Panoply™ Human FAS Over-expressing Stable Cell Line

MicroRNA-25 (miR-25) has been reported to be an important miRNA marker in neural cells and is highly expressed in ischemic brain tissue. However, the precise mechanism and role of miR-25 in cerebral ischemia/reperfusion (I/R) injury require further investigation. Here, researchers established an oxygen-glucose deprivation (OGD) model in human SH-SY5Y and IMR-32 cells to mimic I/R injury and evaluated the role of miR-25 in regulating OGD/reperfusion (OGDR)-induced apoptosis. They found that miR-25 was downregulated in the OGDR model. Overexpression of miR-25 via miRNA mimic transfection significantly inhibited OGDR-induced apoptosis. Furthermore, bioinformatics analysis identified Fas as a target gene of miR-25. Dual-luciferase reporter assays confirmed the interaction between miR-25 and the 3′-untranslated region (UTR) of Fas mRNA. In the OGDR model, overexpression of miR-25 led to downregulation of Fas protein expression. Subsequently, small interfering RNA (siRNA)-mediated knockdown of Fas expression also inhibited OGDR-induced cell apoptosis. Conversely, overexpression of Fas abolished the protective effect of miR-25 on OGDR-induced cells. Taken together, these results suggest that upregulation of miR-25 may inhibit cerebral ischemia-reperfusion injury-induced apoptosis by downregulating Fas/FasL, making it a promising therapeutic target.

Here, researchers conducted MTT and caspase-3 assays to investigate the effects of Fas overexpression and silencing on cell viability and caspase-3 activity. In Fas-overexpressing cells, cell viability was reduced to a level comparable to that in the OGDR group (Figure 1a). Conversely, Fas silencing significantly improved cell viability. Figure 1b shows that caspase-3 activity was enhanced in Fas-overexpressing cells but significantly suppressed by Fas silencing. Taken together, these results demonstrate that Fas overexpression significantly inhibits cell survival, even in the absence of OGDR. Furthermore, Fas silencing exhibited the opposite effect, reversing the effects of OGDR treatment.

Figure 1. Fas overexpression inhibited cell survival, whereas Fas silencing promoted cell survival. (Zhang J, et al., 2016)