Pro-angiogenic effect of SEHM formula on HUVEC cells in vitro and Zebrafish in vivo and action mecha nism study

Abstract

Aim To investigate pro-angiogenic effect of SEHM formula on HUVEC cells in vitro and Zebrafish in vivo and the related action mechanism. Methods VEGFR tyrosine kinase inhibitor II ( VRI)-induced zebrafish intersegmental vessels ( ISVs) damaged model was established to observe the protective effect of SEHM formula on ISVs under fluorescence microscope. The pro-angiogenic effect on subintestinal vessels (SIVs) of water decoction of SEHM formula in zebrafish was observed and the mRNA level of VEGFRs, including fit-i, kdr, kdrl, was measured by real-time PCR. The experimental models of the HUVEC cells were set up and the toxicity and promoting proliferation effect of water decoction of SEHM formula in HUVEC cells were assessed by cell viability assay( Ml]'), and then the levels of Akt, p38, p-p38, p44/42, p-p44/42, VEGFR-1 were detected by Western blot at 6, 12 and 24 h. Results SEHM formula treatment groups could significantly protect VRI-induced zebrafish ISVs loss( P<0. 01) and presented pro-angiogenic effect on zebrafish SIVs obviously ( P<0. 01). The mRNA level of VEGFRs, including flt-l, kdr, kdrl. was up-regulated by SEHM formula treatment group significantly (P<0. 05) compared with VRI group. Compared with control group, and SEHM formula treatment group could apparently promote proliferation in HUVEC cells and up-regulate the level of Akt, p38, p-p38, p44/42, p-p44/42, VEGFR-l. Conclusions Water decoction of SEHM formula could present pro-angiogenic effect on SIVs in zebrafish and promote proliferation of HUVEC cells significantly, and its action mechanism may be related to up-regulating the expression of VEGFR.