In vitro induction, differentiation and seeding of source cells for the endothelialization of small-diameter artificial blood vessel of polyurethane / 中国组织工程研究

ABSTRACT

BACKGROUND: At present, after transplantation of small diameter artificial blood vessel, long-term patency rate is low due to being lacking of endothelial cells for lining and anti-thrombus characters. In some studies,mature endothelial cells were tried to be seeded in the artificial vessel to boost up its anti-thrombus capability so as to improve the long-term patency rate, but we got unsatisfied effect due to the defects of seed cells and scaffolds. Therefore, in clinic, proper seed cells and vascular scaffolds have been searched for improving the long-term low pateney rate in transplantation of small diameter artificial blood vessel.OBJECTIVE: To investigate the feasibility that differentiation of bone marrow mononuclear cells induced in vitro into endothelial-progenitor cells (EPCs) and seed polyurethane small diameter artificial blood vessel so as to provide proper seed cells for endotheliazation of polyurethane small diameter artificial blood vessel.DESIGN: Observation experiment SETTING: Cardivascular Medical Department and Staff Room of Immunology, First Hospital Affiliated to Sun Yat-sen University MATERILAS This experiment was carried out at the First Hospital Affiliated to Sun Yat-sen University from September 2004 to May 2005. About 10 mL of bone marrow from healthy adult volunteers (n=7) was used in this experiment.METHODS: Bone marrow mononuclear cells of healthy adult were collected and put in the fibronectin pre-coated DMEM culture medium, then induced by vascular endothelial growth factor and basic fibroblast growth factor. Induced cells were observed under fluorescence microscope and identified with immunohistochemical staining. The induced and proliferated EPCs were seeded onto the surface of polyurethane small diameter artificial blood vessel. Morphological change was observed under scanning electron microscope.MAIN OUTCOME MEASURES: ① Cellular morphological change.② Staining results of immunohistochemical VWF and CD 34 antibody . ③ Adhesive growth status of EPCs on the polyurethane small diameter artificial blood vessel RESULTS: ① In the vascular endothelial growth factor and basic fibroblast growth factor and other inducers , bone marrow mononuclear cells differentiated into EPCs , presenting typical "spindle-shaped" appearance under an inverted fluorescence microscope and became to form a monolayer that arrayed in "cobblestone-like" ② Immunohistochemical staining showed von willebrand factor(VWF) and CD34 antigen stained positive. ③ Under the scanning electron microscope, surface of polyurethane small diameter artificial blood vessel without seeded cells presented typical polyporous honeycomb-like structure , and the size of hole suited the crawling of EPCs. After seeding the cells, we observed the adhesion, crawling and spreading of the EPCs on the surface of polyurethane small diameter artificial blood vessel. Some EPCs grew into the honeycomb-like holes were seen occasionally.CONCLUSION: Bone marrow mononuclear cells can be induced and differentiated into EPCs, while induced and differentiated EPCs well grow adhesively in the polyurethane small diameter artificial vessels, suggesting that differentiation of bone marrow mononuclear cells induced in vitro into EPCS, which can be used as seed cells for endothelialization of polyurethane small diameter artificial blood vessels.