P38 MAPK信号通路参与BMP-13诱导C3H10T1/2细胞向心肌样细胞分化

 目的 探讨P38MAPK对BMP-13诱导C3H10T1/2细胞向心肌样细胞分化的影响。方法 实验4个部分分组如下：1.BMP-13腺病毒（Ad-BMP-13）对P38MAPK的作用：Ad-BMP-13转染组、Ad-GFP转染组和C3H10空白组。Western blot检测磷酸化P38MAPK（p-P38MAPK）和总P38MAPK（t-P38MAPK）的表达变化，免疫荧光技术定位p-P38MAPK；2.P38MAPK干扰腺病毒（Ad-si-P38）对P38MAPK的作用：si-P38干扰组、si-NC干扰对照组和C3H10空白组。Western blot检测t-P38MAPK的表达；3.Ad-si-P38阻断P38MAPK后对BMP-13诱导分化的影响：si-P38+Ad-BMP-13转染组、si-NC+Ad-BMP-13转染组、si-NC+Ad-GFP转染组和C3H10空白组。Western blot检测cTnT和Cx43的表达，荧光定量PCR检测GATA-4和MEF-2C的mRNA表达；4.SB203580阻断P38MAPK后对BMP-13诱导分化的影响： DMSO+Ad-BMP-13转染组、SB203580(2、5和10μmol/L)+Ad-BMP-13转染组 。荧光定量PCR检测GATA-4和MEF-2C的mRNA表达。结果 BMP-13促进P38MAPK的磷酸化。Ad-si-P38可以有效降低P38MAPK表达水平。Ad-si-P38阻断P38MAPK后BMP-13诱导组cTnT、Cx43表达有明显降低（P<0.05），GATA-4和MEF-2C的表达也有显著降低（P<0.05）。随P38MAPK特异性抑制剂SB203580浓度增加，BMP-13诱导组GATA-4和MEF-2C的表达降低（P<0.05）。结论 Ad-BMP-13可以通过激活P38MAPK信号通路来调控C3H10T1/2细胞向心肌样细胞分化。

P38 MAPK signaling pathway is involved in BMP-13-induced cardiomyocyte-like differentiation from C3H10T1/2 cells

Objective To investigate the role of P38MAPK in BMP-13-induced differentiation of C3H10T1/2 cells into cardiomyocyte-like cells. Methods The four parts of experiment are grouped as follows:1.BMP-13 adenovirus (Ad-BMP-13) on the role of P38MAPK:Ad-BMP-13 transfection group, Ad - GFP transfection group and C3H10 blank group. The phosphorylated P38MAPK (p-P38MAPK) and total P38MAPK (t-P38MAPK) were detected by Western blot . The positioning of p-P38MAPK was detected by immunofluorescence technique;2. P38 MAPK interference adenovirus (Ad-si-P38) on the role of P38MAPK:si-P38 interference group,si-NC control interference group and C3H10 blank group. The t-P38MAPK was detected by Western blot.3. The influence of BMP-13 induced differentiation after Ad-si-P38 blocking P38MAPK signal pathway:si-P38+Ad-BMP-13 transfection group,si-NC+Ad-BMP-13 transfection group,si-NC+Ad-GFP transfection group and C3H10 blank group.The cTnT and Cx43 were detected by Western blot and the GATA-4 and MEF-2C were detected by fluorescent quantitative PCR.4. The influence of BMP-13 induced differentiation after SB203580 blocking P38MAPK signal pathway: DMSO+Ad-BMP-13 transfection group,SB203580(2,5 and 10μmol/L)+ Ad-BMP-13 transfection group.The GATA-4 and MEF-2C were detected by by fluorescent quantitative PCR.Results BMP-13 promoted the P38MAPK phosphorylation. Ad-si-P38 can effectively lower the P38MAPK expression. Ad-si-P38 which can block P38MAPK signal pathway significantly inhibited the BMP-13-induced expression of cTnT,Cx43 (P<0.05) and GATA4,MEF-2C (P<0.05). With the increased concentration of P38MAPK specific inhibitor SB203580, expression of GATA-4,MEF-2C was reduced significantly (P<0.05).Conclusions P38MAPK signal pathway can be activated by Ad-BMP-13 to promote cardiomyocyte -like cells differentiation from C3H10T1/2 cells.