The role of membrane complement regulatory protein in transplantation of bone marrow mesenchymal stem cell

Abstract

Objective To explore the effect of complement activation on bone marrow mesenchymal stem cells (BMSCs) and evaluate the effect after transfection of complement regulatory proteins. Methods Bone marrow aspirate was harvested from 10 cases of patients suffered from fractures. Mesenchymal stem cells were isolated, indentified cultured and then experimented in vitro. The complement cytotoxicity on the mesenchymal stem cells in autologous serum was measured by Europium cytotoxicity assay. The samples were divided into BMSCs group, BMSCs+ autologous human serum (AHS) group and BMSCs+ inactivated autologous human serum (iAHS) group. The complement membrane attack complex (MAC) deposited on the membranes was detected by flow cytometry. Finally, the cytotoxicity on BMSCs was measured after transfected with membrane complement regulatory proteins (mCRPs). All samples were divided into BMSCs with mCRPs untransfected group and BMSCs with mCRPs transfected group. Results More than 95% of cells derived from bone marrow were identified to be mesenchymal stem cells through detection of cell surface markers by flow cytometry. The cytotoxicity of untreated cells was 0.41%±1.48%. BMSCs harvested from the 10 patients all had cytotoxicity after incubated with autologous serum, and the cytotoxicity was 32.59%±2.73%, while cytotoxicity after incubated with complement inactivated autologous serum was 2.59%±3.08%, which was similar to control group. Complement attack complex (MAC) could be detected on the BMSCs incubated with autologous serum, which implied the complement activation. After transfection of mCRPs, the cytotoxicity of autologous serum on transfected cells was decreased. The cytotoxicity of untransfected cells (41.70%±4.47%) had significant difference compared to the cells transfected with CD55 (21.87%±2.19%), the cells transfected with CD59 (18.67%±1.42%), and the cells transfected with CD46+CD55+CD59 (28.43%±2.14%). CD55, CD59 and CD46+ CD55+CD59 transfected groups could impair effectively the cytotoxicity from complement. However, the cytotoxicity impairment was less effective in CD46 transfected cells (39.30%±3.96%), which had no significant difference compared to untransfected cells. Conclusion Membrane complement regulatory proteins could effectively protect bone marrow mesenchymal stem cells from attacks by complement. Key words: Bone marrow; Mesenchymal stem cells; Complement membrane attack complex