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Differential behavior of VEGF receptor expression and response to TNP-470 in two immortalized human endothelial cell
1Department of Surgery, Tokyo Metropolitan Komagome Hospital, Bunkyo-ku, Japan.
Abstract:
Angiogenesis consists of endothelial cell proliferation, migration and tube formation. It is useful to investigate endothelial cell behavior using immortalized endothelial cell lines. We characterized cell growth property, growth factor dependency and response to angioinhibitory drugs; TNP-470, staurosporine, radicicol and genistein, using human umbilical vein endothelial cells (HUVECs) immortalized by human papilloma virus (HPV)-16 E6-E7, named HUVECs/E6-E7, and HUVECs/E6-E7 transformed by v-Ki-ras gene, named HUVECs/E6-E7/ras. The dependency to vascular endothelial cell growth factor (VEGF) and basic fibroblast growth factor (bFGF) for cell proliferation decreased in HUVECs/E6-E7, but were restored in HUVECs/E6-E7/ras. Flow cytometric analysis demonstrated that a VEGF receptor KDR/flk-1 was down-regulated in HUVECs/E6-E7 but not in HUVECs/E6-E7/ras. Expression of another VEGF receptor flt-1 was consistent in all cells including HUVECs, HUVECs/E6-E7 and HUVECs/E6-E7/ras. According to the analysis of the angioinhibitory drugs, HUVECs/E6-E7 was obviously resistant to TNP-470, but HUVECs/E6-E7/ras showed similar response compared to HUVECs which suggests that v-Ki-ras signaling pathway is associated with VEGF receptor expression and make HUVECs/E6-E7 sensitive to TNP-470 by modulating the signal transduction cascade. In conclusion, HPV-16 E6-E7 and v-Ki-ras genes have unique growth properties and these immortalized cells are useful for investigating signal transduction pathways of endothelial cells, and for screening of angioinhibitory drugs.
Insights
Immortalized endothelial cells (HUVECs/E6-E7) exhibit altered growth factor dependency and drug resistance. Co-transformation with v-Ki-ras (HUVECs/E6-E7/ras) restores sensitivity and modulates VEGF receptor expression, aiding angiogenesis research.
Area of Science:
- Endothelial cell biology
- Cancer research
- Molecular oncology
Background:
- Angiogenesis is crucial for endothelial cell proliferation, migration, and tube formation.
- Immortalized endothelial cell lines are valuable tools for studying endothelial cell behavior.
- Human umbilical vein endothelial cells (HUVECs) immortalized with HPV-16 E6-E7 (HUVECs/E6-E7) and v-Ki-ras (HUVECs/E6-E7/ras) were utilized.
Purpose of the Study:
- To characterize the growth properties, growth factor dependency, and drug response of immortalized HUVECs.
- To investigate the role of HPV-16 E6-E7 and v-Ki-ras genes in endothelial cell signaling.
- To assess the utility of these cell lines for screening anti-angiogenic drugs.
Main Methods:
- Cell proliferation assays were performed.
- Growth factor dependency on vascular endothelial cell growth factor (VEGF) and basic fibroblast growth factor (bFGF) was assessed.
- Flow cytometry analyzed VEGF receptor (KDR/flk-1 and flt-1) expression.
- Response to angioinhibitory drugs (TNP-470, staurosporine, radicicol, genistein) was evaluated.
Main Results:
- HUVECs/E6-E7 showed decreased dependency on VEGF and bFGF, while HUVECs/E6-E7/ras restored this dependency.
- KDR/flk-1 was downregulated in HUVECs/E6-E7 but not in HUVECs/E6-E7/ras; flt-1 expression was consistent across cell lines.
- HUVECs/E6-E7 exhibited resistance to TNP-470, whereas HUVECs/E6-E7/ras showed similar sensitivity to HUVECs, suggesting v-Ki-ras involvement in drug response.
Conclusions:
- HPV-16 E6-E7 and v-Ki-ras genes confer unique growth properties to immortalized endothelial cells.
- These cell lines are valuable for investigating endothelial cell signal transduction pathways.
- The characterized cell lines serve as a useful platform for screening anti-angiogenic drugs.