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Figure 4: Effect of pre-miRNA589 on expression ZO-1, E-cadherin, and vimentin in HMrSV5 cells following exposure to TGFβ1(5 ng/ml, 24 hr). Panel (a) and (b) showed the ZO-1 and vimentin protein expression detected by immunofluerence. Fluorescent confocal microscopy for ZO-1 and vimentin confirmed that pretreatment of pre-miR589 normalized the EMT changes induced by TGFβ1 in HMrSV5 cells. Panel (c) and (d) showed that TGFβ1 treatment led a reduction in the mRNA and protein expression of ZO-1 (Panel (c), column 2 and panel (d), column 2). These changes were attenuated by the overexpression of miRNA589 with treatment of pre-miRNA589 (Panel (c), column 3 and panel (d), column 3). The pre-miRNA589 partially reversed the TGFβ1-induced mRNA and protein expression of E-cadherin changes as well as ZO-1 (Panel (e), columns 2 and 3, and panel (f), columns 2 and 3). The level of ZO-1and E-cadherin was decreased, while the level of vimentin mRNA as well as protein were increased in HMrSV5 cells treated with TGFβ1(5 ng/ml, 24 hr) (Panel (g), column 2 and panel (h), column 2), and this effect was abolished following the transfection with pre-miRNA589 (Panel (g), column 3, and panel (h) column 3). *P < 0.05. versus control group, **P < 0.05 versus TGF-β1 treated group. #P < 0.01 versus control group. ##P < 0.01 versus TGF-β1 treated group.