación de Macrófagos/efectos de los fármacos , Activación de Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Minociclina/farmacologíaRESUMEN
Fetal growth restriction (FGR) is a major complication of human pregnancy, frequently resulting from placental vascular diseases and prenatal malnutrition, and is associated with adverse neurocognitive outcomes throughout life. However, the mechanisms linking poor fetal growth and neurocognitive impairment are unclear. Here, we aimed to correlate changes in gene expression induced by FGR in rats and abnormal cerebral white matter maturation, brain microstructure, and cortical connectivity in vivo. We investigated a model of FGR induced by low-protein-diet malnutrition between embryonic day 0 and birth using an interdisciplinary approach combining advanced brain imaging, in vivo connectivity, microarray analysis of sorted oligodendroglial and microglial cells and histology. We show that myelination and brain function are both significantly altered in our model of FGR. These alterations, detected first in the white matter on magnetic resonance imaging significantly reduced cortical connectivity as assessed by ultrafast ultrasound imaging. Fetal growth retardation was found associated with white matter dysmaturation as shown by the immunohistochemical profiles and microarrays analyses. Strikingly, transcriptomic and gene network analyses reveal not only a myelination deficit in growth-restricted pups, but also the extensive deregulation of genes controlling neuroinflammation and the cell cycle in both oligodendrocytes and microglia. Our findings shed new light on the cellular and gene regulatory mechanisms mediating brain structural and functional defects in malnutrition-induced FGR, and suggest, for the first time, a neuroinflammatory basis for the poor neurocognitive outcome observed in growth-restricted human infants. GLIA 2016;64:2306-2320.
Asunto(s)
Lesiones Encefálicas/etiología , Lesiones Encefálicas/patología , Retardo del Crecimiento Fetal/fisiopatología , Microglía/metabolismo , Oligodendroglía/metabolismo , Transcriptoma/fisiología , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Animales , Animales Recién Nacidos , Antígenos/metabolismo , Antígenos CD/metabolismo , Encéfalo/diagnóstico por imagen , Encéfalo/efectos de los fármacos , Lesiones Encefálicas/diagnóstico por imagen , Citocinas/metabolismo , Femenino , Expresión Génica/fisiología , Lipopolisacáridos/farmacología , Proteína Básica de Mielina/metabolismo , Vías Nerviosas/diagnóstico por imagen , Vías Nerviosas/efectos de los fármacos , Factor de Transcripción 2 de los Oligodendrocitos/metabolismo , Embarazo , Proteoglicanos/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND AND PURPOSE: We previously showed that the selective neuronal nitric oxide synthase inhibitor 7-nitroindazole (7-NI) increases cerebral microcirculation in a juvenile ischemic rat model. We address the roles of cyclooxygenase (COX)-elaborated prostaglandins in collateral recruitment and blood supply. METHODS: Fourteen-day-old rats were subjected to ischemia-reperfusion and treated with either PBS or 7-NI (25 mg/kg) at the reperfusion onset. Six-keto-prostaglandin F1α was measured using ELISA. COX-1 and COX-2 and prostaglandin terminal synthesizing enzymes were evaluated using reverse-transcriptase polymerase chain reaction and immunofluorescence. Microvascular blood flow indexes (artery diameter and capillaries number) were measured using sidestream dark-field videomicroscopy in PBS- and 7-NI-treated ischemic rats in the absence or presence of the COX-2 inhibitor NS-398 (5 mg/kg). Cell death was measured with the TUNEL (terminal transferase dUTP nick end labeling) assay and cleaved-caspase-3 immunostaining. RESULTS: Six-keto-prostaglandin F1α and COX-2, associated with a prostaglandin E synthase, were significantly increased in PBS- and 7-NI-treated animals 15 minutes and 1 hour after ischemia-reperfusion, respectively. In contrast and as compared with PBS, 7-NI significantly decreased prostacyclin synthase and cytosolic prostaglandins E synthase mRNA. Selective COX-2 inhibition significantly decreased blood flow indexes and significantly reversed the effects of 7-NI, including the number of TUNEL+- and cleaved-caspase-3+-nuclei. CONCLUSIONS: These results show that the juvenile rat brains mostly respond to ischemia by a COX-2-dependent prostaglandins production and suggest that the transcriptional responses observed under 7-NI facilitate and reorient COX-2-dependent prostaglandins production.
Asunto(s)
6-Cetoprostaglandina F1 alfa/metabolismo , Circulación Cerebrovascular , Inhibidores de la Ciclooxigenasa 2/farmacología , Ciclooxigenasa 2/metabolismo , Microcirculación , Prostaglandina-E Sintasas/metabolismo , Daño por Reperfusión/metabolismo , Animales , Modelos Animales de Enfermedad , RatasRESUMEN
BACKGROUND: Perinatal ischemic stroke is the most frequent form of cerebral infarction in neonates; however, evidence-based treatments are currently lacking. We have previously demonstrated a beneficial effect of sildenafil citrate, a PDE-5 inhibitor, on stroke lesion size in neonatal rat pups. The present study investigated the effects of sildenafil in a neonatal mouse stroke model on (1) hemodynamic changes and (2) regulation of astrocyte/microglia-mediated neuroinflammation. METHODS: Ischemia was induced in C57Bl/6 mice on postnatal (P) day 9 by permanent middle cerebral artery occlusion (pMCAo), and followed by either PBS or sildenafil intraperitoneal (i.p.) injections. Blood flow (BF) velocities were measured by ultrasound imaging with sequential Doppler recordings and laser speckle contrast imaging. Animals were euthanized, and brain tissues were obtained at 72 h or 8 days after pMCAo. Expression of M1- and M2-like microglia/macrophage markers were analyzed. RESULTS: Although sildenafil (10 mg/kg) treatment potently increased cGMP concentrations, it did not influence early collateral recruitment nor did it reduce mean infarct volumes 72 h after pMCAo. Nevertheless, it provided a significant dose-dependent reduction of mean lesion extent 8 days after pMCAo. Suggesting a mechanism involving modulation of the inflammatory response, sildenafil significantly decreased microglial density at 72 h and 8 days after pMCAo. Gene expression profiles indicated that sildenafil treatment also modulates M1- (ptgs2, CD32 and CD86) and M2-like (CD206, Arg-1 and Lgals3) microglia/macrophages in the late phase after pMCAo. Accordingly, the number of COX-2(+) microglia/macrophages significantly increased in the penumbra at 72 h after pMCAo but was significantly decreased 8 days after ischemia in sildenafil-treated animals. CONCLUSIONS: Our findings argue that anti-inflammatory effects of sildenafil may provide protection against lesion extension in the late phase after pMCAo in neonatal mice. We propose that sildenafil treatment could represent a potential strategy for neonatal ischemic stroke treatment/recovery.
Asunto(s)
Isquemia Encefálica/patología , Microglía/efectos de los fármacos , Inhibidores de Fosfodiesterasa/farmacología , Citrato de Sildenafil/farmacología , Animales , Animales Recién Nacidos , Isquemia Encefálica/enzimología , Modelos Animales de Enfermedad , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Microglía/enzimología , Reacción en Cadena de la PolimerasaRESUMEN
The cognitive and behavioral deficits caused by traumatic brain injury (TBI) to the immature brain are more severe and persistent than injuries to the adult brain. Understanding this developmental sensitivity is critical because children under 4 years of age of sustain TBI more frequently than any other age group. One of the first events after TBI is the infiltration and degranulation of mast cells (MCs) in the brain, releasing a range of immunomodulatory substances; inhibition of these cells is neuroprotective in other types of neonatal brain injury. This study investigates for the first time the role of MCs in mediating injury in a P7 mouse model of pediatric contusion-induced TBI. We show that various neural cell types express histamine receptors and that histamine exacerbates excitotoxic cell death in primary cultured neurons. Cromoglycate, an inhibitor of MC degranulation, altered the inflammatory phenotype of microglia activated by TBI, reversing several changes but accentuating others, when administered before TBI. However, without regard to the time of cromoglycate administration, inhibiting MC degranulation did not affect cell loss, as evaluated by ventricular dilatation or cleaved caspase-3 labeling, or the density of activated microglia, neurons, or myelin. In double-heterozygous cKit mutant mice lacking MCs, this overall lack of effect was confirmed. These results suggest that the role of MCs in this model of pediatric TBI is restricted to subtle effects and that they are unlikely to be viable neurotherapeutic targets. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Lesiones Traumáticas del Encéfalo/patología , Mastocitos/patología , Animales , Contusión Encefálica/patología , Caspasa 3/biosíntesis , Caspasa 3/genética , Muerte Celular/efectos de los fármacos , Células Cultivadas , Preescolar , Cromolin Sódico/farmacología , Modelos Animales de Enfermedad , Histamina/farmacología , Humanos , Lactante , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Microglía/metabolismo , Células-Madre Neurales , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Proteínas Proto-Oncogénicas c-kit/genética , Receptores Histamínicos/metabolismoRESUMEN
Several lines of evidence demonstrate that inhaled nitric oxide (iNO) not only acts locally on the pulmonary vasculature but also has remote effects on the mature and developing brain under basal or pathological conditions by modulating cerebral blood flow and microvascularization, white matter maturation, inflammation, and subsequent brain repair. Previously, consistent studies demonstrated that increased levels of guanosine 3',5' cyclic monophosphate (cGMP), the main effector of biological effect induced by nitric oxide (NO), significantly augment proliferation and neuronal differentiation of adult neural progenitor cells (NPCs). In the present study, we ask the question whether iNO could promote the proliferation of NPCs in the uninjured developing brain. We first reported that iNO exposure at a concentration of 20 ppm during the first 7 days of life was associated with a significant but transient elevation of brain cGMP concentration 2 h after the onset of iNO exposure and a subsequent increase in myelin content of the developing white matter at postnatal day (P) 10. Using BrDu labelling and colabelling with specific cell-type markers we found that iNO exposure of rat pups results in an increased NPC proliferation in several layers of the subventricular zone (SVZ) at both early (30 h) and late (P7) time points. These proliferating NPCs were found to be sustainably viable and subsequently differentiated into oligodendroglial cells in the developing white matter and cortex. We also found that NG2 immunoreactivity around vessel walls, labeling pericyte cells, was increased in NO-exposed rat pups in the periventricular SVZ. In conclusion, iNO appears to act on oligodendrocyte progenitor cells, leading to increased density of mature oligodendrocytes and myelin content in the immature rat brain.
Asunto(s)
Encéfalo/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células-Madre Neurales/efectos de los fármacos , Neurotransmisores/farmacología , Óxido Nítrico/farmacología , Oligodendroglía/efectos de los fármacos , Animales , Animales Recién Nacidos , Encéfalo/crecimiento & desarrollo , Modelos Animales de Enfermedad , Femenino , Masculino , Neurotransmisores/administración & dosificación , Óxido Nítrico/administración & dosificación , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND: Fetal growth restriction is the second leading cause of perinatal morbidity and mortality, and neonates with intrauterine growth retardation (IUGR) have increased neurocognitive and neuropsychiatric morbidity. These neurocognitive impairments are mainly related to injury of the developing brain associated with IUGR. Growing evidence from preclinical models of brain injury in both adult and neonatal rodents supports the view that nitric oxide can promote neuroprotection. METHODS: In a model of IUGR induced by protracted gestational hypoxia leading to diffuse white matter injury, we subjected neonatal rats to low dose (5 ppm) but long-lasting (7 d) exposure to inhaled NO (iNO). We used a combination of techniques, including immunohistochemistry, quantitative PCR, and cognitive assessment, to assess neuroprotection. RESULTS: Antenatal hypoxia-induced IUGR was associated with severe neuroinflammation and delayed myelination. iNO exposure during the first postnatal week significantly attenuated cell death and microglial activation, enhanced oligodendroglial proliferation and finally improved myelination. Remarkably, iNO was associated with the specific upregulation of P27kip1, which initiates oligodendrocytic differentiation. Finally, iNO counteracted the deleterious effects of hypoxia on learning abilities. CONCLUSION: This study provides new evidence that iNO could be effective in preventing brain damage and/or enhancing repair of the developing brain.
Asunto(s)
Administración por Inhalación , Fármacos Neuroprotectores/química , Óxido Nítrico/administración & dosificación , Sustancia Blanca/efectos de los fármacos , Animales , Animales Recién Nacidos , Axones/patología , Conducta Animal , Lesiones Encefálicas/patología , Lesiones Encefálicas/prevención & control , Trastornos del Conocimiento/fisiopatología , Modelos Animales de Enfermedad , Femenino , Retardo del Crecimiento Fetal , Hipoxia , Inmunohistoquímica , Aprendizaje , Vaina de Mielina/química , Oligodendroglía/metabolismo , Embarazo , Ratas , Ratas Sprague-Dawley , Sustancia Blanca/patologíaRESUMEN
BACKGROUND AND PURPOSE: The best conceivable treatment for hypoxia-ischemia (HI) is the restoration of blood flow to the hypoxic-ischemic region(s). Our objective was to examine whether boosting NO-cGMP signaling using sildenafil citrate, a phosphodiesterase-type 5 inhibitor, could modify cerebral blood flow and reduce lesions in the developing brain. METHODS: HI was induced in P7 Sprague-Dawley rats by unilateral carotid artery occlusion and hypoxia, and followed by either PBS or sildenafil. Blood-flow velocities were measured by ultrasound imaging with sequential Doppler recordings to evaluate collateral recruitment. Cell death, blood-brain barrier integrity, and glial activation were analyzed by immunohistochemistry. Motor behavior was evaluated using an open-field device adapted to neonatal animals. RESULTS: Sildenafil citrate (10 mg/kg) induced collateral patency, reduced terminal dUTP nick-end labeling-positive cells, reactive astrogliosis, and macrophage/microglial activation at 72 hours and 7 days post-HI. Sildenafil also reduced the number of terminal dUTP nick-end labeling-positive endothelial cells within lesion site. Seven days after HI and sildenafil treatment, tissue loss was significantly reduced, and animals recovered motor coordination. CONCLUSIONS: Our findings strongly indicate that sildenafil citrate treatment, associated with a significant increase in cerebral blood flow, reduces HI damage and improves motor locomotion in neonatal rats. Sildenafil may represent an interesting therapeutic strategy for neonatal neuroprotection.
Asunto(s)
Animales Recién Nacidos/fisiología , Circulación Cerebrovascular/efectos de los fármacos , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Fármacos Neuroprotectores , Inhibidores de Fosfodiesterasa/farmacología , Piperazinas/farmacología , Sulfonas/farmacología , Animales , Análisis de los Gases de la Sangre , Presión Sanguínea/efectos de los fármacos , Presión Sanguínea/fisiología , Muerte Celular/efectos de los fármacos , GMP Cíclico/fisiología , Lateralidad Funcional/fisiología , Hipoxia-Isquemia Encefálica/fisiopatología , Hipoxia-Isquemia Encefálica/psicología , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Inflamación/patología , Activación de Macrófagos/efectos de los fármacos , Microcirculación/efectos de los fármacos , Actividad Motora/efectos de los fármacos , Neuroglía/efectos de los fármacos , Óxido Nítrico/fisiología , Desempeño Psicomotor/efectos de los fármacos , Purinas/farmacología , Ratas , Citrato de SildenafilRESUMEN
OBJECTIVE: Perinatal infections and the systemic inflammatory response to them are critical contributors to white matter disease (WMD) in the developing brain despite the use of highly active antibiotics. Fluoroquinolones including ciprofloxacin (CIP) have intrinsic anti-inflammatory effects. We hypothesized that CIP, in addition to its antibacterial activity, could exert a neuroprotective effect by modulating white matter inflammation in response to sepsis. METHODS: We adapted an Escherichia coli sepsis model to 5-day-old rat pups (P5), to induce white matter inflammation without bacterial meningitis. We then compared the ability of CIP to modulate inflammatory-induced brain damage compared with cefotaxime (CTX) (treatment of reference). RESULTS: Compared with CTX, CIP was associated with reduced microglial activation and inducible nitric oxide synthase (iNOS) expression in the developing white matter in rat pups subjected to E. coli sepsis. In addition to reducing microglial activation, CIP was able to prevent myelination delay induced by E. coli sepsis and to promote oligodendroglial survival and maturation. We found that E. coli sepsis altered the transcription of the guidance molecules semaphorin 3A and 3F; CIP treatment was capable of reducing semaphorin 3A and 3F transcription levels to those seen in uninfected controls. Finally, in a noninfectious white matter inflammation model, CIP was associated with significantly reduced microglial activation and prevented WMD when compared to CTX. INTERPRETATION: These data strongly suggest that CIP exerts a beneficial effect in a model of E. coli sepsis-induced WMD in rat pups that is independent of its antibacterial activity but likely related to iNOS expression modulation.
Asunto(s)
Antiinfecciosos/uso terapéutico , Ciprofloxacina/uso terapéutico , Enfermedades Desmielinizantes/prevención & control , Infecciones por Escherichia coli/complicaciones , Fibras Nerviosas Mielínicas/efectos de los fármacos , Sepsis/complicaciones , Animales , Animales Recién Nacidos , Enfermedades Desmielinizantes/etiología , Enfermedades Desmielinizantes/metabolismo , Enfermedades Desmielinizantes/patología , Infecciones por Escherichia coli/metabolismo , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Fibras Nerviosas Mielínicas/metabolismo , Fibras Nerviosas Mielínicas/patología , Proteínas del Tejido Nervioso/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Semaforina-3A/metabolismo , Sepsis/metabolismo , Estadísticas no ParamétricasRESUMEN
Relative to the cefotaxime-gentamicin combination, the moxifloxacin-cefotaxime combination significantly reduced microglial activation and immature oligodendrocyte cell death and delayed myelination in the developing white matter of neonatal rats with experimental Escherichia coli sepsis. These neuroprotective effects were not due to differences in in vivo bactericidal activities or in the systemic inflammatory responses and could be related to the intrinsic immunomodulatory properties of moxifloxacin. Molecular mechanisms underlying the neuroprotective effect of moxifloxacin remain to be elucidated.
Asunto(s)
Antibacterianos/uso terapéutico , Compuestos Aza/uso terapéutico , Cefotaxima/uso terapéutico , Infecciones por Escherichia coli/tratamiento farmacológico , Gentamicinas/uso terapéutico , Leucoencefalopatías/microbiología , Leucoencefalopatías/prevención & control , Quinolinas/uso terapéutico , Sepsis/tratamiento farmacológico , Animales , Animales Recién Nacidos , Fluoroquinolonas , Moxifloxacino , Ratas , Ratas Sprague-DawleyRESUMEN
White matter damage (WMD) remains the leading cause of cerebral palsy in children born prematurely. The release of an excessive amount of reactive oxygen species is recognized as a risk factor for WMD. We hypothesize that free radical injury during reoxygenation at birth may be harmful to the immature white matter and may underlie, at least in part, the pathogenesis of WMD. We tested this hypothesis in rat pups delivered from normoxic pregnant rats, and by investigating an animal model based on protracted antenatal hypoxia in the pregnant rat and mimicking the main features of human WMD in rat pups. From embryonic day (E)5 to E21, the pregnant rats were placed in a chamber supplied with a gas mixture that either induced hypoxia (FiO(2) = 10%) or maintained normoxia (FiO(2) = 21%). On E21, the dams were removed from the chamber and housed under either normoxia (FiO(2) = 21%), hyperoxia (FiO(2) = 60%) or slowly reoxygenated (FiO(2) from 15% at E21 to 21% at postnatal day 7). Postnatal hyperoxia was associated with a significantly increased density of activated microglial cells (+105%) and TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling)-positive cells (+85%) within the developing white matter. Myelin content (-31%) and mature oligodendrocyte density (-37%) in the normal developing white matter were significantly decreased by postnatal hyperoxia. Postnatal hyperoxia significantly potentiated the myelination delay and oligodendroglial dysmaturation induced by antenatal hypoxia. In contrast, progressive reoxygenation at birth did not induce any change in white matter inflammation, myelination and cell death as compared with normoxic controls, and prevented most of the WMD observed following antenatal hypoxia. This study demonstrates a deleterious effect of hyperoxia at birth on the developing white matter in normal rat pups. Postnatal hyperoxia worsened the WMD induced by antenatal hypoxia. Hyperoxia at birth should be avoided in preterm infants at risk of WMD.
Asunto(s)
Animales Recién Nacidos , Lesiones Encefálicas/etiología , Lesiones Encefálicas/patología , Hiperoxia/complicaciones , Fibras Nerviosas Mielínicas/patología , Oxígeno/efectos adversos , Animales , Parálisis Cerebral/etiología , Parálisis Cerebral/patología , Niño , Modelos Animales de Enfermedad , Femenino , Humanos , Recién Nacido , Recien Nacido Prematuro , Vaina de Mielina/metabolismo , Vaina de Mielina/patología , Factores de Crecimiento Nervioso/genética , Factores de Crecimiento Nervioso/metabolismo , Oligodendroglía/patología , Oligodendroglía/fisiología , Embarazo , Ratas , Ratas Sprague-DawleyRESUMEN
Cerebral stroke, traumatic brain injury, and hypoxic ischemic encephalopathy are among the most frequently occurring brain injuries. A complex pathogenesis, characterized by a synergistic interaction between alterations of the cerebrovascular system, cell death, and inflammation, is at the basis of the brain damage that leads to behavioral and neurodevelopmental disabilities in affected subjects. Sildenafil is a selective inhibitor of the enzyme phosphodiesterase 5 (PDE5) that is able to cross the blood-brain barrier. Preclinical data suggest that sildenafil may be a good candidate for the prevention or repair of brain injury in both adults and neonates. The aim of this review is to summarize the evidence supporting the neuroprotective action of sildenafil and discuss the possible benefits of the association of sildenafil with current therapeutic strategies.
Asunto(s)
Lesiones Encefálicas/tratamiento farmacológico , Neuroprotección , Citrato de Sildenafil/uso terapéutico , Investigación Biomédica Traslacional , Adulto , Humanos , Recién Nacido , Neuroprotección/efectos de los fármacos , Óxido Nítrico/metabolismo , Caracteres Sexuales , Citrato de Sildenafil/farmacologíaRESUMEN
Backgroud: Type-3 metabotropic glutamate (mGlu3) receptors are found in both neurons and glial cells and regulate synaptic transmission, astrocyte function, and microglial reactivity. Here we show that the genetic deletion of mGlu3 receptors amplifies ischemic brain damage and associated neuroinflammation in adult mice. An increased infarct size was observed in mGlu3-/- mice of both CD1 and C57Black strains 24 h following a permanent occlusion of the middle cerebral artery (MCA) as compared to their respective wild-type (mGlu3+/+ mice) counterparts. Increases in the expression of selected pro-inflammatory genes including those encoding interleukin-1ß, type-2 cycloxygenase, tumor necrosis factor-α, CD86, and interleukin-6 were more prominent in the peri-infarct region of mGlu3-/- mice. In contrast, the expression of two genes associated with the anti-inflammatory phenotype of microglia (those encoding the mannose-1-phosphate receptor and the α-subunit of interleukin-4 receptor) and the gene encoding the neuroprotective factor, glial cell line-derived neurotrophic factor, was enhanced in the peri-infarct region of wild-type mice, but not mGlu3-/- mice, following MCA occlusion. In C57Black mice, the genetic deletion of mGlu3 receptors worsened the defect in the paw placement test as assessed in the contralateral forepaw at short times (4 h) following MCA occlusion. These findings suggest that mGlu3 receptors are protective against ischemic brain damage and support the way to the use of selective mGlu3 receptor agonists or positive allosteric modulators in experimental animal models of ischemic stroke.