Leukemia inhibitory factor-transfected embryonic fibroblasts and vascular endothelial growth factor successfully improve the skin substitute wound healing by increasing angiogenesis and matrix production

Abstract

Background and objective : The combined application of cytokines on embryonic fibroblasts and dermal substitute were studied for optimal skin defect coverage. The mechanism of combined treatment of leukemia inhibitory factor (LIF)-transfected embryonic fibroblasts and vascular endothelial growth factor (VEGF) were elucidated and subsequently the in vivo applications of both were tested in an artificial dermal substitute. Methods : Mouse embryonic fibroblast cells, BALB-3T3, were stably transfected with mouse full-length LIF cDNA and added to various doses of VEGF for detection of signaling interaction. LIF-transfected cells and VEGF treatment were tested with pig-tendon derived collagen dermal substitute in the backs of BALB/c male mice up to for 14 days. Results : LIF-transfected cells as well as vector-transfected fibroblasts significantly proliferated by 1, 10, or 100 ng VEGF on days 3 and 5. Erk mitogen-activated protein (MAP) kinase phosphorylation was observed from 1 to 30 min in LIF-transfected and 10 ng of VEFG, and 1 to 60 min in LIF-transfected and 100 ng VEFG treatments. The cellular fibronectin levels also increased in LIF-transfected cells with 10 and 100 ng VEGF additions. In in vivo analyses, LIF-transfected embryonic fibroblasts with 50 μg of VEGF markedly enhanced collagen I expression and CD34 angiogenic marker on days 7 and 14. Conclusion : LIF transfection and VEGF treatment enhanced phosphorylated-Erk-MAP kinase in vitro. In vivo study revealed that the combined application of LIF transfection of embryonic fibroblasts with an angiogenic factor such as VEGF in the template of a dermal substitute induced greater skin collagen production and angiogenesis in the dermal substitute.