sFGFR3的原核表达及其对乳腺癌细胞增殖的抑制作用

成纤维细胞生长因子(FGFs)是一类高效的细胞分裂、成形促进剂,以及血管生成的诱导物。FGF信号通路是与癌症的发生和发展紧密相关的。可溶性成纤维细胞生长因子受体(sFGFRs)能够与FGFs相结合,从而阻断它们的信号。为了研究sFGFR3对乳腺癌细胞增殖的抑制作用,作者利用从人类胎盘组织中提取的总RNA,通过逆转录聚合酶链式反应技术(RT-PCR),扩增了FGFR3IIIc的胞外域(sFGFR3)。克隆了sFGFR3片段并将其插入到pET3c载体中,然后在大肠杆菌BL21(DE3)中以包涵体的形式进行表达。通过凝胶层析和肝素亲和层析,分离和纯化了成功重折叠的蛋白。sFGFR3的重折叠率为10.2%,纯化后的蛋白其纯度高达95%。3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)处理后的细胞扩增分析结果显示,sFGFR3能够有效抑制乳腺癌细胞BT549的增殖,且其抑制不具有剂量依赖性。

Prokaryotic Expression of Soluble Ectodomain of Human Fibroblast Growth Factor Receptor 3IIIc and its Inhibitory Effect on the Proliferation of Breast Cancer Cells

Fibroblast growth factors (FGFs) are potent mitogens,morphogens,and inducers of angiogenesis. FGF signal-pathway is associated with cancer development and progression. Soluble forms of the fibroblast growth factor receptors (sFGFRs) can bind to FGFs to block their signals. To study the inhibitory effect of sFGFR3 on the proliferation of breast cancer cells,the ectodomain of FGFR3IIIc (sFGFR3) was amplified by extracting total RNA from human placental tissue and using reverse-transcriptase polymerase chain reaction (RT-PCR) . sFGFR3 was cloned into a pET3c vector and expressed in the form of inclusion bodies in Escherichia coli strain BL21 (DE3 ) . Successfully refolded proteins were isolated by gel chromatography and purified by heparin affinity chromatography. The refolded yield of sFGFR3 was 10. 2% ,with purity as high as 95%. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay showed that sFGFR3 was effective on inhibiting the proliferation of BT549 breast cancer cells.