Low-dose metronomic chemotherapy with cisplatin: can it suppress angiogenesis in H22 hepatocarcinoma cells?

Abstract

Low-dose chemotherapy drugs can suppress tumours by restraining tumour vessel growth and preventing the repair of damaged vascular endothelial cells. Cisplatin is a broad-spectrum, cell cycle-non-specific drug, but has serious side effects if used at high doses. There have been few reports on the anti-angiogenic effects of low-dose cisplatin and hence the effect of low-dose metronomic (LDM) chemotherapy on the proliferation and neovascularization of H22 hepatocarcinoma cells is discussed in this research. The influence of LDM chemotherapy with cisplatin on human umbilical vascular endothelial cells (HUVECs) and proliferation of the HepG(2) human hepatocarcinoma cell line were measured using MTT assays. The LDM group was treated with cisplatin 0.6 mg/kg/day; the control group with saline 0.2 ml; the maximum tolerated dose (MTD) group with cisplatin 9 mg/kg/day. Vascular endothelial growth factor (VEGF) and matrix metallopeptidase 2 (MMP-2) were detected using immunohistochemical staining. A chicken chorio-allantoic membrane (CAM) model was used to check the inhibitory effect of LDM chemotherapy with cisplatin on neovascularization in vivo. Low-dose cisplatin inhibited HUVEC proliferation in a dose- and time-dependent manner, but was ineffective in inhibiting HepG(2) cell proliferation. Tumour growth was delayed in mice receiving LDM cisplatin, without apparent body weight loss, compared with mice that received MTD cisplatin. Microvessel density and expression of VEGF and MMP-2 were much lower in mice receiving LDM cisplatin than in the control and MTD groups. Continuous low-dose cisplatin suppressed CAM angiogenesis in vivo. LDM chemotherapy with cisplatin can inhibit the growth of blood vessel endothelial cells in vitro and shows anti-angiogenic ability in vivo.

Figure 1

The growth curve of tumour volume (V/mm³) in each group.

Figure 2

Immunohistochemistry staining in H22 hepatocarcinoma cells. (a) CD34 staining in H22 hepatocarcinoma cells. Groups include cells receiving maximum tolerable doses (MTD) and low-dose metronomic (LDM) amounts of cisplatin and control group (×400). (b) VEGF staining in H22 hepatocarcinoma cells (×400). (c) MMP-2 staining in H22 hepatocarcinoma cells (×400). The microvessel density and VEGF and MMP-2 levels in the LDM group were significantly lower than those in the control and MTD groups.