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  • The transport of glutamate by


    The transport of glutamate by EAAT2 from the extracellular fluid into either astrocytes or endothelial cells is an unfavorable and energy-consuming process. This energy is provided by a coupled co-transport of three sodium ions, one proton, and one glutamate molecule in the counter-transport of one potassium ion. Notably, the transporters also function as anion-selective channels [4]. The critical role of EAAT2 in controlling brain glutamate homeostasis has led to the development of different therapeutic strategies to reduce the excitotoxic damage by glutamate after stroke. β-lactam ibuprofen msds such as ceftriaxone have been described to be transcriptional activators of EAAT2. Due to the increase in EAAT2 gene expression and transport activity, treatment with antibiotics results in facilitated glutamate uptake by astroglial cells and thus neuronal protection against ischemic insult [3,5]. The brain-to-blood glutamate efflux mechanism mediated by endothelial EAAT2 has also permitted the development of a new generation of protective drugs against ischemic glutamate toxicity, namely, blood glutamate-grabbers. These blood glutamate-grabbers can metabolize and thus reduce the glutamate concentration in the blood. This leads to a larger glutamate gradient between the brain and blood, facilitating the efflux of extracellular brain glutamate via endothelial cells into the blood. In the blood, glutamate is metabolized by the activity of glutamate oxaloacetate transaminase 1 (GOT1), which catalyzes the transformation of oxaloacetate and glutamate into aspartate and α-ketoglutarate. Thus, the administration of both oxaloacetate and/or recombinant GOT1 (rGOT1) in ischemic animal models reduces glutamate in both the blood and the brain, which improves functional recovery after an ischemic lesion [9,[12], [13], [14], [15], [16], [17]]. The protective efficacy of this strategy has been shown in different types of ischemic animal models and has been validated in humans by pharmacological [18] and non-pharmacological approaches such as peritoneal dialysis [19]. It has also been tested in other pathological models associated with an increase in glutamate in the brain, such as traumatic brain injury [20], subarachnoid hemorrhage [21], glioma [22], amyotrophic lateral sclerosis [23], or Alzheimer's disease [24], with successful results. On the basis of the substantial increase in glutamate uptake in astrocytes by the overexpression of EAAT2 and the promising efficacy of the brain-to-blood glutamate efflux mechanism [3,9], we hypothesized that combining EAAT2 expression in therapeutic cells for systemic administration might accomplish an alternative cell-based glutamate-grabbing therapy, i.e., by combining certain intrinsic cell properties with excitotoxic protection. Therefore, EAAT2-encoding cDNA was introduced and functionally expressed in mesenchymal stem cells (MSCs), which are among the best candidates for stem cell therapy for ischemic stroke owing to their ability to release growth factors, as well as their immunomodulatory capacities [25]. For independent control experiments, EAAT2 was introduced into human embryonic kidney 293 (HEK 293) cells, which do not interfere with ischemic damage per se. The expression and functionality of EAAT2 in both cell types were evaluated by several in vitro assays, and the blood glutamate-grabbing activity was tested in ischemic animals and compared with that resulting from oxaloacetate treatment.
    Experimental procedures
    Discussion Blood/brain glutamate-grabbing is an emerging protective strategy to reduce the ibuprofen msds excitotoxic effect of the elevated extracellular glutamate that accumulates in the brain during the acute phase of ischemic stroke [9,18]. This novel protective strategy has been tested by several laboratories in different ischemic animal models [13,16,[35], [36], [37], [38]], using the two most common pharmacological approaches, i.e., oxaloacetate and rGOT1 application. The protective mechanism has also been validated by non-pharmacological approaches like peritoneal dialysis, used to filter the blood and remove excess glutamate [19,39,40]. All these studies imply that blood glutamate reduction holds great potential as an effective protective strategy to reduce the neurotoxic increase in glutamate in the brain. On the basis of the high specificity of excitatory amino acid transporters, e.g., EAAT2, for glutamate uptake, we developed a novel cell-based blood/glutamate-scavenging approach by introducing the coding sequence of EAAT2 into MSCs, combining the intrinsic properties of these cells with excitotoxic protection. Here, we compared the blood glutamate-grabbing efficacy of this cell-based strategy with the “gold standard,” i.e., oxaloacetate treatment.