RESUMEN
Cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathyis a rare form of inherited cerebral small vessel disease associated with mutations in the high-temperature requirement serine peptidase A1 gene. As of now, only about 50 cases have been reported. In 2012, our group reported a family with a novel mutant of the high-temperature requirement serine peptidase A1 gene in China for the first time. To further explore the molecular pathogenesis of cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy, a recombination mouse model expressed human high-temperature requirement serine peptidase A1 gene mutant identified by our group was generated using the Donor & Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 system and termed the Mut-high-temperature requirement serine peptidase A1 geneL364P mouse model. Results show that Mut-high-temperature requirement serine peptidase A1 geneL364P mice present similar pathological characteristics to patients with cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy, suggesting that the Mut-high-temperature requirement serine peptidase A1 geneL364P mouse model was generated successfully. Moreover, apoptosis was induced in mouse brain vascular smooth muscle cells derived from Mut-high-temperature requirement serine peptidase A1 geneL364P mice. In summary, the cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy mouse model described in this study will be beneficial to demonstrate the pathological mechanism of cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy and provide new therapeutic targets for clinical treatment.
Asunto(s)
Alopecia/genética , Encéfalo/irrigación sanguínea , Infarto Cerebral/genética , Serina Peptidasa A1 que Requiere Temperaturas Altas/genética , Leucoencefalopatías/genética , Mutación , Enfermedades de la Columna Vertebral/genética , Alopecia/enzimología , Alopecia/patología , Animales , Apoptosis , Células Cultivadas , Infarto Cerebral/enzimología , Infarto Cerebral/patología , Predisposición Genética a la Enfermedad , Serina Peptidasa A1 que Requiere Temperaturas Altas/metabolismo , Leucoencefalopatías/enzimología , Leucoencefalopatías/patología , Ratones Endogámicos C57BL , Ratones Transgénicos , Músculo Liso Vascular/enzimología , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/enzimología , Miocitos del Músculo Liso/patología , Fenotipo , Enfermedades de la Columna Vertebral/enzimología , Enfermedades de la Columna Vertebral/patologíaRESUMEN
Recent works highlight the therapeutic potential of targeting cyclic guanosine monophosphate (cGMP)-dependent pathways in the context of brain ischemia/reperfusion injury (IRI). Although cGMP-dependent protein kinase I (cGKI) has emerged as a key mediator of the protective effects of nitric oxide (NO) and cGMP, the mechanisms by which cGKI attenuates IRI remain poorly understood. We used a novel, conditional cGKI knockout mouse model to study its role in cerebral IRI. We assessed neurological deficit, infarct volume, and cerebral perfusion in tamoxifen-inducible vascular smooth muscle cell-specific cGKI knockout mice and control animals. Stroke experiments revealed greater cerebral infarct volume in smooth muscle cell specific cGKI knockout mice (males: 96 ± 16 mm3; females: 93 ± 12 mm3, mean±SD) than in all control groups: wild type (males: 66 ± 19; females: 64 ± 14), cGKI control (males: 65 ± 18; females: 62 ± 14), cGKI control with tamoxifen (males: 70 ± 8; females: 68 ± 10). Our results identify, for the first time, a protective role of cGKI in vascular smooth muscle cells during ischemic stroke injury. Moreover, this protective effect of cGKI was found to be independent of gender and was mediated via improved reperfusion. These results suggest that cGKI in vascular smooth muscle cells should be targeted by therapies designed to protect brain tissue against ischemic stroke.
Asunto(s)
Infarto Cerebral/enzimología , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/metabolismo , Músculo Liso Vascular/enzimología , Miocitos del Músculo Liso/enzimología , Daño por Reperfusión/enzimología , Accidente Cerebrovascular/enzimología , Animales , Infarto Cerebral/genética , Infarto Cerebral/patología , Proteína Quinasa Dependiente de GMP Cíclico Tipo I/genética , Femenino , Masculino , Ratones , Ratones Noqueados , Músculo Liso Vascular/patología , Miocitos del Músculo Liso/patología , Daño por Reperfusión/genética , Daño por Reperfusión/patología , Accidente Cerebrovascular/genética , Accidente Cerebrovascular/patologíaRESUMEN
There is a strong association between lipoprotein-associated phospholipase A2 (Lp-PLA2) levels and atherosclerosis-related diseases. The aim of this study was to investigate the role of Lp-PLA2 in the ischemic stroke and further offer clinical evidence that measuring Lp-PLA2 helps predict the risk of stroke occurrence and recurrence. A total of 328 hospitalized patients were recruited, including 179 cases of acute cerebral infarction (ACI) and 149 non-ACI controls. The serum level of Lp-PLA2 in ACI was significantly higher than non-ACI. The serum level of Lp-PLA2 in the recurrence of ACI was significantly higher than the nonrecurrence. The serum levels of Lp-PLA2 in large-artery atherosclerosis subtype were the highest among the subtypes of the Trial of Org 10172 in Acute Stroke Treatment and non-ACI controls. The level of Lp-PLA2 in large-artery atherosclerosis and the cardioembolism group was statistically significantly higher than that of the control cases. There was no statistically significant difference between the small-vessel occlusion group and the control cases. The present study confirmed that the elevated Lp-PLA2 level can be a risk factor for ischemic stroke in the Chinese population. The serum level of Lp-PLA2 may be a predictive factor for the recurrence of ACI.
Asunto(s)
Infarto Cerebral/sangre , Infarto Cerebral/enzimología , Fosfolipasas A2/sangre , Adulto , Biomarcadores/sangre , Infarto Cerebral/terapia , China , Femenino , Hospitalización , Humanos , Arteriosclerosis Intracraneal/sangre , Arteriosclerosis Intracraneal/enzimología , Arteriosclerosis Intracraneal/terapia , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Recurrencia , Factores de RiesgoRESUMEN
Increased plasma levels of neuron-specific enolase (NSE) are related to damage of neurons and neuroendocrine cells. We aimed to investigate elevation of NSE after elective percutaneous coronary intervention (PCI) on the prediction of silent cerebral infarct (SCI). Study population consisted of 2 groups of patients. Group 1 included 92 consecutive patients with normal coronary angiograms, whereas group 2 consisted of 89 patients who underwent elective coronary stenting. NSE levels were studied before and 12 hours after the procedure. Elevation of >0.12 µg/L was considered as SCI. Forty-seven of 181 study patients (26%) had SCI after the procedure. NSE elevation was significantly more prevalent in patients with PCI than that of controls. Elevation of NSE was observed in 42% of patients who underwent elective PCI (n = 37) and 11% of the normal coronary artery group (n = 10) (p <0.001). The incidence of SCI was higher in active smokers and patients who had history of myocardial infarction (MI) (55% vs 10%, p <0.001 for active smokers and 40% vs 8%, p <0.001 for history of MI, respectively). Multivariate analysis demonstrated history of smoking (odds ratio [OR] 9.9; 95% confidence interval [CI] 3.7 to 26.9; p <0.001) and previous MI (OR 4.4; 95% CI 1.7 to 11.4; p = 0.01) as independent predictors of SCI. For patients who underwent elective PCI, NSE levels after procedure increases. Invasive coronary procedures have risk of SCIs, even in patients with normal coronary arteries. In conclusion, increased diagnosis of SCIs might improve understanding of their relation with invasive cardiac procedures, facilitate to prevent occurrence of silent microemboli and decrease the risk of adverse neurologic events.
Asunto(s)
Infarto Cerebral/enzimología , Estenosis Coronaria/cirugía , Intervención Coronaria Percutánea/efectos adversos , Fosfopiruvato Hidratasa/sangre , Medición de Riesgo/métodos , Stents , Biomarcadores/sangre , Infarto Cerebral/epidemiología , Infarto Cerebral/etiología , Estenosis Coronaria/enzimología , Femenino , Estudios de Seguimiento , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Pronóstico , Factores de Riesgo , Factores de Tiempo , Turquía/epidemiologíaRESUMEN
OBJECTIVE: To elucidate the molecular mechanism of mutant HTRA1-dependent cerebral small vessel disease in heterozygous individuals. METHODS: We recruited 113 unrelated index patients with clinically diagnosed cerebral small vessel disease. The coding sequences of the HTRA1 gene were analyzed. We evaluated HTRA1 protease activities using casein assays and oligomeric HTRA1 formation using gel filtration chromatography. RESULTS: We found 4 heterozygous missense mutations in the HTRA1 gene (p.G283E, p.P285L, p.R302Q, and p.T319I) in 6 patients from 113 unrelated index patients and in 2 siblings in 2 unrelated families with p.R302Q. The mean age at cognitive impairment onset was 51.1 years. Spondylosis deformans was observed in all cases, whereas alopecia was observed in 3 cases; an autopsied case with p.G283E showed arteriopathy in their cerebral small arteries. These mutant HTRA1s showed markedly decreased protease activities and inhibited wild-type HTRA1 activity, whereas 2 of 3 mutant HTRA1s reported in cerebral autosomal-recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL) (A252T and V297M) did not inhibit wild-type HTRA1 activity. Wild-type HTRA1 forms trimers; however, G283E and T319I HTRA1, observed in manifesting heterozygotes, did not form trimers. P285L and R302Q HTRA1s formed trimers, but their mutations were located in domains that are important for trimer-associated HTRA1 activation; in contrast, A252T and V297M HTRA1s, which have been observed in CARASIL, also formed trimers but had mutations outside the domains important for trimer-associated HTRA1 activation. CONCLUSIONS: The mutant HTRA1s observed in manifesting heterozygotes might result in an impaired HTRA1 activation cascade of HTRA1 or be unable to form stable trimers.
Asunto(s)
Alopecia/enzimología , Alopecia/genética , Infarto Cerebral/enzimología , Infarto Cerebral/genética , Heterocigoto , Leucoencefalopatías/enzimología , Leucoencefalopatías/genética , Mutación Missense , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Enfermedades de la Columna Vertebral/enzimología , Enfermedades de la Columna Vertebral/genética , Alopecia/diagnóstico por imagen , Alopecia/patología , Encéfalo/diagnóstico por imagen , Encéfalo/enzimología , Encéfalo/patología , Infarto Cerebral/diagnóstico por imagen , Infarto Cerebral/patología , Cromatografía en Gel , Dimerización , Familia , Serina Peptidasa A1 que Requiere Temperaturas Altas , Humanos , Leucoencefalopatías/diagnóstico por imagen , Leucoencefalopatías/patología , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Modelos Moleculares , Linaje , Análisis de Secuencia de ADN , Enfermedades de la Columna Vertebral/diagnóstico por imagen , Enfermedades de la Columna Vertebral/patologíaRESUMEN
The aim of the present study was to evaluate the effect of electroacupuncture (EA) on cognitive function following cerebral ischemiareperfusion (I/R) injury, and to clarify the mechanism through which Rho GTPase is associated with EA analgesia modulation of dendritic spine plasticity. Rats were randomly divided into three groups: The sham surgery group, the middle cerebral artery occlusion (MCAO) model of ischemia group, and the MCAO with EA (MCAO+EA) treatment group. The MCAO+EA group received treatment with EA at points of Baihui (DU20) and Shenting (DU24) following surgery. It was demonstrated that treatment with EA significantly (P<0.05) protected the cognitive function of rats from impairment caused by cerebral I/R injury. Furthermore, EA treatment increased the density of dendritic spines in the hippocampus of cerebral I/Rinjured rats. Simultaneously, EA increased the expression of cell division cycle 42, Rasrelated C3 botulinum toxin substrate 1 and Factin proteins. By contrast, EA treatment inhibited the expression of Ras homologous member A. Collectively, these findings suggest that Rho GTPases and dendritic spine plasticity are critical in mediating the effects of EA treatment at the points of Shenting and Baihui, and that EA protects against impairment of cognitive function following ischemic stroke.
Asunto(s)
Disfunción Cognitiva/terapia , Espinas Dendríticas/fisiología , Electroacupuntura , Plasticidad Neuronal , Daño por Reperfusión/terapia , Animales , Infarto Cerebral/enzimología , Infarto Cerebral/etiología , Infarto Cerebral/patología , Infarto Cerebral/terapia , Disfunción Cognitiva/enzimología , Disfunción Cognitiva/etiología , Masculino , Aprendizaje por Laberinto , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/enzimología , Proteínas de Unión al GTP rho/metabolismoRESUMEN
OBJECTIVE: To observe the effect of electroacupuncture (EA) intervention on the neurological function and the expression change of Slit-Robo GTPase-activating protein-1 (srGAP 1) and cell division-cycle 42 (Cdc 42) in the cortex of rats with cerebral ischemic injury (CIRI) , so as to explore the mechanism of EA in the management of cerebral infarction. METHODS: A total of 48 male Sprague Dawley (SD) rats were randomly and equally divided into control, model, non-acupoint EA and EA groups (n = 12/group). The CIRI model was established based on the modified Zea Longa method. EA intervention was applied for 30 min, once a day for 14 days. Modified neurologic severity scores (mNSS) were assessed on day 1,3,7 and 14 after mode- ling. Immunofluorescence assay was used to detect the immunoactivity and distribution of srGAP 1 and Cdc 42 in the cortical ischemic region. Western blot was employed to detect the expression of srGAP 1 and Cdc 42 in the affected cortex. RESULTS: The mNSS displayed that the neurological score in the EA group was significantly lower than that in the model group and non-acupoint EA group at the 7th d and 14th d (P<0. 01). Immunofluorescence results showed that cerebral srGAP 1 and Cdc 42 were ex- pressed mainly in the cytoplasm. The fluorescence intensity of srGAP 1 of the EA group was significantly lower than that of the model group and non-acupoint EA group(P<0. 01). Meanwhile the fluorescence intensity of Cdc 42 of the EA group was markedly higher than that in the model group and non-acupoint EA group(P<0. 01). Western blot assay indicated that the expression level of srGAP 1 in the model group was significantly higher than that of the control group( P<0. 01) ,and that of the EA group was much lower than those of the model group and non-acupoint EA group(P<0. 01). There was no significant difference of srGAP 1 expression levels between the non-acupoint EA group and the model group(P>0. 05). Additionally, the protein expression of Cdc 42 in the model group was slightly higher than that of the control group(P>0. 05), and that of the EA group was significantly higher than those of the model group and non-acupoint EA group(P<0. 01). There was no significant difference of Cdc 42 expression levels between the non-acupoint EA group and the model group(P>0. 05). CONCLUSION: Cerebral infarction induced increase of cerebral srGAP 1 and decrease of Cdc 42 can be reversed by acupoint EA intervention in CIRI rats, which may be responsible for its effect in improving impaired neurological function after cerebral infarction.
Asunto(s)
Infarto Cerebral/terapia , Electroacupuntura , Proteínas Activadoras de GTPasa/genética , Proteína de Unión al GTP cdc42/genética , Animales , Infarto Cerebral/enzimología , Infarto Cerebral/genética , Modelos Animales de Enfermedad , Proteínas Activadoras de GTPasa/metabolismo , Humanos , Masculino , Ratas , Ratas Sprague-Dawley , Proteína de Unión al GTP cdc42/metabolismoRESUMEN
OBJECTIVE: To observe the effect of electroacupuncture (EA) intervention on expression and content of protein kinase C (PKC) in the middle cerebral artery in acute cerebral infarction (ACI) rats so as to explore its mechanism underlying improvement of ACI. METHODS: Wistar rats were randomly divided into normal control (n = 6), sham operation (n = 30), ACI model (n = 30), and EA (n = 30) groups, and the latter three groups were further divided into 0. 5 h, 1 h, 3 h, 6 h and 12 h subgroups (n = 6 in each subgroup). The ACI model was established by occlusion of the middle cerebral artery (MCAO). EA (15 Hz, 1 mA) was applied to "Shuigou" (GV 26) for 20 min. The PKC expression levels and activity in the vascular smooth muscle of the middle cerebral artery were detected using immunohistochemistry and ELISA, respectively. RESULTS: In comparison with the control group, the immunoactivity and activities of PKC in the middle cerebral artery tissue at 0. 5 h, 1 h, 3 h, 6 h and 12 h were significantly increased in the model group (P<0. 05). After EA intervention, the expression levels and activities of PKC at the 5 time-points were markedly down-regulated in comparison with the model group at the same corresponding time-point (P<0. 05). No significant changes of PKC expression and activity were found in the sham operation group (P>0. 05). CONCLUSION: EA intervention can up-regulate the immunoactivity and activity of PKC in the vascular smooth muscle of the middle cerebral artery in ACI rats, which may contribute to its effect in improving ACI by relieving arterial spasm.
Asunto(s)
Puntos de Acupuntura , Infarto Cerebral/terapia , Electroacupuntura , Arteria Cerebral Media/enzimología , Proteína Quinasa C/metabolismo , Animales , Infarto Cerebral/enzimología , Modelos Animales de Enfermedad , Humanos , Masculino , Ratas , Ratas WistarRESUMEN
BACKGROUND: Cerebral infarction due to thrombosis leads to the most common type of stroke and a likely cause of age-related cognitive decline and dementia. Endothelial nitric oxide synthase (eNOS) generates NO, which plays a crucial role in maintaining vascular function and exerting an antithrombotic action. Reduced eNOS expression and eNOS polymorphisms have been associated with stroke and Alzheimer's disease (AD), the most common type of dementia associated with neurovascular dysfunction. However, direct proof of such association is lacking. Since there are no reports of complete eNOS deficiency in humans, we used heterozygous eNOS(+/-) mice to mimic partial deficiency of eNOS, and determine its impact on cerebrovascular pathology and perfusion of cerebral vessels. RESULTS: Combining cerebral angiography with immunohistochemistry, we found thrombotic cerebral infarctions in eNOS(+/-) mice as early as 3-6 months of age but not in eNOS(+/+) mice at any age. Remarkably, vascular occlusions in eNOS(+/-) mice were found almost exclusively in three areas: temporoparietal and retrosplenial granular cortexes, and hippocampus this distribution precisely matching the hypoperfused areas identified in preclinical AD patients. Moreover, progressive cerebral amyloid angiopaphy (CAA), blood brain barrier (BBB) breakdown, and cognitive impairment were also detected in aged eNOS(+/-) mice. CONCLUSIONS: These data provide for the first time the evidence that partial eNOS deficiency results in spontaneous thrombotic cerebral infarctions that increase with age, leading to progressive CAA and cognitive impairments. We thus conclude that eNOS(+/-) mouse may represent an ideal model of ischemic stroke to address early and progressive damage in spontaneously-evolving chronic cerebral ischemia and thus, study vascular mechanisms contributing to vascular dementia and AD.
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Barrera Hematoencefálica/fisiología , Angiopatía Amiloide Cerebral/genética , Infarto Cerebral/genética , Trastornos del Conocimiento/genética , Óxido Nítrico Sintasa de Tipo III/fisiología , Envejecimiento , Animales , Angiopatía Amiloide Cerebral/patología , Infarto Cerebral/enzimología , Infarto Cerebral/patología , Infarto Cerebral/psicología , Circulación Cerebrovascular , Trastornos del Conocimiento/enzimología , Trastornos del Conocimiento/patología , Progresión de la Enfermedad , Endotelio Vascular/fisiopatología , Regulación de la Expresión Génica , Heterocigoto , Hipocampo/irrigación sanguínea , Hipocampo/enzimología , Hipocampo/patología , Aprendizaje por Laberinto , Trastornos de la Memoria/enzimología , Trastornos de la Memoria/genética , Trastornos de la Memoria/patología , Ratones , Ratones Mutantes Neurológicos , Proteínas del Tejido Nervioso/biosíntesis , Proteínas del Tejido Nervioso/genética , Óxido Nítrico Sintasa de Tipo III/deficiencia , Óxido Nítrico Sintasa de Tipo III/genética , Especificidad de Órganos , Lóbulo Parietal/irrigación sanguínea , Lóbulo Parietal/enzimología , Lóbulo Parietal/patología , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Lóbulo Temporal/irrigación sanguínea , Lóbulo Temporal/enzimología , Lóbulo Temporal/patologíaRESUMEN
Histone deacetylases (HDACs) constitute a super-family of enzymes grouped into four major classes (Class I-IV) that deacetylate histone tails leading to chromatin condensation and gene repression. Whether stroke-induced oligodendrogenesis is related to the expression of individual HDACs in the oligodendrocyte lineage has not been investigated. We found that 2 days after stroke, oligodendrocyte progenitor cells (OPCs) and mature oligodendrocytes (OLGs) were substantially reduced in the peri-infarct corpus callosum, whereas at 7 days after stroke, a robust increase in OPCs and OLGs was observed. Ischemic brains isolated from rats sacrificed 7 days after stroke were used to test levels of individual members of Class I (1 and 2) and Class II (4 and 5) HDACs in white matter oligodendrocytes during stroke-induced oligodendrogenesis. Double immunohistochemistry analysis revealed that stroke substantially increased the number of NG2+OPCs with nuclear HDAC1 and HDAC2 immunoreactivity and cytoplasmic HDAC4 which were associated with augmentation of proliferating OPCs, as determined by BrdU and Ki67 double reactive cells after stroke. A decrease in HDAC1 and an increase in HDAC2 immunoreactivity were detected in mature adenomatous polyposis coli (APC) positive OLGs, which paralleled an increase in newly generated BrdU positive OLGs in the peri-infarct corpus callosum. Concurrently, stroke substantially decreased the acetylation levels of histones H3 and H4 in both OPCs and OLGs. Taken together, these findings demonstrate that stroke induces distinct profiles of Class I and Class II HDACs in white matter OPCs and OLGs, suggesting that the individual members of Class I and II HDACs play divergent roles in the regulation of OPC proliferation and differentiation during brain repair after stroke.
Asunto(s)
Histona Desacetilasas/biosíntesis , Histona Desacetilasas/genética , Oligodendroglía/enzimología , Accidente Cerebrovascular/enzimología , Sustancia Blanca/enzimología , Acetilación , Animales , Diferenciación Celular , Proliferación Celular , Infarto Cerebral/enzimología , Infarto Cerebral/patología , Regulación Enzimológica de la Expresión Génica/genética , Histonas/metabolismo , Infarto de la Arteria Cerebral Media/patología , Masculino , Ratas , Ratas Wistar , Células Madre/enzimologíaRESUMEN
Recently, it has become widely known that rehabilitative training after stroke brings about some improvement of paralysis and disability; however, not much is known about the relationship between paralysis recovery and the participation of plasticity-related molecules. Hence, the localization and level of expression of several proteins in the cerebral cortex of rat groups with/without voluntary exercise using a running wheel after photo thrombotic infarction were examined in this study. In behavioral evaluation, the mean latency until falling from a rotating rod in the group with voluntary exercise at 6 days after infarction was significantly longer than that in the group without exercise. Immunohistochemical localization of c-Fos protein after behavioral test occurred in the area surrounding the infarction core in the exercise group. In protein expression analysis, protein kinase C (PKC), growth-associated protein 43 (GAP43) and phosphorylated at serine 41 GAP43 (p-GAP43) were significantly increased after voluntary exercise compared with those in rats without exercise. Expression of PKC immunoreactivity was observed in layer III of the perilesional cortex in rats with exercise, and the intracellular localization in the pyramidal neurons was mainly translocated to the plasma membrane. The expression and localization of these proteins may be related to the underlying mechanisms of exercise-induced paralysis recovery, that is, neuronal plasticity and remodeling of cortical connections through the phosphorylation of GAP43 by interaction with PKC. In the present study, the participation of at least some of the modulators associated with the improvement of motor deficit adjacent to the brain lesion might have been detected.
Asunto(s)
Infarto Cerebral/enzimología , Infarto Cerebral/rehabilitación , Condicionamiento Físico Animal , Proteína Quinasa C/metabolismo , Recuperación de la Función/fisiología , Animales , Western Blotting , Corteza Cerebral/enzimología , Inmunohistoquímica , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: Mutations in the high-temperature requirement A serine peptidase 1 (HTRA1) gene were studied in a Chinese family with cerebral autosomal recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL). METHODS: Exons 1-9 of the HTRA1 gene were amplified and bidirectionally sequenced in a Chinese family with CARASIL. Mutation effects were analysed by three-dimensional modelling of the serine protease HTRA1 protein. RESULTS: The proband was found to be homozygous for a novel missense mutation (c.854 C > T) identified in exon 4 of the HTRA1 gene; the parents of the proband were heterozygous for the same missense mutation. This c.854 C > T mutation resulted in a change from proline to leucine (p.P285L) in serine protease HTRA1, and was absent in 260 control chromosomes. Three-dimensional models showed that the change from proline to leucine (p.P285L) could attenuate the hydrogen bond between S284 and S287 residues, which might affect function of serine protease HTRA1. CONCLUSION: Discovery of a novel missense mutation (c.854C>T) associated with CARASIL expands the known CARASIL-related mutations in HTRA1.
Asunto(s)
Alopecia/genética , Infarto Cerebral/genética , Leucoencefalopatías/genética , Modelos Moleculares , Mutación Missense , Serina Endopeptidasas/genética , Enfermedades de la Columna Vertebral/genética , Adulto , Alopecia/enzimología , Alopecia/etnología , Alopecia/patología , Sustitución de Aminoácidos , Pueblo Asiatico , Secuencia de Bases , Estudios de Casos y Controles , Infarto Cerebral/enzimología , Infarto Cerebral/etnología , Infarto Cerebral/patología , Análisis Mutacional de ADN , Exones , Familia , Femenino , Heterocigoto , Serina Peptidasa A1 que Requiere Temperaturas Altas , Homocigoto , Humanos , Enlace de Hidrógeno , Leucoencefalopatías/enzimología , Leucoencefalopatías/etnología , Leucoencefalopatías/patología , Masculino , Datos de Secuencia Molecular , Linaje , Serina Endopeptidasas/química , Serina Endopeptidasas/metabolismo , Enfermedades de la Columna Vertebral/enzimología , Enfermedades de la Columna Vertebral/etnología , Enfermedades de la Columna Vertebral/patologíaRESUMEN
BACKGROUND: Matrix metalloproteinases (MMPs) are key players in proteolytic blood-brain barrier (BBB) disruption during ischemic stroke, leading to vascular edema, hemorrhagic transformation and infiltration by leukocytes. Their effect is dampened by the endogenous tissue inhibitors of metalloproteinases (TIMPs). The respective cellular source of specific MMPs and TIMPs during BBB breakdown is still under investigation. METHODS: We analyzed the MMP and TIMP release of human brain microvascular endothelial cells (BMECs) under oxygen glucose deprivation (OGD). Cultured human BMECs (the hCMEC/D3 cell line) were subjected to OGD (6, 12, 18 and 24 h). Gene expression of MMP-2, MMP-9, TIMP-1 and TIMP-2 were serially measured by quantitative real time-PCR and compared to ELISA-detected cell culture medium levels. RESULTS: OGD induced a significant and long-lasting increase in MMP-2 gene expression, reaching a plateau after 12 h. Medium protein levels of MMP-2 were correspondingly elevated at 12 h of OGD. The MMP-9 synthesis rate was detectable at very low levels and remained unaffected by OGD. TIMP-1 gene expression and secretion declined under OGD, whereas both expression and secretion of TIMP-2 remained stable. Contrary to the respective gene expression rate, medium levels of MMP-2, TIMP-1 and TIMP-2 started a simultaneous decline after 12 h of OGD. This is most likely due to an impaired synthesis and enhanced consumption rate under OGD. CONCLUSIONS: The objective of our study was to determine the contribution of human BMECs to the MMP metabolism under in vitro OGD conditions simulating ischemic stroke. Our results suggest that human BMECs switch to a proinflammatory state by means of an enhanced production of MMP-2, attenuated release of TIMP-1, and unaffected production of TIMP-2. Thus, human BMECs might participate in the MMP-mediated BBB breakdown during ischemic stroke. However, our data does not support human BMECs to be a source of MMP-9.
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Isquemia Encefálica/enzimología , Células Endoteliales/enzimología , Metaloproteinasa 2 de la Matriz/metabolismo , Inhibidor Tisular de Metaloproteinasa-1/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Barrera Hematoencefálica/metabolismo , Células Cultivadas , Infarto Cerebral/enzimología , HumanosRESUMEN
We made a case-control study to investigate a possible association between ALOX5AP-SG13S114A/T, COX-2-765G/C, and COX-1-50C/T polymorphisms with cerebral infarction in a Chinese population. A total of 411 cases with cerebral infarction were included; 411 controls matched for age, gender, and risk factors were also selected. The ALOX5AP-SG13S114A/T (rs10507391), COX-2-765G/C (rs20417), and COX-1-50C/T (rs3842787) polymorphisms were determined using PCR-RFLP. The generalized multifactor dimensionality reduction method was employed to detect gene-gene interactions. Based on single-gene analysis, there were no significant differences in the genotype and allele frequency distributions of ALOX5AP-SG13S114A/T, COX-2-765G/C, and COX-1-50C/T between the cerebral infarction group and controls. However, in those cases carrying ALOX5AP-SG13S114AA as well as COX-2-765CC, the risk of cerebral infarction increased significantly by 2.84 times (95%CI = 1.324-6.543). The single-gene ALOX5AP-SG13S114A/T, COX-2-765G/C, and COX-1-50C/T polymorphisms appear not to be associated with the development of cerebral infarction in Chinese populations. However, the interaction between ALOX5AP-SG13S114AA and COX-2-765CC apparently increases susceptibility to cerebral infarction.
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Proteínas Activadoras de la 5-Lipooxigenasa/genética , Pueblo Asiatico/genética , Infarto Cerebral/genética , Ciclooxigenasa 2/genética , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Polimorfismo de Nucleótido Simple/genética , Anciano , Estudios de Casos y Controles , Infarto Cerebral/enzimología , China , Ciclooxigenasa 1/genética , Epistasis Genética , Femenino , Frecuencia de los Genes/genética , Humanos , Modelos Logísticos , Masculino , Modelos Genéticos , Reducción de Dimensionalidad Multifactorial , Reproducibilidad de los ResultadosRESUMEN
We previously found that xanthotoxol, one of the major active ingredients in Cnidium monnieri (L.) Cusson, exerts protective effects in a rat model of focal cerebral ischemia/reperfusion injury by alleviating brain edema, inhibiting the neutrophil infiltration, and decreasing the expression of intercellular adhesion molecule-1 (ICAM-1) and E-selectin. The present study was designed to further determine the possible mechanisms of action of neuroprotective properties of xanthotoxol after cerebral ischemia. Transient focal cerebral ischemia/reperfusion model in male Sprague-Dawley rats was induced by 2-h middle cerebral artery occlusion followed by 24-h reperfusion. Xanthotoxol (5 and 10 mg/kg) or vehicle were administered intraperitoneally at 1 and 12 h after the onset of ischemia. At 24 h after reperfusion, we assessed the effect of xanthotoxol on the blood-brain barrier (BBB) permeability, the production of pro-inflammatory mediators such as interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, IL-8, nitric oxide (NO), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and the p65 subunit of the transcription factor, nuclear factor-κB (NF-κB) in the cortex after ischemic insult. The results showed that xanthotoxol treatment significantly attenuated BBB disruption, reduced the IL-1ß, TNF-α, IL-8 and NO level, and attenuated the iNOS activity compared with vehicle-treated animals. Further, xanthotoxol treatment also significantly prevented the ischemia/reperfusion-induced increase in the protein expression of iNOS, COX-2, and the nuclear NF-κB p65. These results, taken together with those of our previous study, suggest that the neuroprotection may be attributed to the ability of xanthotoxol to attenuate the expression of pro-inflammatory mediators and thereby inhibit the inflammatory response after cerebral ischemia.
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Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/patología , Furocumarinas/uso terapéutico , Inflamación/patología , Fármacos Neuroprotectores/uso terapéutico , Animales , Barrera Hematoencefálica/efectos de los fármacos , Barrera Hematoencefálica/patología , Edema Encefálico/complicaciones , Edema Encefálico/tratamiento farmacológico , Edema Encefálico/enzimología , Edema Encefálico/patología , Isquemia Encefálica/complicaciones , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/enzimología , Corteza Cerebral/patología , Infarto Cerebral/complicaciones , Infarto Cerebral/tratamiento farmacológico , Infarto Cerebral/enzimología , Infarto Cerebral/patología , Ciclooxigenasa 2/metabolismo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Furocumarinas/farmacología , Inflamación/complicaciones , Masculino , Fármacos Neuroprotectores/farmacología , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Permeabilidad/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Daño por Reperfusión/complicaciones , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/enzimología , Daño por Reperfusión/patología , Factor de Transcripción ReIA/metabolismoRESUMEN
The type 2 cannabinoid receptor (CB2R) was recently shown to mediate neuroprotection in ischemic injury. However, the role of CB2Rs in the central nervous system, especially neuronal and glial CB2Rs in the cortex, remains unclear. We, therefore, investigated anti-ischemic mechanisms of cortical CB2R activation in various ischemic models. In rat cortical neurons/glia mixed cultures, a CB2R agonist, trans-caryophyllene (TC), decreased neuronal injury and mitochondrial depolarization caused by oxygen-glucose deprivation/re-oxygenation (OGD/R); these effects were reversed by the selective CB2R antagonist, AM630, but not by a type 1 cannabinoid receptor antagonist, AM251. Although it lacked free radical scavenging and antioxidant enzyme induction activities, TC reduced OGD/R-evoked mitochondrial dysfunction and intracellular oxidative stress. Western blot analysis demonstrated that TC enhanced phosphorylation of AMP-activated protein kinase (AMPK) and cAMP responsive element-binding protein (CREB), and increased expression of the CREB target gene product, brain-derived neurotrophic factor. However, TC failed to alter the activity of either Akt or extracellular signal-regulated kinase, two major CB2R signaling pathways. Selective AMPK and CREB inhibitors abolished the neuroprotective effects of TC. In rats, post-ischemic treatment with TC decreased cerebral infarct size and edema, and increased phosphorylated CREB and brain-derived neurotrophic factor expression in neurons. All protective effects of TC were reversed by co-administration with AM630. Collectively, these data demonstrate that cortical CB2R activation by TC ameliorates ischemic injury, potentially through modulation of AMPK/CREB signaling, and suggest that cortical CB2Rs might serve as a putative therapeutic target for cerebral ischemia.
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Proteínas Quinasas Activadas por AMP/metabolismo , Lesiones Encefálicas/enzimología , Lesiones Encefálicas/patología , Isquemia Encefálica/patología , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Receptor Cannabinoide CB2/metabolismo , Transducción de Señal , Proteínas Quinasas Activadas por AMP/antagonistas & inhibidores , Animales , Lesiones Encefálicas/tratamiento farmacológico , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/enzimología , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Catalasa/metabolismo , Muerte Celular/efectos de los fármacos , Corteza Cerebral/patología , Infarto Cerebral/enzimología , Infarto Cerebral/patología , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/antagonistas & inhibidores , Depuradores de Radicales Libres/metabolismo , Glucosa/deficiencia , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Oxígeno/farmacología , Fosforilación/efectos de los fármacos , Sesquiterpenos Policíclicos , Ratas , Ratas Sprague-Dawley , Receptor Cannabinoide CB2/agonistas , Sesquiterpenos/farmacología , Sesquiterpenos/uso terapéutico , Transducción de Señal/efectos de los fármacos , Superóxido Dismutasa/metabolismoRESUMEN
Because arginase and nitric oxide (NO) synthases (NOS) compete to degrade l-arginine, arginase plays a crucial role in the modulation of NO production. Moreover, the arginase 1 isoform is a marker of M2 phenotype macrophages that play a key role in tissue remodeling and resolution of inflammation. While NO has been extensively investigated in ischemic stroke, the effect of stroke on the arginase pathway is unknown. The present study focuses on arginase expression/activity and localization before and after (1, 8, 15 and 30 days) the photothrombotic ischemic stroke model. This model results in a cortical lesion that reaches maximal volume at day 1 post-stroke and then decreases as a result of astrocytic scar formation. Before stroke, arginase 1 and 2 expressions were restricted to neurons. Stroke resulted in up-regulation of arginase 1 and increased arginase activity in the region centered on the lesion where inflammatory cells are present. These changes were associated with an early and long-lasting arginase 1 up-regulation in activated macrophages and astrocytes and a delayed arginase 1 down-regulation in neurons at the vicinity of the lesion. A linear positive correlation was observed between expressions of arginase 1 and glial fibrillary acidic protein as a marker of activated astrocytes. Moreover, the pattern of arginase 1 and brain-derived neurotrophic factor (BDNF) expressions in activated astrocytes was similar. Unlike arginase 1, arginase 2 expression was not changed by stroke. In conclusion, increased arginase 1 expression is not restricted to macrophages in inflammation elicited by stroke but also occurs in activated astrocytes where it may contribute to neuroplasticity through the control of BDNF production.
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Arginasa/metabolismo , Encéfalo/enzimología , Accidente Cerebrovascular/enzimología , Animales , Arginasa/genética , Astrocitos/metabolismo , Encéfalo/metabolismo , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Infarto Cerebral/enzimología , Infarto Cerebral/metabolismo , Expresión Génica , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Macrófagos/metabolismo , Masculino , Neuronas/metabolismo , Ratas , Ratas Wistar , Accidente Cerebrovascular/metabolismoRESUMEN
Perinatal hypoxic ischemic (HI) brain injury is a leading cause of long-term neurological handicap in newborn babies. Recently, excessive activity of matrix metalloproteinases (MMPs), and in particular MMP-9, has been implicated in the aetiology of HI injuries to the immature brain. Our previous study suggested that MMP-9 may be involved in the development of the delayed injury processes following HI injury to the developing brain. Given this, we therefore propose that MMP-9 may be a useful target for rescue therapies in the injured developing brain. To address this, we chose to use SB-3CT, a highly selective inhibitor that is known to target only MMP-2 and MMP-9, to attenuate the elevated MMP-9 activity seen following HI injury to the developing brain. Twenty-one-day-old postnatal Wistar rats were subjected to unilateral carotid artery occlusion followed by exposure to hypoxia (8% oxygen for 1 h). SB-3CT (50 mg/kg body weight in 25% dimethyl sulphoxide/75% polyethylene glycol) or an equal volume of vehicle or saline diluent was then administered intraperitoneally at 2, 5 and 14 h following the insult. Gelatin zymography revealed that pro-MMP-9 levels were significantly reduced at 6 h following hypoxic ischaemia (p ≤ 0.05). However, our results showed that despite significantly inhibiting brain pro-MMP-9 activity after hypoxic ischaemia, SB-3CT failed to confer significant neuroprotection in postnatal day 21 rats 3 days after an HI insult. Further investigations are warranted using a recently reported selective water-soluble version of SB-3CT or another MMP-9 selective inhibitor to resolve the role of MMP-9 in the aetiology of HI injury in the developing brain.
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Encéfalo/crecimiento & desarrollo , Inhibidores Enzimáticos/farmacología , Compuestos Heterocíclicos con 1 Anillo/farmacología , Hipoxia-Isquemia Encefálica/enzimología , Metaloproteinasa 9 de la Matriz/efectos de los fármacos , Sulfonas/farmacología , Animales , Conducta Animal/efectos de los fármacos , Peso Corporal/efectos de los fármacos , Infarto Cerebral/tratamiento farmacológico , Infarto Cerebral/enzimología , Cromatografía Líquida de Alta Presión , Inhibidores Enzimáticos/farmacocinética , Compuestos Heterocíclicos con 1 Anillo/farmacocinética , Hipoxia-Isquemia Encefálica/complicaciones , Metaloproteinasa 2 de la Matriz/metabolismo , Proteínas del Tejido Nervioso/biosíntesis , Enfermedades del Sistema Nervioso/etiología , Enfermedades del Sistema Nervioso/prevención & control , Ratas , Ratas Wistar , Sulfonas/farmacocinética , Espectrometría de Masas en TándemRESUMEN
Cytochrome P450 epoxygenase metabolites of arachidonic acid, EETs, have multiple cardiovascular effects, including reduction of blood pressure, protection against myocardial ischemia-reperfusion injury, and attenuation of endothelial apoptosis. This study investigated the hypothesis that transgenic mice with endothelial overexpression of CYP2J2 (Tie2-CYP2J2-Tr) would be protected against global cerebral ischemia induced by bilateral common carotid artery occlusion (BCCAO) and action mechanisms of EETs on cerebral ischemia in cultures of astrocytes exposed to oxygen-glucose deprivation (OGD). Tie2-CYP2J2-Tr mice had significantly increased CYP2J2 expression, increased 14,15-EET production, increases regional cerebral blood flow (rCBF) and microvascular density, decreased ROS production, decreased brain infarct size and apoptosis after ischemia compared to wild type mice, these were associated with increased activation of the PI3K/AKT and apoptosis-related protein in ischemic brain. Addition of exogenous EETs or CYP2J2 transfection attenuated OGD-induced apoptosis in astrocytes via activation of PI3K/AKT and anti-apoptosis pathways. However, these effects were reduced by pretreatments with inhibitor of the PI3K (LY294002) and 14,15-EET (14,15-EEZE), respectively. These results indicate that CYP2J2 overexpression exerts marked neuroprotective effects against ischemic injury by a mechanism linked to increased level of circulating EETs and increases CBF and reduction of apoptosis.
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Isquemia Encefálica/enzimología , Infarto Cerebral/enzimología , Oclusión Coronaria/enzimología , Sistema Enzimático del Citocromo P-450/genética , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido 8,11,14-Eicosatrienoico/metabolismo , Ácido 8,11,14-Eicosatrienoico/farmacología , Animales , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/antagonistas & inhibidores , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Astrocitos/citología , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Isquemia Encefálica/genética , Isquemia Encefálica/fisiopatología , Hipoxia de la Célula , Células Cultivadas , Infarto Cerebral/genética , Infarto Cerebral/fisiopatología , Circulación Cerebrovascular , Oclusión Coronaria/genética , Oclusión Coronaria/fisiopatología , Citocromo P-450 CYP2J2 , Sistema Enzimático del Citocromo P-450/metabolismo , Expresión Génica/efectos de los fármacos , Glucosa/deficiencia , Masculino , Ratones , Ratones Transgénicos , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
OBJECTIVE: To observe the effect of electroacupunctur (EA) of Shuigou(GV 26)on the expression of vascular protein kinase C (PKC) in the focal ischemia cerebral tissue in rats so as to study its dynamic regulation mechanism. METHODS: A total of 78 Wistar rats were randomly divided into model group (n = 24), EA group (n = 24), sham operation (sham) group (n = 24) and control group (n = 6). The first 3 groups were further divided into 0. 5 h, 1 h, 3 h and 6 h subgroups, with 6 cases in each. Cerebral ischemia (CI) model was established by occlusion of the middle cerebral artery. EA (15 Hz, 1.0 mA) was applied to "Shuigou" (GV 26) for 20 min. The anterior, middle and posterior cerebral arteries on the infarct side of the brain were collected for assaying the expression of PKC protein by using Western blot, and the formalin-fixed artery samples were embedded with paraffin, followed by sectioning and staining with immunohistochemistry. RESULTS: Compared with the control group, the expression levels of cerebral vascular smooth PKC protein shown by both immunohistochemistry and Western blot were upregulated significantly at the time-points of 0.5 h, 1 h, 3 h and 6 h after CI in the model group (P<0.05, P<0.01). In comparison with the model group, the expression levels of PKC protein in the cerebral vascular smooth muscle shown by the aforementioned two methods were down-regulated considerably in the EA group (P<0.01, P<0.05). No statistic differences were found between the control and sham groups in the expression of vascular PKC protein at different time-points (P>0.05). CONCLUSION: EA Intervention can significantly inhibit PKC expression in the cerebral vascular smooth muscle in CI rats, which may be responsible for its efficacy in improving ischemic stroke.