Figure 7. miR-26a attenuates neointimal formation in a rat model of autogenous jugular vein graft.

(A) LV3-mediated overexpression of miR-26a (LV3-miR-26a) significantly reduced neointimal formation in vivo. Representative Elastica van Gieson -stained jugular veins from rats treated with LV3 or LV3-miR-26a 4 weeks after autogenous jugular graft. Black arrows indicate internal elastic membrane. Cross-sectional areas of the intima and media were marked with N and M respectively. (B) The effect of miR-26a on vascular neointimal lesion formation in rat jugular veins 4 weeks after autogenous jugular graft as quantitated by neointima/media (N/M) ratio. *P < 0.05 represents statistical significance compared with LV3-NC. (C) LV3-miR-26a (10⁸ pfu/ml) or LV3-NC (10⁸ pfu/ml) was suspended in 50 μl Pluronic F127 gel (BASF, 30% wt/vol) and was applied around the grafted-jugular vein. Jugular veins were harvested 4 weeks after graft. Total RNAs were extracted, and the expression of miR-26a was detected by qRT-PCR (n = 4). The expression of miR-26a in rat jugular vein transduced with LV3-NC was undetectable (ND). *P < 0.05 represents statistical significance compared with LV3-NC. (D) Representative immunofluorescence staining of PCNA in rat jugular veins 4 weeks after ligation injury. Green indicates PCNA, which represents proliferating cells; blue is the cell nuclear staining by DAPI. (E) Quantification of PCNA-positive cells showed that fewer cells were proliferating in the grafted-vein treated with LV3-miR-26a compared with LV3-treated vessels. *P < 0.05 represents statistical significance compared with LV3-NC. (F) Representative immunofluorescence staining of MAPK6 in rat jugular vein 4 weeks after ligation injury. Green indicates MAPK6 staining, and blue is nuclear staining by DAPI. (G) Quantification of MAPK6-positive cells showed that MAPK6 was downregulated in LV3-miR-26a-treated vessels compared with LV3-treated vessels. *P < 0.05 represents statistical significance compared with LV3-NC.