Objective: The discovery of vascular endothelial growth factor C (VEGF-C) and VEGF receptor-3 (VEGFR-3) has started to provide an understanding of the molecular mechanisms of lymphangiogenesis. The homeobox gene prox1 has been proven to specify lymphatic endothelial cells (ECs) from blood ECs. We investigated the process of lymphatic EC (LEC) differentiation using embryonic stem (ES) cells.
Methods And Results: VEGFR-2+ cells derived from ES cells differentiated into LECs at day 3 on OP9 stromal cells defined by the expression of prox1, VEGFR-3, and another lymphatic marker podoplanin. VEGFR-2+ cells gave rise to LYVE-1+ embryonic ECs, which were negative for prox1 on day 1 but turned to prox1+ LECs by day 3. VEGFR-3-Fc or Tie2-Fc, sequestering VEGF-C or angiopoietin1 (Ang1), suppressed colony formation of LECs on OP9 cells. However, addition of VEGF-C and Ang1 in combination with VEGF to the culture of VEGFR-2+ cells on collagen-coated dishes failed to induce LECs. LEC-inducing activity of OP9 cells was fully reproduced on paraformaldehyde-fixed OP9 cells with the conditioned medium.
Conclusions: We succeeded in differentiating LECs from ES cells and revealed the requirements of VEGF-C, Ang1, and other unknown factors for LEC differentiation.
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| http://dx.doi.org/10.1161/01.ATV.0000225770.57219.b0 | DOI Listing |
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