Effects of hypoxia inducible factor-1α on expression levels of MLCK, p-MLC and ZO-1 of rat endothelial cells.

OBJECTIVE:To examine the aberrant expression of endothelial permeability associated proteins including MLCK, p-MLC and ZO-1 in presence of different levels of hypoxia-inducible factor 1 alpha (HIF-1α). METHODS:We established monolayer vascular endothelial cell model with the primary rat endothelial cells. Over-expressed or under-expressed HIF-1α cell lines were made by endothelial cells transfected with plasmid vector constructed with HIF-1α gene or HIF-1α-specific short hairpin RNA (shRNA). Levels of mRNA and protein of MLCK, p-MLC and ZO-1 were determined using Real-Time PCR and Western blot. All data were analyzed using by One-Way ANOVA method and LSD. RESULTS:We successfully cultured the rat endothelial primary cells for four days. The mRNA and protein levels of MLCK and p-MLC were significantly increased in the HIF-1α over-expression group than that in the blank control group and the empty plasmid GV230 group (P＜0.05). ZO-1 was significantly lower in the HIF-1α over-expression group than that in the blank control group and the GV230 group. On the contrary, the mRNA and protein levels of MLCK and p-MLC were significantly lower in the HIF-1α under-expression group than that in the blank control group and the shRNA-NC group (P＜0.05). ZO-1 was significantly higher in the HIF-1α low-expression group than that in the blank control group and the shRNA-NC group. CONCLUSION:HIF-1α positively regulates the expression of MLCK and p-MLC and negatively regulates the expression of ZO-1 in rat monolayer endothelial cells.

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