Sulf2促进淋巴管生成的实验研究

Role of Sulf2 in Promoting Lymphangiogenesis

目的研究硫酸酯酶2(Heparan sulfate 2,Sulf2)在淋巴管生成中的作用。方法根据Sulf2 cDNA序列,设计并构建pCDH-Flag-Sulf2真核表达载体,慢病毒转染293FT细胞,获得合成纯化的rSulf2。通过流式分析、成管试验、裸鼠耳部淋巴管生成模型等实验,研究rSulf2对于淋巴管内皮细胞(Lymphatic endothelial cells,LECs)细胞周期、凋亡、成管能力和淋巴管生成的影响。结果构建的pCDH-Flag-Sulf2真核表达载体,经酶切和测序证明构建完全正确。Western blot检测证明293FT细胞转染pCDH-Flag-Sulf2真核表达载体后,rSulf2表达增高。合成纯化的rSulf2可以减少无血清培养液诱导的LECs细胞的凋亡,增加活细胞的比例,但对细胞周期无显著影响,rSulf2还可以增加淋巴管内皮细胞的体外成管能力及增加裸鼠耳部淋巴管的数量。结论 Sulf2可以抑制LECs细胞的凋亡,对淋巴管生成起到促进作用。

Objective To explore the role of Sulfatase2 （Sulf2） in promoting lymphangiogenesis. Methods According to Sulf2 cDNA sequence, pCDH-Flag-Sulf2 eukaryotic expression vector was designed and constructed, then transfected into 293FF cell line to obtain and purify rSulf2. The functions of LECs were examined and compared with control group by flow cytometry, matrigel assay and mice ear lymphangiogenesis model to figure out the effects of rSulf2 on cell cycles, apoptosis, tube formation ability and lymphangiogenesis in LECs. Results The construction of pCDH-Flag-Sulf2 vector was proved correctly by restriction enzyme digestion and sequencing test. Sulf2 expression increased in 293FF cells showed by Western blot. Purified Sulf2 reduced no serum medium induced apoptosis and increased the number of living cells, but it had no effect on cell cycles in LECs. Sulf2 also promoted the tube formation ability in vitro and the number of lymphatic vessels in nude mice ear. Conclusion Sulf2 can inhibit LECs apoptosis and promote lymphangiogenesis.