RESUMO
OBJECTIVES: Pathological increases in asymmetric dimethylarginine, an endogenous nitric oxide synthase inhibitor, have been implicated in endothelial dysfunction and vascular diseases. Reduced nitric oxide early after traumatic brain injury may contribute to hypoperfusion. Currently, methods to quantify asymmetric dimethylarginine in the cerebrospinal fluid have not been fully explored. We aimed to develop and validate a method to determine asymmetric dimethylarginine in the cerebrospinal fluid of a pediatric traumatic brain injury population and to use this method to assess the effects of 1) traumatic brain injury and 2) therapeutic hypothermia on this mediator. DESIGN, SETTING, AND PATIENTS: An ancillary study to a prospective, phase II randomized clinical trial of early hypothermia in a tertiary care pediatric intensive care unit for children with Traumatic brain injury admitted to Children's Hospital of Pittsburgh. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: A UPLC-MS/MS method was developed and validated to quantitate asymmetric dimethylarginine. A total of 56 samples collected over 3 days with injury onset were analyzed from the cerebrospinal fluid of consented therapeutic hypothermia (n = 9) and normothermia (n = 10) children. Children undergoing diagnostic lumbar puncture (n = 5) were enrolled as controls. Asymmetric dimethylarginine was present at a quantifiable level in all samples. Mean asymmetric dimethylarginine levels were significantly increased in normothermic Traumatic brain injury children compared with that in control (0.19 ± 0.08 µmol/L and 0.11 ± 0.02 µmol/L, respectively, p = 0.01), and hypothermic children had significantly reduced mean asymmetric dimethylarginine levels (0.11 ± 0.05 µmol/L) vs. normothermic (p = 0.03) measured on day 3. Patient demographics including age, gender, and nitric oxide levels (measured as nitrite and nitrate using liquid chromatography coupled with Griess reaction) did not significantly differ between normothermia and hypothermia groups. Also, nitric oxide levels did not correlate with asymmetric dimethylarginine concentrations. CONCLUSIONS: Asymmetric dimethylarginine levels were significantly increased in the cerebrospinal fluid of traumatic brain injury children. Early hypothermia attenuated this increase. The implications of attenuated asymmetric dimethylarginine on nitric oxide synthases activity and regional cerebral blood flow after traumatic brain injury by therapeutic hypothermia deserve future study.
Assuntos
Arginina/análogos & derivados , Lesões Encefálicas/líquido cefalorraquidiano , Lesões Encefálicas/terapia , Hipotermia Induzida , Adolescente , Arginina/líquido cefalorraquidiano , Estudos de Casos e Controles , Líquido Cefalorraquidiano/química , Criança , Pré-Escolar , Cromatografia Líquida , Feminino , Humanos , Lactente , Masculino , Nitratos/líquido cefalorraquidiano , Nitritos/líquido cefalorraquidiano , Espectrometria de Massas em TandemRESUMO
BACKGROUND: Endothelin-1 (ET-1) is a potent vasoconstrictor implicated in the pathogenesis of vasospasm and delayed cerebral ischemia (DCI) in aneurysmal subarachnoid hemorrhage (aSAH) patients. The aim of this study was to investigate the relationship between cerebrospinal fluid (CSF) ET-1 levels and angiographic vasospasm and DCI. METHODS: Patients with aSAH were consented (n = 106). Cerebral vasospasm was determined by angiography. DCI was determined by transcranial Doppler (TCD) results and/or angiogram results with corresponding clinical deterioration. CSF ET-1 levels over 14 days after the initial insult was quantified by ELISA. ET-1 analysis included a group-based trajectory analysis and ET-1 exposure rate during 24, 48, and 72 h prior to, as well as 72 h post angiography, or clinical deterioration. RESULTS: Trajectory analysis revealed two distinct groups of subjects with 56% of patients in the low ET-1 trajectory group (mean at day 1 = 0.31 pg/ml; SE = 0.04; mean at day 14 = 0.41 pg/ml; SE = 0.15) and 44% of patients in the high ET-1 trajectory group (mean at day 1 = 0.65 pg/ml; SE = 0.08; mean at day 14 = 0.61 pg/ml; SE = 0.06). Furthermore, we observed that ET-1 exposure rate 72 h before angiography and clinical spasm was a significant predictor of both angiographic vasospasm and DCI, whereas, ET-1 exposure after angiography and clinical spasm was not associated with either angiographic vasospasm or DCI. CONCLUSION: Based on these results we conclude that ET-1 concentrations are elevated in a sub-group of patients and that the acute (72 h prior to angiography and clinical neurological deterioration), but not chronic, elevations in CSF ET-1 concentrations are indicative of the pathogenic alterations of vasospasm and DCI in aSAH patients.
Assuntos
Isquemia Encefálica/líquido cefalorraquidiano , Endotelina-1/líquido cefalorraquidiano , Hemorragia Subaracnóidea/líquido cefalorraquidiano , Hemorragia Subaracnóidea/complicações , Vasoespasmo Intracraniano/líquido cefalorraquidiano , Adulto , Idoso , Isquemia Encefálica/diagnóstico , Isquemia Encefálica/etiologia , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Fatores Socioeconômicos , Fatores de Tempo , Vasoespasmo Intracraniano/diagnóstico , Vasoespasmo Intracraniano/etiologiaRESUMO
N-hydroxy-N'-(4-n-butyl-2-methylphenyl)formamidine (HET0016) is a potent inhibitor of 20-hydroxyeicosatetraenoic acid (20-HETE) formation by specific cytochrome P450 isoforms. Previous studies have demonstrated that administration of HET0016 inhibits brain formation of 20-HETE and reduces brain damage in a rat model of thromboembolic stroke. Delineation of the dose, concentration, and neuroprotective effect relationship of HET0016 has been hampered by the relative insolubility of HET0016 in aqueous solutions and the lack of information concerning the mechanism and duration of HET0016 inhibition of brain 20-HETE formation. Therefore, it was the purpose of this study to develop a water-soluble formulation of HET0016 suitable for intravenous (i.v.) administration and to determine the time course and mechanism of brain 20-HETE inhibition after in vivo dosing. In this study we report that HET0016 is a noncompetitive inhibitor of rat brain 20-HETE formation, which demonstrates a tissue concentration range for brain inhibition. In addition, we demonstrate that complexation of HET0016 with hydroxypropyl-beta-cyclodextrin results in increased aqueous solubility of HET0016 from 34.2 +/- 31.2 to 452.7 +/- 63.3 microg/ml. Administration of the complex as a single HET0016 i.v. dose (1 mg/kg) rapidly reduced rat brain 20-HETE concentrations from 289 to 91 pmol/g. Collectively, these data demonstrate that the i.v. formulation of HET0016 rapidly penetrates the rat brain and significantly inhibits 20-HETE tissue concentrations. These results will enable future studies to determine biopharmaceutics of HET0016 for inhibition of 20-HETE after cerebral ischemia.