That MSCs release IL upon LPS stimulation or other stressful

That MSCs release IL-6 upon LPS stimulation or other stressful stimuli like H2O2 [43] is well-documented [44]. However, there are conflicting reports on the potential effects of this cytokine in stroke. Some studies have found that the external administration of IL-6 in ischemic animal models leads to an improvement in the animal outcome [45], while we recently reported IL-6 to be associated with poor outcome in a study on 4112 stroke patients [46], a finding that is corroborated by the present findings. We cannot exclude that an alteration in the release of other mesenchymal factors or cytokines could contribute to reduce the beneficial effect of transfected MSCs in ischemic animals. From our study, we can deduce that the experimental manipulation of mesenchymal cells induces an alteration of their intrinsic properties and reduces their beneficial properties, which is an important issue for studies focusing on the therapeutic effects of MSCs. We previously evaluated the fate and biodistribution of MSCs after i.v. administration in the same ischemic animal model as we have used in this study [30]. We observed that the benefits observed after i.v. delivery of MSCs were not attributable to the cell engraftment in the p2y inhibitor as MSCs did not reach the brain tissue, and positively labeled cells were found to be distributed mainly along the lungs. Therefore, we can assume that the effect of MSCs on glutamate and infarct reduction was mediated by a systemic effect and not the engraftment and dedifferentiation into cerebral EAAT2-expressing cells, as astrocytes or endothelial cells. This study had some limitations that should be taken into consideration. One of the main limitations of protective drugs against glutamate excitotoxicity after stroke is the narrow therapeutic time window, mainly because the increase in glutamate release appears immediately after stroke; therefore, treatments against glutamate excitotoxicity should be administered as soon as possible but within a clinically relevant time window. In this study, cell infusions were applied upon reperfusion, only 75 min after occlusion onset. This could be considered an early treatment regimen, which might be impractical in the clinic setting. However, although the metabolic rate in rats is faster than that in humans (~6.4 times faster [47]), making the time window clinically acceptable, further studies should determine the time window of these cell therapies. In summary, our results show that cell-based therapies using the administration of EAAT2-expressing cells to ischemia-lesioned animal models is a novel alternative to reduce glutamate neuronal excitotoxicity. Although further studies are necessary to optimize the EAAT2 expression in mesenchymal cells, this study opens up avenues for combining a protective strategy with a cell therapy for cerebral ischemia.
Author contributions
Conflict of interest
Acknowledgement This study was partially supported by grants from Instituto de Salud Carlos III (PI13/00292 and PI17/0054), Spanish Research Network on Cerebrovascular Diseases RETICS-INVICTUS (RD12/0014), Fundación Mutua Madrileña; the Ministry of Economy and Competitiveness of Spain (SAF2014-56336-R), Xunta de Galicia (Programa de Desarrollo Precomercial de los resultados de investigación del Sistema Público de Salud de Galicia_ PRIS); and the European Union program FEDER. Furthermore, F. Campos (CP14/00154) and T. Sobrino (CP12/03121 and CPII17/00027) are recipients of a research contract from Miguel Servet Program of Instituto de Salud Carlos III. Funders had not any role in study design, data collection, data analysis, interpretation, writing of the report. We thank Prof. C. Fahlke (Institute of Complex Systems – Cellular Biophysics (ICS-4), Forschungszentrum Jülich, Germany) for assistance and for providing the pRcCMVmYFPEAAT2 plasmid.
Introduction Neurotransmitter transporters (NTTs) regulate neurotransmission by removing their respective substrates from the synaptic space into presynaptic elements or adjacent glial cells. There are two structurally distinct families of NTTs: the glutamate/neutral amino acid transporters (SLC1); and the neurotransmitter-sodium symporters (SLC6) (Torres and Amara, 2007). Both families differ radically in the mechanism of translocation of their substrates, although as secondary active transporters, they share some biophysical properties such as dependency on the electrochemical gradient of sodium. In addition, to a greater or lesser degree, both families have decoupled movement of charges, with a channel-like mechanism of operation (Divito et al., 2017; Sonders et al., 1997). Among the transporters of the SLC1 family, GLT-1 stands out for its abundance, and though it is expressed mainly in glial cells, neuronal forms have been identified that are estimated to represent 5–10% of total GLT-1 (Rimmele and Rosenberg, 2016). On the other hand, the SLC6 family includes transporters for GABA, glycine and biogenic amines, among others. One of the transporters of biogenic amines is the dopamine transporter (DAT), which is expressed exclusively in dopaminergic neurons whose cell bodies are in the midbrain with terminals mainly in the striatum, and more sparingly in the cortex and other areas of the brain. DAT has a key function in dopaminergic neurotransmission because the reuptake of dopamine by this high affinity transporter is crucial to determine the intensity and duration of dopamine signaling at synapses (Torres et al., 2003).