From eight independent experiments and are expressed as fold changes reasonably to nontreated microglia.Differences among the three various groups at every time point have been obtained by oneway ANOVA BET-IN-1 Formula followed by Bonferroni posthoc correction.p .and p .vs.nontreated cells; ## p .vs.treatment with exosomes from wt NSC MNs.Frontiers in Neuroscience www.frontiersin.orgMay Volume ArticlePinto et al.MNMicroglia Exosomal Trafficking in ALSFIGURE Exosomes from NSC motor neurons (MNs) mutated in GA (mSOD) establish a sustained and marked decrease within the N microglia PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/21536721 phagocytic potential.N microglial cells have been incubated for , , and h with exosomes (Exos) from wildtype (wt) NSC MNs and mSOD NSC MNs (Nwt Exos and NmSOD Exos, respectively), as indicated in strategies.Nontreated cells had been considered as manage.(A) Representative final results of a single experiment, showing engulfed latex beads (in green) by the Iba stained (in red) microglia with nuclei labeled by Hoechst dye (in blue).(B) Results are expressed as percentage of cells, fairly to the total number of microglia, showing ingested beads.Benefits are imply (SEM) from eight independent experiments.Differences amongst the 3 distinctive groups at each time point were obtained by oneway ANOVA followed by Bonferroni posthoc correction.# p .vs.exosomes from wt MNs.Scale bar represents .upregulated and maintained until h interaction, differently from the above pointed out inflammatory mediators, in cells exposed to exosomes from mSOD NSC MNs, also disappearing just after h incubation (Figures F,G).Determined by these data we may assume that exosomes from the mSOD NSC MNs transiently switch N microgliainto a M polarized cell (Durafourt et al Chhor et al).Considering that early or late NFB activation was shown to induce different sets of genes, by respectively encoding TNF, IL, MMP, or cell surface receptors, adhesion molecules and signal adapters (Tian et al), we next evaluated the effects created on the expression of cell surface receptors.Frontiers in Neuroscience www.frontiersin.orgMay Volume ArticlePinto et al.MNMicroglia Exosomal Trafficking in ALSExosomes from mSOD NSC MNs Lead to a Delayed Upregulation of Receptors Involved in N Microglia Response to StimuliTo decide no matter if late NFB activation in microglia treated with mSOD exosomes was linked using the improved expression of membrane surface receptors, like TREM, RAGE, and TLR, we evaluated their gene expression levels within a timedependent manner.Indeed, microglia was shown to express many receptors able to effectively respond to external stimuli (Pocock and Kettenmann,).TREM receptor has been identified as a potential regulator on the microglial phenotype (Stefano et al) and found elevated in the spinal cord of ALS patients and SODGA mice (Cady et al).As depicted in Figure A, improved expression of TREM gene in N microglia was evident following h incubation with each wt NSC MNs and mSOD MNsderived exosomes, even though some fluctuations had been observed overtime.TREM overexpression has been associated with suppression of neuroinflammation and microglia M polarization connected with enhanced phagocytic ability (Painter et al Jiang et al).RAGE is also a receptor identified elevated in association with mSOD (Shibata et al).Within the present study, it can be clear its net elevation only in the N microglia treated for h with exosomes from mSOD MNs (Figure B, p .vs.wt NSC MNs, and p .vs.nontreated N microglia).Besides RAGE, elevation of TLR was also identified in.