| Objective To observe the effect of Buyang Huanwu Decoction on proliferation,migration and ability of lumen formation in human brain microvascular endothelial cells(HBMECs)and explore the possible mechanism of BYHWD promoting angiogenesis via the expression of miR-210/VEGF.Methods Prepare Buyang Huanwu Decoction containing serum and culture human microvascular endothelial cells in vitro.Different concentration of BYHWD serum(5%、10%、20%)was added to HBMECs and the viability was detected by MTT method at different time(12、24、36、48、72 h).cells migration of HBMECs was detected by cell wound scratch test method.tube formation of HBMECs was detected by matrigel tube formation assay method the effect of BYHWD on expression of miR-210 was detected by qRT-PCRthe miR-210 inhibitor was transfected into HBMECs by lipofectamine 2000 and transfection efficiency was determined by qRT-PCR.After miR-210 inhibitor was transfected,the cell viability was detected by MTT assay,The migration ability of HBMECs was detected by cell wound scratch test and Transwell cell migration.The effect of BYHWD on tube formation of HBMECs was detected by matrigel tube formation assay.④After miR-210 inhibitor was transfected,qRT-PCR and Western Blot assay were used to detect the expression of miR-210、VEGF/VEGF2 to explore the possible mechanism.Results The HBMECs were cultured in vitro:cells were adherent grown well,single layer was paving stone arrangement,morphology was uniform and stable;MTT results suggest that compared with blank serum group,BYHWD serum containing HBMCEs proliferation,and incubated with 10%BYHWD and within 36 h was the best;cell wound scratch test and Transwell cell migration results showed that 10%BYHWD containing serum significantly enhanced the migration of HBMECs(P<0.05);Compared with blank serum group,qRT-PCR results showed that BYHWD containing serum could significantly increase the expression of miR-210 miRNA,BYHWD+ miR-210 inhibitor could induce expression of miR-210 miRNA(P<0.01);Compared with BYHWD group,the MTT assay showed that BYHWD+miR-210 inhibitor group inhibited the proliferation of HBMECs(P<0.01)and the number of tube formation was decreased(P<0.01);cell wound scratch test and Transwell cell migration showed that Compared with blank serum group,the migration ability of HBMECs was significantly increased;BYHWD+miR-210 inhibitor can decreased mRNA and protein expression of VEGF/VEGFR2(P<0.05).Conclusion BYHWD containing serum can upregulate miR-210 expression to promote proliferation and enhance cell migrationn of HBMECs,contribute to alumina formation to enhances angiogenesis,the possible mechanisms are related to upregulate expression of VEGF/VEGFR2. |
