Pericyte-based collagen scaffold for vascular tissue engineering

摘要

Pericytes are a perivascular cell population embedded on the capillary wall in a shared basement membrane with the endothelium. During vasculogenesis and angiogenesis, endothelial cells secrete and/or present cell bound molecular cues that stimulate pericytes to proliferate, migrate and attach to nascent capillary tubes. It has been previously developed cell sheet engineering using temperature-responsive culture dishes in order to avoid traditional tissue engineering approaches. In this study our aim is to isolate human umbilical cord vein pericytes then differentiate into smooth muscle cells and fibroblasts then co-culture these cells on 3D collagen scaffold that will be used for generating tissue engineering vascular grafts. Pericytes were enzymatically isolated from human umbilical cord vein and were purified by using MACs cell separation with CD146 microbeads. They were cultured in fibroblast growth medium and smooth muscle growth medium for 21 days on poly(N-isopropylacrylamide) to obtain cell sheets. Cell differentiation was confirmed by immunofluorescence staining and flow cytometry analysis (fibroblast markers; Tenascin-C and Collagen type Ⅰ, smooth muscle cell markers; Caldesmon, Calponin and Alpha smooth muscle actin). Collagen gel was prepared with human collagen type Ⅰ (3mg/mL) + 10X DMEM (adjustment pH of mixture to 7.2-7.6 using sterile 0.1 M NaOH). Collagen gel was covered by cell sheets (sandwich system) and vascular graft was incubated in 37oC for 7 days (Figure 1). The results indicated that perivascular cells could differentiate into fibroblast and smooth muscle and formed vascular graft with collagen gel. As a conclusion, differentiated pericytes offer an alternative cell source for constructing tissue engineered vascular graft. The authors wish to thank The Scientific and Technological Research Council of Turkey (Project Number 113S815) for their financial support.