Inhibitory effect of tenomodulin versus ranibizumab on in vitro angiogenesis

Abstract

Aim: To evaluate anti-angiogenic effect of tenomodulin (TNMD) and ranibizumab on cell proliferation and capillary-like morphogenesis of vascular endothelial cells under the stimulation of vascular endothelial growth factor (VEGF) in vitro.

Methods: The effects of TNMD and ranibizumab on VEGF-induced proliferation of human umbilical vein endothelial cells (HUVECs) were evaluated by MTT assay, and the effects of TNMD and ranibizumab on capillary-like structures formed by HUVECs under the stimulation of VEGF were examined in culture. Capillary-like morphogenesis of HUVECs was quantitatively evaluated, and total lengths of tube-like structures per field were measured in a masked way.

Results: HUVECs with both ranibizumab and TNMD protein showed MTT reduction in VEGF-stimulated cell proliferation as expected, while MTT absorbance in the HUVECs with TNMD was significantly declined than that with ranibizumab (P<0.01). The capillary-like structures formed by HUVECs were markedly impaired by the presence of both TNMD and ranibizumab in the culture medium. The total length of the capillary-like structures per field was significantly shorter in the medium with TNMD than that of ranibizumab (P<0.01). The inhibitory effect of TNMD on tube formation in vitro angiogenesis was significantly stronger than that of ranibizumab.

Conclusion: TNMD may have stronger inhibitory effect than ranibizumab on in vitro angiogenesis.

Keywords: angiogenesis; inhibitory effect; proliferation; ranibizumab; tenomodulin.

Figure 1. VEGF screened for optimum concentration of cell proliferation.

Figure 2. Inhibitory effect of TNMD and lucentis on VEGF-induced endothelial proliferation

The suppressive effect of TNMD and lucentis on VEGF-induced endothelial proliferation was evaluated by measurement of MTT assay.

Figure 3. Suppressive effect of TNMD/lucentis on VEGF-induced endothelial proliferation and on in vitro angiogenesis (×100)

The tube morphogenesis of HUVECs in different conditioned culture medium. A: 1% FBS containing medium as positive control; B: Medium including 1% FBS and 100 ng/mL VEGF; C: Medium including 1% FBS, 100 ng/mL VEGF and lucentis 1 µg/mL; D: Medium including 1% FBS, 100 ng/mL VEGF and TNMD 10 µg/mL. Bar chart: quantitative assessment of the length of the capillary structures by image computer analysis. Data are means±SD of three independent experiments. The total length of HUVECs incubated in the medium containing TNMD had significant differences with that of in the other three different conditioned culture medium (P<0.01).