Matrix-embedded endothelial cells are protected from the uremic milieu

Abstract

Background. Endothelial cells (ECs) embedded in 3D matrices [matrix-embedded endothelial cells (MEECs)] of denatured collagen implanted around vascular access anastomoses preserve luminal patency. MEEC implant efficacy depends on embedded EC health. As the uremic milieu inhibits proliferation and induces apoptosis of ECs, we examined whether uremia might impact MEECs. Methods. ECs grown on 2D gelatin-coated polystyrene tissue culture plates (gTCPS) or in MEEC were treated with sera pooled from 20 healthy control or uremic patients with end-stage renal disease. EC viability was examined using 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide assay, cell counting and Trypan blue exclusion. Media conditioned (CM) with 2 and 3D-supported ECs were examined for its potential to inhibit vascular smooth muscle cell (vSMC) proliferation using ³ [H] thymidine incorporation and cyclin D1 expression. ECs grown on gTCPS were treated with uremic serum filtered through matrices to examine if matrices retain uremic toxins or whether EC effects were cell mediated. Results. Uremic serum significantly reduced viability and number of live, and increased dead ECs when grown on gTCPS, but not in MEECs. EC survival correlated with vSMC inhibition. While CM from ECs grown in gTCPS with uremic serum inhibited vSMC proliferation no better than uremic serum alone (22 versus 27%), MEEC CM inhibited vSMC proliferation by 47% (P = 0.0004). Cyclin D1 expression tracked with indices of vSMC proliferation. There was no significant difference in EC viability between EC treated with matrix-filtered or unfiltered uremic serum. Conclusion. The viability, number and efficacy of MEECs were preserved in uremic serum compared to those of ECs on gTCPS. MEECs are protected from uremic toxicity, not from retention of uremic toxins by matrices, but likely from intrinsic changes in EC sensitivity to uremia. MEECs implanted at vascular access sites should inhibit neointimal hyperplasia in uremia. This study underscores the robustness of matrix embedding as a cell protectant, especially in hostile environments like uremia.