OBJECTIVE To observe the effects of Huangqi-Ezhu-Chonglou combination (HEC) on the permeability and tight junction of human umbilical vein endothelial cells (HUVEC) and to explore the mechanism of HEC on inhibiting the hematogenous metastasis of colon cancer cells (HCT116).
METHODS Control serum and HEC-medicated serum were prepared. The co-culture system of HUVEC-HCT116 was constructed and divided into single culture control group, co-culture model group and HEC low, medium and high dose (2.1, 4.2, 8.4 g ·kg-1) groups. In addition, HUVEC single culture control group was set up. Cell proliferation was detected by CCK-8 method; FITC-dextran method and transwell method were used to detect the permeability of HUVEC and the number of cross endothelial migration of HCT116 cells; The distribution of ZO-1 protein in HUVEC was detected by immunofluorescence; The protein expressions of RhoA, ROCK and ZO-1 were detected by Western blot.
RESULTS The proliferation of HUVEC was inhibited after 48 h of treatment with different doses of HEC medicated serum (P < 0.01). Compared with the single culture control group, FITC-dextran penetration and the number of transendothelial migration of HCT116 in the co-culture model group significantly increased (P < 0.01), the expression of ZO-1 was down-regulated (P < 0.01), and the expressions of RhoA and ROCK were up-regulated(P < 0.01); Compared with the co-culture model group, the FITC-dextran penetration and the number of transendothelial migration of HCT116 in the HEC group significantly decreased (P < 0.05), the expression of ZO-1 protein was up-regulated (P < 0.05), and the expressions of RhoA and ROCK were down-regulated in a dose-dependent manner (P < 0.05).
CONCLUSION HEC may regulate the expression of tight junction associated proteins ZO-1 in HUVEC by inhibiting RhoA/ROCK pathway, reduce vascular endothelial permeability, and then inhibit colon cancer hematogenous metastasis.
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