瘢痕疙瘩分泌性卷曲相关蛋白2与骨母细胞特异性因子2的相互作用

The interaction between secreted frizzled-related protein 2 and osteoblast-specific factor 2 in keloid

目的验证分泌性卷曲相关蛋白2（SFRP2）与骨母细胞特异性因子2（OSF-2）的相互作用。方法构建能在哺乳动物细胞中表达带人膜联蛋白（HA）标签的OSF-2融合蛋白重组载体pCMV—HA—OSF-2，经酶切鉴定确认后，与Myc—SFRP2重组真核表达载体pCMV-HA-SFRP2单独或共转染人HK293细胞，利用免疫共沉淀技术及蛋白质印迹法验证OSF-2与SFRP2的相互作用。结果重组载体经酶切电泳后，显示近800bp处的条带为酶切后的目的片段SFRP2编码基因，近2500bp处的条带为酶切后目的基因OSF-2。表明pCMV-Myc-SFRP2和pCMV-HA—OSF-2均构建成功。HK293细胞单独转染pCMV—Myc—SFRP2或pCMV-HA-OSF-2后，未检测到HA—OSF-2的表达；当pCMV—Myc—SFRP2和pCMV—HA，OSF-2共转染后，经Myc抗体进行免疫沉淀可见HA—OSF-2表达。结论HA—OSF-2的重组载体能在哺乳动物细胞中表达，HA—OSF-2与SFRP2间存在相互作用。

Objective To verify the interaction between secreted frizzled-related protein 2 （SFRP2） and osteoblast-specific factor 2 （OSF-2）. Methods HA-tagged OSF-2 fusion protein recombinant vector pCMV-HA-OSF-2, which could express in mammal ceils was constructed, then identified by enzyme-cutting and transfected into human kidney 293 （HK293） cells with or without Myc-SFRP2 recombinant eukaryotic expression vector pCMV-HA-SFRP2. The interaction between SFRP2 and OSF-2 was detected through coimmunoprecipitation and Western blotting. Results In electrophoresis bath, target fragment of SFRP2 coding gene with 800 bp and target gene OSF-2 with 2500 bp could be seen respectively after enzyme-cutting, which showed that pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 were constructed successfully. No HA-OSF-2 expression was detected after pCMV-Myc-SFRP2 or pCMV-HA-OSF-2 transfection. Whereas, HA-OSF-2 expressed by Myc antibody immunoprecipitation after pCMV-Myc-SFRP2 and pCMV-HA-OSF-2 co- transfection. Conclusions HA-OSF-2 recombinant vector can express in mammal cells. Interaction exists between HA-OSF-2 and SFRP2.