Synergistic effect of bevacizumab and celecoxib on angiogenesis in vitro using human umbilical vein endothelial cells.

Angiogenesis is the underlying cause of a large number of neoplastic diseases. It is necessary for tumor metastasis, and without it the tumor cannot grow or metastasize. This study aimed to determine the synergistic effect of bevacizumab and celecoxib on angiogenesis using human umbilical vein endothelial cells (HUVEC) as an in vitro model. HUVEC were isolated from the umbilical cord by enzymatic digestion using collagenase type IV. HUVEC characterization was done by flow cytometry using cell surface markers CD31, CD105, CD146, and CD45. HUVEC were treated with bevacizumab, celecoxib, and the combination of both drugs and the cell viability was assessed using MTT assay. The formation of capillary-like endotubes for angiogenesis was analyzed using a tube formation assay by measuring the total length of capillary tubes and branch points. Morphologically, HUVEC showed a typical cobblestone appearance using inverted-phase contrast microscopy and were further evaluated using flow cytometry, which showed positive expression for cell surface markers CD31, CD105, CD146, and negative for CD45. Celecoxib, bevacizumab, and the combination of both drugs showed a dose-dependent inhibition on HUVEC viability. Celecoxib inhibited total tube length by 15% and branch points by 16.5%. Bevacizumab inhibited total tube length by 34% and branch points by 49%. When the two drugs were combined, the total tube length was reduced due to synergism by 68% and branch points by 80%, and the difference was found to be statistically significant (p < 0.001). Bevacizumab and celecoxib have a synergistic effect in inhibiting in vitro angiogenesis and their combination achieved more strong inhibition than either drug alone.