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. 2021 Aug 30;17(4):824–831. doi: 10.4103/1673-5374.322466

Figure 2.

Figure 2

Inhibition of MEG3 suppresses OGD/R-induced HT22 autophagy.

Control group: Normal cultured HT22 cells; OGD/R group: HT22 cells were reoxygenated for 48 hours after OGD treatment for 6 hours; OGD/R + si-NC group: HT22 cells were transfected with si-NC before OGD/R treatment; OGD/R + si-MEG3 group: HT22 cells were transfected with si-MEG3 before OGD/R treatment; OGD/R + RAPA group: HT22 cells were exposed to RAPA before OGD/R treatment; OGD/R + si-MEG3 + RAPA group: HT22 cells were transfected with si-MEG3 and then exposed to RAPA before treatment with OGD/R. (A) MEG3 expression was quantified by qRT-PCR and normalized to the control group. (B, C) ATG7, Beclin1, and LC3II/I protein levels were evaluated by western blotting. The target protein expression was normalized to the control group. (D, E) MDC staining was used to assess cell autophagy (arrows) in HT22 cells (scale bar: 100 μm; original magnification 200×). si-MEG3 transfection increased cell autophagy, and MEG3 knockdown decreased cell autophagy. (F) Cell viability was examined by the CCK8 assay. Data are representative of three independent experiments (mean ± SD). ***P < 0.001, vs. control group; &P < 0.05, &&P < 0.01, vs. OGD/R group; ##P < 0.01, ###P < 0.001, vs. OGD/R + si-NC group; $P < 0.05, $$P < 0.01, $$$P < 0.001, vs. OGD/R + si-MEG3 group (one-way analysis of variance followed by the least significant difference post hoc test). ATG7: Autophagy-related gene 7; CCK8: Cell Counting Kit-8; CI/RI: cerebral ischemia-reperfusion injury; LC3: light chain 3; MDC: monodansylcadaverine; MEG3: maternally expressed gene 3; OGD/R: oxygen and glucose deprivation/reoxygenation; qRT-PCR: quantitative real-time PCR; RAPA: rapamycin.