目的 筛选并分离云南重楼水提物中能与血管内皮生长因子(VEGF)蛋白结合的物质, 并探究其对VEGF的刺激作用。方法 采用固相化VEGF亲和柱色谱的方法筛选云南重楼水提物中的VEGF结合因子, HPLC法进一步分离纯化该因子成分, 并采用MTT法以VEGF敏感型细胞株HepG2为模型验证该因子活性, 采用UV、RRLC-QTOF检测, 推测该因子可能的成分组成。结果 从云南重楼水提物中筛选到能与VEGF蛋白结合的因子(命名为因子B³), HPLC分离纯化后因子B³可以促进VEGF敏感型细胞HepG2的增殖, 而对VEGF不敏感型细胞HEK293的增殖没有影响。因子B³和VEGF蛋白对HepG2细胞的增殖促进作用存在叠加效应;而这种效应可以被VEGF抗体阻断;因子B³本身对HepG2细胞的增殖作用也可以被VEGF抗体阻断。通过UV和RRLC-QTOF检测, 推测因子B³可能为皂苷类成分。结论 云南重楼中分离得到的因子B³具有促进VEGF功能的活性。

Objective To screen and isolate vascular endothelial growth factor (VEGF)-binding factors from Paris polyphylla var. yunnanensis, and explore their property for stimulating VEGF activity. Methods The VEGF-binding factors from water extract of P. polyphylla var. yunnanensis were screened by VEGF-affinity chromatography and further purified by HPLC method. Their activity on proliferation of VEGF-dependent cells was determined by MTT analysis with sensitive cell line HepG2 as model. We also predicted the possible components according to RRLC-QTOF and UV results. Results The VEGF-binding factors screened from the water extract of P. polyphylla var. yunnanensis were named as factor B³ which included three compounds and could induce the proliferation of VEGF-dependent cell line HepG2 but not the VEGF-independent cell line HEK293. Further studies indicated that factor B³ had an additive effect with VEGF to induce the proliferation of HepG2, and the additive effect could be attenuated by VEGF antibody. In addition, the proliferation of HepG2 cells induced by factor B³ alone could also be attenuated by VEGF antibody. Furthermore, based on the results of RRLC-QTOF and UV analysis, we predicted that factor B³ are probably members of saponin family. Conclusion The factor B³ isolated from P. polyphylla var. yunnanensis has the property to stimulate VEGF activity.

浙江农林大学科研发展基金(2012FR017);浙江省自然科学基金资助项目(LQ14H280007)