关键词: pEGFP-N2-BMP2, 基因转染, P19细胞, 心肌细胞, 免疫组织化学

Abstract: Objective To identify occurrence of cardiomyocyte differentiation from P19 cells transfected with pEGFP-N2-BMP2 eukaryotic expression plasmid. Methods P19 cells were divided into transfected and non-transfected groups.In the transfected group,the cells with stably expressed of BMP2 as being transfected with pEGFP-N2-BMP2 eukaryotic expression plasmid and then were cultured in suspension. For 4 days, and the formed aggregates were transferred to Petri dish for adherent culture.On days 4, 8, 12, and 16 of the adherent culture, the expressions of cardiac troponin T（cTnT）and α-cardiac actin were detected with immunocytochemistry and Western blotting,while the gene expression of GATA-4 and NKx2-5 was detected with semiquantitative RT-PCR. Results In the transfected group, no expression of cTnT and α-cardiac actin was found on day 4, and the expression of these 2 proteins existed on days 8, 12, and 16 by both immunocytochemistry and Western blotting. The expression of two proteins was gradually increased with the prolong of time. There was a obvious difference between every two different drawing materials time point(EM>P/EM>0.01). The expression of GATA-4 and NKx2-5 was appeared with a weak expression at 4 days after transfection, as time went by, at 8 days, 12 days and 16 days after transfection, the expression of two gene intensified gradually. Also,the expression of two genes at every two different drawing materials time point had a significantly difference (EM>P/EM>0.01). In the non-transfected group, neither the protein expression of cTnT and α-cardiac actin nor the gene expression of GATA-4 and NKx2-5 was found. Conclusion Stable expression of BMP2 gene can induce cardiomyocyte differentiation from P19 cells. Possibly, BMP2 regulates the transcrip

Key words: pEGFP-N2-BMP2, Gene transfection, P19 cell, Cardiomyocyte, Immunohistochemistry