NDT Advance Access published online on November 16, 2004
Nephrology Dialysis Transplantation, doi:10.1093/ndt/gfh572
© 2004 by European Renal Association - European Dialysis and Transplant Association
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1 Department of Medicine and Clinical Science, Gunma University Graduate School of Medicine, Maebashi, Japan
* To whom correspondence should be addressed. Background. There are conflicting results regarding the role of periendothelial mural cells in angiogenesis. In the current study, we investigated the role of mesangial cells (MCs) in endothelial vascularization by using a three-dimensional co-culture system in basement-membrane reconstruct gel (Matrigel). Methods. Human umbilical vein endothelial cells (ECs) and human MCs were used. In the contact co-culture system, ECs and MCs were mixed and then plated together onto Matrigel. In the non-contact co-culture system, MCs were cultured within an intercup chamber, which prevented direct physical contact with the ECs on the Matrigel but allowed both cell types to share culture medium. To visualize ECs and MCs, the cells were labelled with two different fluorescent dyes prior to the co-culture. A capillary-like network formation was observed under a fluorescent microscope and confocal microscope, and the length of the network formation was quantified by the image analyzer. Results. ECs barely formed capillary-like networks when cultured alone in growth factor-free medium. However, ECs cultured with MCs in a contact condition remarkably formed capillary-like networks (9.10±0.96 vs 0.20±0.07 mm/mm2 at 6 h, contact vs ECs alone, P<0.001). Direct contact between ECs and MCs was clearly demonstrated by confocal microscopy. Differentiation into branching capillary-like structures was also observed in the non-contact co-culture system (3.02±1.21 mm/mm2 at 6 h, P<0.001 vs ECs alone), but less prominently than in the contact co-culture condition. Vascular endothelial growth factor (VEGF) was secreted from MCs, as determined by enzyme-linked immunosorbent assay and immunofluorescent study. Adding neutralizing antibodies against VEGF into the co-culture system partially inhibited capillary network formation. Conclusions. Our data indicate that MCs help ECs differentiate toward vascularization, in which the direct cell-cell contact between ECs and MCs plays an important role. VEGF is a mediator in this process.
Accepted October 6, 2004
Original Articles
Mesangial cells stimulate differentiation of endothelial cells to form capillary-like networks in a three-dimensional culture system
Tokuyuki Kitahara, E-mail: tkita1{at}jcom.home.ne.jp
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