Skip to main content
. Author manuscript; available in PMC: 2019 Oct 29.
Published in final edited form as: Cell Rep. 2019 Sep 17;28(12):3105–3119.e7. doi: 10.1016/j.celrep.2019.08.036

Figure 5. miR-21 Regulates Selected Target Genes In Vitro in Primary Neonatal Microglia.

Figure 5.

(A) In comparison with scrambled control, transfection of miR-21-mimic in primary neonatal miR-21-null mice microglia results in significant downregulation of miR-21 target genes (Btg2, Nfat5, and Pcdc4) normalized to β-Actin.

(B) Transfection of miR-21-mimic in primary neonatal miR-21-null mice microglia did not affect a control gene Gaphd (not targeted by miR-21).

(C) Schematic overview of EV isolation using differential centrifugation. EVs from conditioned media cultured with GL261 cells (EV) and unconditioned media (UcM) were subjected to differential centrifugation. Collected EV pellet was resuspended in PBS and added to miR-21-null microglia.

(D) EVs from conditioned media (EV) and unconditioned media (UcM) were isolated and added to miR-21-null primary microglia followed by 24 h incubation. Increased levels of miR-21 were detected in microglia exposed to GL261-derived EV, compared with control media, with Ct > 40 considered baseline.

(E and F) Fold expression of miR-21 target genes (Bmpr2, Btg2, Kbtbd2, Nfat5, Pdcd4, Pten, Rhob, Smad7, and Ski) (E) and (F) Gapdh, a gene not targeted by miR-21, normalized to β-Actin in miR-21-null microglia exposed to GL261-derived EVs, compared with UcM.

Data represent three independent experiments and are presented as the mean with SEM (error bars). **p < 0.01 and ***p < 0.001, unpaired t test and multiple t test.