RESUMEN
Traumatic brain injury leads to a highly orchestrated immune- and glial cell response partially responsible for long-lasting disability and the development of secondary neurodegenerative diseases. A holistic understanding of the mechanisms controlling the responses of specific cell types and their crosstalk is required to develop an efficient strategy for better regeneration. Here, we combine spatial and single-cell transcriptomics to chart the transcriptomic signature of the injured male murine cerebral cortex, and identify specific states of different glial cells contributing to this signature. Interestingly, distinct glial cells share a large fraction of injury-regulated genes, including inflammatory programs downstream of the innate immune-associated pathways Cxcr3 and Tlr1/2. Systemic manipulation of these pathways decreases the reactivity state of glial cells associated with poor regeneration. The functional relevance of the discovered shared signature of glial cells highlights the importance of our resource enabling comprehensive analysis of early events after brain injury.
Asunto(s)
Lesiones Encefálicas , Heridas Punzantes , Animales , Ratones , Masculino , Proteína Ácida Fibrilar de la Glía/metabolismo , Neuroglía/metabolismo , Lesiones Encefálicas/metabolismo , Corteza Cerebral/metabolismo , Heridas Punzantes/complicaciones , Heridas Punzantes/metabolismoRESUMEN
Astrogliosis after brain trauma can have a significant impact on functional recovery. However, little is known about the mechanisms underlying astrocyte proliferation and subsequent astrogliosis. In this study, we established a cortical stab wound injury mouse model and observed dramatic astrocyte activation and nerve growth factor receptor (p75NTR) upregulation near the lesion. We also found profound alterations in the cell cycle of astrocytes near the lesion, with a switch from a mitotically quiescent (G0) phase to the G2/M and S phases. However, no changes in the level of astrocyte apoptosis were observed. Cell cycle progression to the G2/M and S phases and CDK2 protein levels in response to cortical stab wound was inhibited after p75NTR knockdown in mouse astrocytes. Conversely, p75NTR overexpression in mouse astrocytes was sufficient in promoting cell cycle progression. In conclusion, our results suggested that p75NTR upregulation in astrocytes after brain injury induces cell cycle entry by promoting CDK2 expression and promoting astrocyte proliferation. Our findings provided a better understanding of astrocytic responses after cortical stab wound injury in mice.
Asunto(s)
Astrocitos , Heridas Punzantes , Animales , Astrocitos/metabolismo , Proliferación Celular , Gliosis/patología , Ratones , Proteínas del Tejido Nervioso , Receptores de Factores de Crecimiento , Receptores de Factor de Crecimiento Nervioso/metabolismo , Heridas Punzantes/metabolismo , Heridas Punzantes/patologíaRESUMEN
We examined the mechanism how 2-carba-cyclic phosphatidic acid (2ccPA), a lipid mediator, regulates neuronal apoptosis in traumatic brain injury (TBI). First, we found 2ccPA suppressed neuronal apoptosis after the injury, and increased the immunoreactivity of tenascin-C (TN-C), an extracellular matrix protein by 2ccPA in the vicinity of the wound region. 2ccPA increased the mRNA expression levels of Tnc in primary cultured astrocytes, and the conditioned medium of 2ccPA-treated astrocytes suppressed the apoptosis of cortical neurons. The neuroprotective effect of TN-C was abolished by knockdown of TN-C. These results indicate that 2ccPA contributes to neuroprotection via TN-C from astrocytes in TBI.
Asunto(s)
Astrocitos/metabolismo , Lesiones Traumáticas del Encéfalo/metabolismo , Fármacos Neuroprotectores/uso terapéutico , Ácidos Fosfatidicos/fisiología , Tenascina/metabolismo , Animales , Apoptosis/efectos de los fármacos , Astrocitos/efectos de los fármacos , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Células Cultivadas , Corteza Cerebral/citología , Medios de Cultivo Condicionados/farmacología , Femenino , Proteína Ácida Fibrilar de la Glía/biosíntesis , Proteína Ácida Fibrilar de la Glía/genética , Inyecciones Intraperitoneales , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos ICR , Neuronas/efectos de los fármacos , Neuronas/patología , Ácidos Fosfatidicos/farmacología , Ácidos Fosfatidicos/uso terapéutico , Interferencia de ARN , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Tenascina/antagonistas & inhibidores , Tenascina/genética , Heridas Punzantes/tratamiento farmacológico , Heridas Punzantes/metabolismoRESUMEN
Recent findings suggest a significant role of the brain-derived neurotrophic factor (BDNF) as a mediator of brain regeneration following a stab injury in zebrafish. Since BDNF has been implicated in many physiological processes, we hypothesized that these processes are affected by brain injury in zebrafish. Hence, we examined the impact of stab injury on oxidative stress and apoptosis in the adult zebrafish brain. Stab wound injury (SWI) was induced in the right telencephalic hemisphere of the adult zebrafish brain and examined at different time points. The biochemical variables of oxidative stress insult and transcript levels of antioxidant genes were assessed to reflect upon the oxidative stress levels in the brain. Immunohistochemistry was performed to detect the levels of early apoptotic marker protein cleaved caspase-3, and the transcript levels of pro-apoptotic and anti-apoptotic genes were examined to determine the effect of SWI on apoptosis. The activity of antioxidant enzymes, the level of lipid peroxidation (LPO) and reduced glutathione (GSH) were significantly increased in the injured fish brain. SWI also enhanced the expression of cleaved caspase-3 protein and apoptosis-related gene transcripts. Our results indicate induction of oxidative stress and apoptosis in the telencephalon of adult zebrafish brain by SWI. These findings contribute to the overall understanding of the pathophysiology of traumatic brain injury and adult neurogenesis in the zebrafish model and raise new questions about the compensatory physiological mechanisms in response to traumatic brain injury in the adult zebrafish brain.
Asunto(s)
Lesiones Encefálicas/fisiopatología , Regeneración Cerebral/fisiología , Estrés Oxidativo/fisiología , Animales , Apoptosis/genética , Encéfalo/metabolismo , Lesiones Encefálicas/genética , Lesiones Encefálicas/metabolismo , Lesiones Traumáticas del Encéfalo , Regeneración Cerebral/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Modelos Animales de Enfermedad , Regeneración Nerviosa/fisiología , Neurogénesis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Telencéfalo/lesiones , Telencéfalo/metabolismo , Telencéfalo/fisiopatología , Heridas Punzantes/metabolismo , Heridas Punzantes/fisiopatología , Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
The present study proves that rapid and demarcating astroglial reactions are confined to birds and mammals. To understand the function of post-lesion astroglial reaction, the phylogenetical aspects are also to be investigated. Considering the regenerative capabilities, reptiles represent an intermediate position between the brain regeneration-permissive fishes and amphibians and the almost non-permissive birds and mammals. Damage is followed by a rapid astroglial reaction in the mammalian and avian brain, which is held as an impediment of regeneration. In other vertebrates the reactions were usually observed following long survival periods together with signs of regeneration, therefore they can be regarded as concomitant phenomena of regeneration. The present study applies short post-lesion periods comparable to those seen in mammals and birds for astroglial reactions. Two species of lizards were used: gecko (leopard gecko, Eublepharis macularius, Blyth, 1854) and agama (bearded dragon, Pogona vitticeps, Ahl, 1926). The gecko brain is rich in GFAP whereas the agama brain is quite poor in this. Crocodilia, the closest extant relatives of birds were represented in this study by Cuvier's dwarf caiman (Paleosuchus palpebrosus, Cuvier, 1807). The post-lesion astroglial reactions of crocodilians have never been investigated. The injuries were stab wounds in the telencephalon. The survival periods lasted 3, 7, 10 or 14 days. Immunoperoxidase reactions were performed applying anti-GFAP, anti-vimentin and anti-nestin reagents. No rapid and demarcating astroglial reaction resembling that of mammalian or avian brains was found. Alterations of the perivascular immunoreactivities of laminin and ß-dystroglycan as indicators of glio-vascular decoupling proved that the lesions were effective on astroglia. The capability of rapid and demarcating astroglial reaction seems to be confined to mammals and birds and to appear by separate, parallel evolution in them.
Asunto(s)
Astrocitos/patología , Lesiones Traumáticas del Encéfalo/patología , Encéfalo/patología , Heridas Punzantes/patología , Animales , Astrocitos/metabolismo , Evolución Biológica , Aves , Encéfalo/metabolismo , Lesiones Traumáticas del Encéfalo/metabolismo , Modelos Animales de Enfermedad , Femenino , Proteína Ácida Fibrilar de la Glía/metabolismo , Lagartos , Masculino , Mamíferos , Nestina/metabolismo , Especificidad de la Especie , Factores de Tiempo , Vimentina/metabolismo , Heridas Punzantes/metabolismoRESUMEN
The healing of acute wounds is vital to humans and is a well-orchestrated process that involves systemic and local factors. However, there is a lack of effective and safe clinical therapies. The collagen triple helix repeat containing 1 (CTHRC1) protein is a type of exocrine protein that has been recently reported to contribute to tissue repair. Our aim is to validate the promoting effects of CTHRC1 on the healing of acute wounds and to elucidate the underlying molecular mechanism. Therefore, we first established acute wound healing mouse models and confirmed that CTHRC1 accelerates the healing process of acute wounds. Then, we characterized wound macrophages using a polyvinylalcohol (PVA) sponge model and used Western blotting to investigate the molecular mechanism. We found that CTHRC1 increased the M2 macrophage population and the TGF-ß expression level as a result of the activation of the TGF-ß and Notch pathways, which eventually contributed to the promotion of wound healing. Inhibition of the Notch pathway showed attenuated M2 macrophage recruitment, and it decreased the TGF-ß expression level. These results substantiate our hypothesis that CTHRC1 promotes wound healing by recruiting M2 macrophages and regulating the TGF-ß and Notch pathways.
Asunto(s)
Proteínas de la Matriz Extracelular/farmacología , Macrófagos/efectos de los fármacos , Receptores Notch/metabolismo , Piel/lesiones , Factor de Crecimiento Transformador beta/metabolismo , Cicatrización de Heridas/efectos de los fármacos , Heridas Punzantes/tratamiento farmacológico , Animales , Línea Celular , Modelos Animales de Enfermedad , Humanos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Recombinantes/farmacología , Transducción de Señal , Piel/inmunología , Piel/metabolismo , Cicatrización de Heridas/inmunología , Heridas Punzantes/inmunología , Heridas Punzantes/metabolismoRESUMEN
The reparative ability of the central nervous system varies widely in the animal kingdom. In the mammalian brain, the regenerative mechanisms are very limited and newly formed neurons do not survive longer, probably due to a non-suitable local environment. On the opposite, fish can repair the brain after injury, with fast and complete recovery of damaged area. The brain of zebrafish, a teleost fish widely used as vertebrate model, also possesses high regenerative properties after injury. Taking advantage of this relevant model, the aim of the present study was to investigate the role of brain-derived neurotrophic factor (BDNF) in the regenerative ability of adult brain, after stab wound telencephalic injury. BDNF is involved in many brain functions and plays key roles in the repair process after traumatic brain lesions. It has been reported that BDNF strengthens the proliferative activity of neuronal precursor cells, facilitates the neuronal migration toward injured areas, and shows survival properties due to its anti-apoptotic effects. BDNF mRNA levels, assessed by quantitative PCR and in situ hybridization at 1, 4, 7, and 15 days after the lesion, were increased in the damaged telencephalon, mostly suddenly after the lesion. Double staining using in situ hybridization and immunocytochemistry revealed that BDNF mRNA was restricted to cells identified as mature neurons. BDNF mRNA expressing neurons mostly increased in the area around the lesion, showing a peak 1 day after the lesion. Taken together, these results highlight the role of BDNF in brain repair processes and reinforce the value of zebrafish for the study of regenerative neurogenesis.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/metabolismo , Telencéfalo/lesiones , Telencéfalo/metabolismo , Proteínas de Pez Cebra/metabolismo , Animales , Modelos Animales de Enfermedad , Lateralidad Funcional , Masculino , Regeneración Nerviosa/fisiología , Neuronas/metabolismo , Neuronas/patología , ARN Mensajero/metabolismo , Telencéfalo/patología , Heridas Punzantes/metabolismo , Heridas Punzantes/patología , Pez CebraRESUMEN
Previous research by the authors on an animal model showed that bloodstains can contain additional information about their somatic origin in the form of wound cells. Bloodstains produced by a gunshot wound to the head were distinguished from bloodstains produced by a gunshot wound to the chest by testing the stains for a brain microRNA marker. In this study, the effectiveness of the technique was examined on blood drops shed externally from a stab wound to the liver of rat carcasses. Specifically, investigations were conducted on the liver microRNA marker, rno-mir-122-3p, with the QIAGEN miScript System, and PCR analysis. Between the two stabbing methods used, 67% of the scalpel blades and 57% of the blood drops tested positive for rno-mir-122-3p; however, other samples tested negative giving inconclusive results as to the wound-of-origin. The amount of the liver cells in the bloodstains appeared to be related to the extent of trauma.
Asunto(s)
Manchas de Sangre , MicroARNs/genética , Heridas Punzantes/metabolismo , Traumatismos Abdominales/metabolismo , Animales , Patologia Forense , Marcadores Genéticos , Hígado/lesiones , Hígado/metabolismo , MicroARNs/metabolismo , Modelos Animales , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Traumatismos Torácicos/metabolismoRESUMEN
Repetitive transcranial magnetic stimulation (rTMS), a non-invasive form of brain stimulation, has shown experimental and clinical efficacy in a range of neuromodulatory models, even when delivered at low intensity (i.e. subthreshold for action potential generation). After central nervous system (CNS) injury, studies suggest that reactive astrocytes and microglia can have detrimental but also beneficial effects; thus modulating glial activity, for example through application of rTMS, could potentially be a useful therapeutic tool following neurotrauma. Immunohistochemistry was used to measure the effect of low intensity rTMS (LI-rTMS) on GFAP (astrocyte), IBA1 (microglial), and CS56 (proteoglycan) expression in a unilateral penetrating cortical stab injury model of glial scarring in young adult and aged male and female C57BL6/J mice. Mice received contralateral low frequency, ipsilateral low frequency, ipsilateral high frequency or sham LI-rTMS (4-5mT intensity), for two weeks following injury. There was no significant difference in the overall volume of tissue containing GFAP positive (+) astrocytes, IBA1+ microglia, or proteoglycan expression, between sham and LI-rTMS-treated mice of all ages and sex. Importantly however, the density of GFAP+ astrocytes and IBA1+ microglia immediately adjacent to the injury was significantly reduced following ipsilateral low and high frequency stimulation in adult and aged females (p≤0.05), but was significantly increased in adult and aged males (p≤0.05). LI-rTMS effects were generally of greater magnitude in aged mice compared to young adult mice. These results suggest that sex differences need to be factored into therapeutic rTMS protocols. In particular, more work analyzing frequency and intensity specific effects, especially in relation to age and sex, is required to determine how rTMS can best be used to modify glial reactivity and phenotype following neurotrauma.
Asunto(s)
Corteza Cerebral/lesiones , Corteza Cerebral/patología , Neuroglía/patología , Estimulación Magnética Transcraneal/métodos , Heridas Punzantes/patología , Heridas Punzantes/terapia , Envejecimiento , Animales , Astrocitos/metabolismo , Astrocitos/patología , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Corteza Cerebral/metabolismo , Femenino , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Procesamiento de Imagen Asistido por Computador , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Neuroglía/metabolismo , Caracteres Sexuales , Heridas Punzantes/metabolismoRESUMEN
Following brain injury astrocytes change into a reactive state, proliferate and grow into the site of lesion, a process called astrogliosis, initiated and regulated by changes in cytoplasmic Ca2+ . Transient receptor potential canonical (TRPC) channels may contribute to Ca2+ influx but their presence and possible function in astrocytes is not known. By RT-PCR and RNA sequencing we identified transcripts of Trpc1, Trpc2, Trpc3, and Trpc4 in FACS-sorted glutamate aspartate transporter (GLAST)-positive cultured mouse cortical astrocytes and subcloned full-length Trpc1 and Trpc3 cDNAs from these cells. Ca2+ entry in cortical astrocytes depended on TRPC3 and was increased in the absence of Trpc1. After co-expression of Trpc1 and Trpc3 in HEK-293 cells both proteins co-immunoprecipitate and form functional heteromeric channels, with TRPC1 reducing TRPC3 activity. In vitro, lack of Trpc3 reduced astrocyte proliferation and migration whereas the TRPC3 gain-of-function moonwalker mutation and Trpc1 deficiency increased astrocyte migration. In vivo, astrogliosis and cortex edema following stab wound injury were reduced in Trpc3-/- but increased in Trpc1-/- mice. In summary, our results show a decisive contribution of TRPC3 to astrocyte Ca2+ signaling, which is even augmented in the absence of Trpc1, in particular following brain injury. Targeted therapies to reduce TRPC3 channel activity in astrocytes might therefore be beneficial in traumatic brain injury.
Asunto(s)
Astrocitos/metabolismo , Señalización del Calcio/fisiología , Corteza Cerebral/lesiones , Gliosis/metabolismo , Canales Catiónicos TRPC/metabolismo , Animales , Astrocitos/patología , Edema Encefálico/etiología , Edema Encefálico/metabolismo , Edema Encefálico/patología , Movimiento Celular/fisiología , Proliferación Celular/fisiología , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Modelos Animales de Enfermedad , Gliosis/etiología , Gliosis/patología , Células HEK293 , Humanos , Masculino , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Canales Catiónicos TRPC/genética , Canal Catiónico TRPC6 , Heridas Punzantes/metabolismo , Heridas Punzantes/patologíaRESUMEN
Compelling evidence now points to the critical role of the cytoskeleton in neurodegeneration. In the present study, using an immunohistochemical approach, we have shown that cortical stab injury (CSI) in adult Wistar rats significantly affects temporal pattern of expression of neurofilament proteins (NFs), a major cytoskeleton components of neurons, and microtubule-associated proteins (MAP2). At 3 days post-injury (dpi) most of the NFs immunoreactivity was found in pyknotic neurons and in fragmentized axonal processes in the perilesioned cortex. These cytoskeletal alterations became more pronounced by 10dpi. At the subcellular level CSI also showed significant impact on NFs and MAP-2 expression. Thus, at 3dpi most of the dendrites disappeared, while large neuronal somata appeared like open circles pointing to membrane disintegration. Conversely, at 10dpi neuronal perikarya and a few new apical dendrites were strongly labeled. Since aberrant NF phosphorylation is a pathological hallmark of many human neurodegenerative disorders, as well as is found after stressor stimuli, the present results shed light into the expression of neurofilaments after the stab brain injury.
Asunto(s)
Lesiones Encefálicas/metabolismo , Corteza Cerebral/metabolismo , Regulación de la Expresión Génica , Proteínas Asociadas a Microtúbulos/biosíntesis , Proteínas del Tejido Nervioso/biosíntesis , Proteínas de Neurofilamentos/biosíntesis , Heridas Punzantes/metabolismo , Animales , Lesiones Encefálicas/patología , Corteza Cerebral/patología , Masculino , Ratas , Ratas Wistar , Heridas Punzantes/patologíaRESUMEN
The timing of skin wounds is one of the most challenging problems in forensic pathology. In the first minutes or hours after infliction, histological examination fails to determine whether a wound was sustained before or after death. The aim of this study was to evaluate the use of three immunohistochemical markers (FVIIIra, CD15, and tryptase) for the interpretation of the timing of cutaneous stab wounds. We evaluated these markers in intravital wounds from autopsy cases (n = 12) and surgical specimens (n = 58). As controls, we used normal skin samples from autopsies (n = 8) and an original ex vivo surgical human model of recent postmortem wounds (n = 24). We found overexpression of FVIIIra in 100 % of vital wounds, but also in 53 % of the controls. The number of CD15-positive cells was higher in wound margins than in internal controls (p < 0.0001) and was significantly correlated with the time interval between incision and devascularization (p = 0.0005; minimal time for positivity, 9 min). Using the anti-tryptase antibody, we found that the mast cell degranulation rate was higher in wound margins (p < 0.0001) and correlated with the time interval (minimal time, 1 min). The sensitivity and specificity for the diagnosis of vitality were respectively 100 and 47 % for FVIIIra, 47 and 100 % for CD15, and 60 and 100 % for tryptase. The inter-observer agreement coefficients were 0.68 for FVIIIra, 0.90 for CD15, and 0.46 for tryptase. Finally, we demonstrated that these markers were not reliable in putrefied or desiccated specimens. In conclusion, CD15 and tryptase, but not FVIIIra, may be useful markers for differentiating recent antemortem from postmortem injuries.
Asunto(s)
Antígeno Lewis X/metabolismo , Piel/metabolismo , Triptasas/metabolismo , Heridas Punzantes/metabolismo , Heridas Punzantes/patología , Factor de von Willebrand/metabolismo , Biomarcadores/metabolismo , Estudios de Casos y Controles , Degranulación de la Célula , Patologia Forense , Hemorragia/patología , Humanos , Inmunohistoquímica , Mastocitos/patología , Neutrófilos/metabolismo , Cambios Post Mortem , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Piel/lesiones , Piel/patología , Factores de Tiempo , Cicatrización de HeridasRESUMEN
During injury to the central nervous system (CNS), astrocytes and microglia proliferate and migrate around the lesion sites. Recently, it has been reported that one of the water channels, aquaporin-4 (AQP4) is seemed to have a role in astroglial migration and glial scar formation caused by brain injury, although its molecular mechanism is largely unknown. In the present study, we examined the expression profiles in wild-type (WT) and AQP4-deficient (AQP4/KO) mice after a stab wound to the cerebral cortex. Three days after the stab wound, AQP4 expression was observed in activated microglia around the lesion site as well as in astrocytes. A microarray analysis revealed that 444 genes around the lesion site were upregulated 3 days after the wounding in WT mice. Surprisingly, most of these up-regulations were significantly attenuated in AQP4/KO mice. Real-time RT-PCR and immunofluorescence showed that osteopontin (OPN) expression around the lesion site was much lower in AQP4/KO mice than in WT mice. Moreover, the up-regulation of pro-inflammatory cytokines was significantly attenuated in AQP4/KO mice. Taken together, these results suggest that AQP4 plays an important role in immunological function in concert with OPN under pathological conditions in the CNS.
Asunto(s)
Acuaporina 4/metabolismo , Corteza Cerebral/lesiones , Osteopontina/metabolismo , Heridas Punzantes/metabolismo , Animales , Acuaporina 4/genética , Astrocitos/metabolismo , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Perfilación de la Expresión Génica , Inflamación/genética , Inflamación/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/metabolismo , Osteopontina/genética , Regulación hacia Arriba , Heridas Punzantes/inmunología , Heridas Punzantes/patologíaRESUMEN
Reactive gliosis, in which astrocytes as well as other types of glial cells undergo massive proliferation, is a common hallmark of all brain pathologies. Brain-type fatty acid-binding protein (FABP7) is abundantly expressed in neural stem cells and astrocytes of developing brain, suggesting its role in differentiation and/or proliferation of glial cells through regulation of lipid metabolism and/or signaling. However, the role of FABP7 in proliferation of glial cells during reactive gliosis is unknown. In this study, we examined the expression of FABP7 in mouse cortical stab injury model and also the phenotype of FABP7-KO mice in glial cell proliferation. Western blotting showed that FABP7 expression was increased significantly in the injured cortex compared with the contralateral side. By immunohistochemistry, FABP7 was localized to GFAP(+) astrocytes (21% of FABP7(+) cells) and NG2(+) oligodendrocyte progenitor cells (62%) in the normal cortex. In the injured cortex there was no change in the population of FABP7(+)/NG2(+) cells, while there was a significant increase in FABP7(+)/GFAP(+) cells. In the stab-injured cortex of FABP7-KO mice there was decrease in the total number of reactive astrocytes and in the number of BrdU(+) astrocytes compared with wild-type mice. Primary cultured astrocytes from FABP7-KO mice also showed a significant decrease in proliferation and omega-3 fatty acid incorporation compared with wild-type astrocytes. Overall, these data suggest that FABP7 is involved in the proliferation of astrocytes by controlling cellular fatty acid homeostasis.
Asunto(s)
Astrocitos/metabolismo , Corteza Cerebral/lesiones , Corteza Cerebral/metabolismo , Proteínas de Unión a Ácidos Grasos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Heridas Punzantes/metabolismo , Animales , Animales Recién Nacidos , Antígenos/metabolismo , Astrocitos/patología , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos/genética , Ácidos Grasos Omega-3/metabolismo , Expresión Génica , Gliosis/metabolismo , Gliosis/patología , Homeostasis , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Células-Madre Neurales/metabolismo , Células-Madre Neurales/patología , Oligodendroglía/metabolismo , Oligodendroglía/patología , Proteoglicanos/metabolismo , Heridas Punzantes/genética , Heridas Punzantes/patologíaRESUMEN
NG2 cells are an abundant glial cell type in the adult brain. They are distinct from astrocytes, mature oligodendrocytes, and microglia. NG2 cells generate oligodendrocytes and a subpopulation of protoplasmic astrocytes in the ventral forebrain during development. To determine whether NG2 cells generate reactive astrocytes in the lesioned brain, stab wound injury was created in adult NG2creBAC:ZEG double transgenic mice, in which enhanced green fluorescent protein (EGFP) is expressed in NG2 cells and their progeny, and the phenotype of the EGFP(+) cells was analyzed at 10 and 30 days post lesion (dpl). The majority (>90%) of the reactive astrocytes surrounding the lesion that expressed glial fibrillary acidic protein (GFAP) lacked EGFP expression, and conversely the majority (>90%) of EGFP(+) cells were GFAP-negative. However, 8% of EGFP(+) cells co-expressed GFAP at 10 dpl. Most of these EGFP(+) GFAP(+) cells were morphologically distinct from hypertrophic reactive astrocytes and exhibited weak GFAP expression. NG2 was detected in a fraction of the EGFP(+) GFAP(+) cells found at 10 dpl. By 30 dpl the number of EGFP(+) GFAP(+) cells had decreased more than four-fold from 10 dpl. A similar transient appearance of EGFP(+) GFAP(+) cells with simple morphology was observed in NG2creER™:ZEG double transgenic mice in which EGFP expression had been induced in NG2 cells prior to injury. NG2 cell-specific deletion of the oligodendrocyte lineage transcription factor Olig2 using NG2creER™:Olig2(fl/fl) :ZEG triple transgenic mice did not increase the number of EGFP(+) reactive astrocytes. These findings suggest that NG2 cells are not a major source of reactive astrocytes in the neocortex.
Asunto(s)
Astrocitos/citología , Diferenciación Celular/fisiología , Neocórtex/lesiones , Neuroglía/citología , Animales , Astrocitos/metabolismo , Lesiones Encefálicas/metabolismo , Recuento de Células , Proteína Ácida Fibrilar de la Glía/metabolismo , Inmunohistoquímica , Ratones , Ratones Transgénicos , Neocórtex/citología , Neocórtex/metabolismo , Neuroglía/metabolismo , Heridas Punzantes/metabolismoRESUMEN
INTRODUCTION: Although the involvement of plasminogen in liver repair has been reported, its roles are still poorly understood. Here, we investigated the role of plasminogen in accumulations of macrophages and neutrophils after liver injury in mice with gene deficient of plasminogen (Plg(-/-)) or its wild type (Plg(+/+)). MATERIALS AND METHODS: Mice received traumatic liver injury caused by stabbing on the lobe or hepatic ischemia-reperfusion, and the damaged sites were histologically analyzed. RESULTS: After the traumatic liver injury, both the stab wound and the damaged tissue were decreased until day 7 in the Plg(+/+) mice. In contrast, both the stab wound and the damaged tissue were still remained until day 7 in the Plg(-/-) mice. On day 4 after traumatic liver injury, macrophages were abundant at the surrounding area of the damaged site in the Plg(+/+) mice. However, the macrophage accumulation was impaired in the Plg(-/-) mice. After hepatic ischemia-reperfusion injury, macrophage accumulation and decrease in the damaged tissue were also observed in the Plg(+/+) mice until day 7. In contrast, these responses were also impaired in the Plg(-/-) mice. Furthermore, neutrophil accumulation at the surrounding area of the damaged site was also impaired in the Plg(-/-) mice on day 4 after both liver traumatic liver injury and hepatic ischemia-reperfusion injury. CONCLUSIONS: Our data indicate that plasminogen plays a crucial role in macrophage accumulation together with the neutrophil accumulation after liver injury in both models, which may be essential for triggering the subsequent healing responses including decrease in the damaged tissue.
Asunto(s)
Hígado/lesiones , Hígado/metabolismo , Macrófagos/metabolismo , Macrófagos/patología , Plasminógeno/metabolismo , Heridas Punzantes/metabolismo , Heridas Punzantes/patología , Animales , Hígado/patología , Ratones , Ratones Noqueados , Plasminógeno/genética , Regeneración/fisiologíaRESUMEN
During a variety of insults to the brain adenine nucleotides are released in large quantities from damaged cells, triggering local cellular and biochemical responses to injury. Different models of brain injury reveal that the local increase in adenine nucleotides levels is followed by a compensatory up-regulation of ectonucleotidase enzymes that catalyze sequential hydrolysis of ATP to ADP, AMP and adenosine. However, recent studies imply that changes in adenine nucleotides release may also occur in the areas distant from the site of direct damage. Therefore, in the present study we have used the model of cortical stab injury to analyze extracellular ATP, ADP and AMP hydrolysis in the membrane preparations obtained from the brain regions that were not subjected to direct tissue damage. The brain regions analyzed were contralateral cortex, hippocampus, caudate nucleus, thalamus and hypothalamus. It was evidenced that cortical stab injury induced early widespread decrease in AMP hydrolysis in all brain areas tested, except in the hypothalamus, without changes in ATP hydrolysis. These findings imply that brain injury affects global extracellular adenine nucleotide and nucleoside levels, consequently affecting neuronal function in the regions distant to the primary damage.
Asunto(s)
5'-Nucleotidasa/metabolismo , Lesiones Encefálicas/enzimología , Heridas Punzantes/enzimología , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Lesiones Encefálicas/metabolismo , Lesiones Encefálicas/patología , Membrana Celular/metabolismo , Espacio Extracelular/metabolismo , Hidrólisis , Masculino , Ratas , Ratas Wistar , Heridas Punzantes/metabolismo , Heridas Punzantes/patologíaRESUMEN
Basic fibroblast growth factor (bFGF) is a highly conserved and ubiquitously distributed mitogen, and much is known at the molecular level. However the available information about in vivo distribution in human tissues and expression changes in relation to causes of death is not sufficient. The present study investigated 35 autopsy cases, comprising five cases for each cause of death: acute myocardial infarction/ischemia (AMI), mechanical asphyxiation, blunt brain injury, drowning, hypothermia, intoxication and sharp instrument injury. The bFGF immunopositivity was detected mainly in interstitial cells and sporadically in cardiomyocytes in the heart, in macrophages in the lung, astrocytes in the brain, mainly in the sinusoidal Kupffer cells and partly in the hepatocytes in the liver, red pulp of the spleen, pancreatic islets and proximal convoluted tubules and corpuscles in the kidney. Immunopositivity was frequently detected in the lung and liver for AMI and hypothermia, and in the kidney for AMI, mechanical asphyxiation, drowning and injuries, but was not evident in the kidney for hypothermia. Positivity in these tissues varied by case in other causes of death. High positivity in the brain was seen for intoxication, but AMI, mechanical asphyxiation and drowning showed lower positivity. For the heart, spleen and pancreas, there was no evident difference among the causes of death. These findings suggested that bFGF expression in the lung, liver, kidney and brain varies depending on the cause of death, and is useful for investigating deaths.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/metabolismo , Adulto , Anciano , Asfixia/metabolismo , Astrocitos/metabolismo , Encéfalo/citología , Encéfalo/metabolismo , Lesiones Encefálicas/metabolismo , Ahogamiento/metabolismo , Femenino , Patologia Forense , Hepatocitos/metabolismo , Humanos , Hipotermia/metabolismo , Inmunohistoquímica , Islotes Pancreáticos/metabolismo , Riñón/metabolismo , Macrófagos del Hígado/metabolismo , Macrófagos Alveolares/metabolismo , Masculino , Persona de Mediana Edad , Infarto del Miocardio/metabolismo , Miocitos Cardíacos/metabolismo , Intoxicación/metabolismo , Bazo/metabolismo , Heridas Punzantes/metabolismoRESUMEN
We have analysed the expression of the endogenous angiogenesis inhibitor endostatin/collagen XVIII following stab wound injury and observed a highly significant (p<0.0001) lesional accumulation confined to areas of pan-necrotic injury and developing secondary damage. Maximal cell numbers were detected at Day 14, declining until Day 21 after injury. Further, endostatin/collagen XVIII(+) monocytic cells accumulated in Virchow-Robin spaces where they formed cell clusters. Besides being prevailingly localised to ED1(+) activated microglia/macrophages, endostatin/collagen XVIII expression was also detected by subendothelial surrounding vessels in the lesioned area. Late and prolonged accumulation of endostatin/collagen XVIII(+) microglia/macrophages and increased numbers of endostatin/collagen XVIII(+) subendothelial cells/vessels in areas of vascular pruning and regression, point to a role in the termination of the transient angiogenic response, linked to a "late" inflammatory milieu.
Asunto(s)
Inhibidores de la Angiogénesis/metabolismo , Lesiones Encefálicas/metabolismo , Encéfalo/metabolismo , Colágeno Tipo XVIII/metabolismo , Endostatinas/metabolismo , Heridas Punzantes/metabolismo , Animales , Vasos Sanguíneos/fisiopatología , Encéfalo/irrigación sanguínea , Encéfalo/patología , Lesiones Encefálicas/patología , Lesiones Encefálicas/fisiopatología , Macrófagos/metabolismo , Macrófagos/patología , Masculino , Microglía/metabolismo , Microglía/patología , Necrosis , Ratas , Ratas Sprague-Dawley , Distribución Tisular , Heridas Punzantes/patología , Heridas Punzantes/fisiopatologíaRESUMEN
The mechanisms by which neural and glial progenitor cells in the adult brain respond to tissue injury are unknown. We studied the responses of these cells to stab wound injury in rats and in two transgenic mouse models in which Y/GFP is driven either by Sox2 (a neural stem cell marker) or by Talpha-1 (which marks newly born neurons). The response of neural progenitors was low in all nonneurogenic regions, and no neurogenesis occurred at the injury site. Glial progenitors expressing Olig2 and NG2 showed the greatest response. The appearance of these progenitors preceded the appearance of reactive astrocytes. Surprisingly, we found evidence of the translocation of the transcription factor Olig2 into cytoplasm in the first week after injury, a mechanism that is known to mediate the differentiation of astrocytes during brain development. Translocation of Olig2, down-regulation of NG2, and increased glial fibrillary acidic protein expression were recapitulated in vitro after exposure of glial progenitors to serum components or bone morphogentic protein by up-regulation of Notch-1. The glial differentiation and Olig2 translocation could be blocked by inhibition of Notch-1 with the gamma-secretase inhibitor DAPT. Together, these data indicate that the prompt maturation of numerous Olig2(+) glial progenitors to astrocytes underlies the repair process after a traumatic injury. In contrast, neural stem cells and neuronal progenitor cells appear to play only a minor role in the injured adult CNS.