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Objective: To investigate recombinant human TIMP-1 (125I-rhTIMP-1) half-life in blood and its distribution in rat brain tissue after cerebral ischemia/reperfusion as part of a therapeutic development paradigm. Method: A suture model of the middle cerebral artery occlusion was used. 125I-labeled rhTIMP-1 at 60 μg/kg (11·23 μCi/μg) was administered to rats intravenously at the beginning of reperfusion. Blood and brain tissue were collected. The radioactivity was detected with a gamma counter and analyzed by autoradiography. Results: The blood half-life T1/2 of 125I-rhTIMP-1 was 42·2 hours. Thirty minutes after 125I-rhTIMP-1 administration, an increased accumulation of 125I-rhTIMP-1 in the ischemic hemisphere was observed. The maximum brain tissue concentration Cmax was 26·1 ng/g at 1·5 hours in the striatum and 13·9 ng/g at 5 hours in the cortex when the uptake percentage of brain tissue to blood was 6·1±0·4 and 6·7±2·1%, respectively. The cortex and striatum elimination half-lives T1/2 were 45·3 and 39·2 hours, respectively. Electrophoretic analysis of ischemic samples for 125I-rhTIMP-1 showed a clear 28 kDa band 1·5 hours after 125I-rhTIMP-1 administration in the cortex and striatum. The intensity of the 28 kDa band decreased after 3·0 hours of the administration. Some 125I-rhTIMP-1 maintained its molecular integrity for 8·5 hours in ischemic striatum after reperfusion. Discussion: 125I-labeled rhTIMP-1 was distributed quickly into ischemic brain tissue and had a slow elimination in both blood and brain tissue. These results, along with other studies suggesting therapeutic benefits, will aid in the development of TIMP-1 for protecting ischemic stroke. [ABSTRACT FROM AUTHOR]

Objective: The discovery of IL-7Rα polymorphisms implicated in the pathogenesis of multiple sclerosis has highlighted the importance of interleukin 7 (IL-7) in central nervous system diseases. Hypoxia affects neurological disease states in part by modulating expression of many early and late response genes. The present work used cultured PC12 cells to investigate the effect of hypoxia on IL-7 expression. Method: PC12 cells were cultured in Dulbecco's modified Eagle's medium (DMEM)/F12 medium. RNA was isolated and reverse transcriptase–polymerase chain reaction (RT-PCR) was run to quantify messenger RNA (mRNA) change. Western blots were used to assess IL-7 protein change in the medium. Extracellular free Ca2+ was removed by using Ca2+-free DMEM/F12 with 1 mM ethylene glycol tetraacetic acid for 45 minutes before the start of hypoxia. Results: Exposure of PC12 cells to 1% oxygen for 6 hours decreased IL-7 mRNA by 77% using RT-PCR (p<0.01). Exposure to 1% oxygen for 24 hours decreased IL-7 protein in the medium by 21% (p<0.05). As hypoxia duration increased (2, 4, 6 and 24 hours) or oxygen concentrations decreased (10%, 5% and 1%), IL-7 mRNA expression progressively decreased. Removal of extracellular free Ca2+ completely prevented these hypoxia-induced decreases of IL-7 mRNA. Discussion: Since IL-7 exhibits trophic properties in developing brain, down-regulation of IL-7 by hypoxia may contribute to hypoxia-induced injury to neural cells. [ABSTRACT FROM AUTHOR]

Objective: Worldwide, cerebral vasospasm after subarachnoid hemorrhage (SAH) has an estimated morbidity and mortality of 1.2 million annually. While it has long been suspected that reactive oxygen species play a major role in the etiology of cerebral vasospasm after SAH, promising results in animal work were not borne out in human clinical trials, despite intensive research effort. The purpose of this study is to investigate the role of glutathione peroxidase in the SAH cerebrospinal fluid milieu. Methods: We utilized commercially available kits for the quantitation of glutathione peroxidase 1 (glutathione peroxidase) activity and oxygen radical capacity and sodium dodecyl sulfate polyacrylamide gel electrophoresis with Western blotting with specific antibodies to human glutathione peroxidase to determine the enzyme content of the cerebrospinal fluid samples. Human cerebrospinal fluid was obtained in an Institutional Review Board-exempt manner for this study in the following groups: control (no SAH), CSFC (SAH but no vasospasm on angiography) and CSFV (SAH with clinical and angiographic vasospasm). Results: We found that glutathione peroxidase activity is significantly higher in CSFV compared with CSFC, and this is reflected in a higher total oxidative capacity in CSFV. Despite similar levels of glutathione peroxidase protein, CSFV had significantly greater activity than CSFC. Discussion: These results further elucidate previous research from this laboratory, showing increased oxidative stress in CSFV compared with CSFC. In conclusion, there appears to be increased glutathione peroxidase activity in CSFV, despite there being increased levels of oxidative stress markers, suggesting overwhelming oxidative stress may play a role in cerebral vasospasm after SAH. [ABSTRACT FROM AUTHOR]

Objective: Recanalization therapies for ischemic stroke have been slow to change clinical practice because of perceived and published risks of hemorrhage associated with lytic administration. We quantified alfimeprase in an acute ischemia–reperfusion model, as compared with recombinant tissue plasminogen activator, with hemorrhagic transformation as the primary endpoint and infarction volume and blood–brain barrier permeability as secondary endpoints. Methods: Five groups were studied in a blinded fashion: alfimeprase at doses of 0.03 (n=8), 0.1 (n=11) and 0.3 mg/kg (n=8); recombinant tissue plasminogen activator at 1 mg/kg (n=9); carrier infused controls (n=9). The middle cerebral artery was occluded for 5 hours followed by removal of the suture for reperfusion. Drugs were infused immediately following reperfusion over a 10-minute period. Approximately 24 hours later, the animals were anesthetized and decapitated, and the brains were rapidly harvested and frozen. Serial brain sections were obtained and inspected for hemorrhages. Infarction and blood–brain barrier permeability were also evaluated in additional experiments in control, 0.1 mg/kg alfimeprase and 1 mg/kg recombinant tissue plasminogen activator-treated rats. Results: The hemorrhagic transformation frequency, neurological deficit and the mortality rate of alfimeprase were significantly lower than for recombinant tissue plasminogen activator at the 0.03 mg/kg dose and not statistically different at the higher doses. Infarction and blood–brain barrier permeability were not significantly different among control, 0.1 mg/kg alfimeprase and recombinant tissue plasminogen activator. Discussion: In this model, alfimeprase, a new fibrinolytic agent, exhibits a profile comparable to recombinant tissue plasminogen activator. [ABSTRACT FROM AUTHOR]

Summary: N-acetylaspartate (NAA) is an intermediary metabolite that is found in relatively high concentrations in the human brain. More specifically, NAA is so concentrated in the neurons that it generates one of the most visible peaks in nuclear magnetic resonance (NMR) spectra, thus allowing NAA to serve as “a neuronal marker”. However, to date there is no generally accepted physiological (primary) role for NAA. Another molecule that is found at similar concentrations in the brain is glutamate. Glutamate is an amino acid and neurotransmitter with numerous functions in the brain. We propose that NAA, a six-carbon amino acid derivative, is converted to glutamate (five carbons) in an energetically favorable set of reactions. This set of reactions starts when aspartoacylase converts the six carbons of NAA to aspartate and acetate, which are subsequently converted to oxaloacetate and acetyl CoA, respectively. Aspartylacylase is found in astrocytes and oligodendrocytes. In the mitochondria, oxaloacetate and acetyl CoA are combined to form citrate. Requiring two steps, the citrate is oxidized in the Kreb’s cycle to α-ketoglutarate, producing NADH. Finally, α-ketoglutarate is readily converted to glutamate by transaminating the α-keto to an amine. The resulting glutamate can be used by multiple cells types to provide optimal brain functional and structural needs. Thus, the abundant NAA in neuronal tissue can serve as a large reservoir for replenishing glutamate in times of rapid or dynamic signaling demands and stress. This is beneficial in that proper levels of glutamate serve critical functions for neurons, astrocytes, and oligodendrocytes including their survival. In conclusion, we hypothesize that NAA conversion to glutamate is a logical and favorable use of this highly concentrated metabolite. It is important for normal brain function because of the brain’s relatively unique metabolic demands and metabolite fluxes. Knowing that NAA is converted to glutamate will be important for better understanding myriad neurodegenerative diseases such as Canavan’s Disease and Multiple Sclerosis, to name a few. Future studies to demonstrate the chemical, metabolic and pathological links between NAA and glutamate will support this hypothesis. [Copyright &y& Elsevier]