HKa轻链D5区及其合成肽对MDA-MB-231细胞增殖和凋亡的影响

Effect of Synthetic Peptides Derived from HKa D5 on Proliferation and Apoptosis of MDA-MB-231 Cells

目的探讨Hka轻链D5区对肿瘤细胞增殖和凋亡的影响及其发挥活性作用的功能区。方法通过制备HKaD5区重组蛋白(r-HRD)及其D5区合成肽P-5、P-5n、P-5m、P-5c,采用WST法和AnnexinⅤ/PI双标记法检测HKa轻链D5区及其合成肽对MDA-MB-231细胞增殖与凋亡的影响。结果合成肽P-5、P-5n、P-5m能抑制MDA-MB-231细胞增殖,诱导MDA-MB-231细胞凋亡;P-5c无抑制细胞增殖及诱导细胞凋亡的作用。相差显微镜观察细胞形态改变,发现肽P-5、P-5n、P-5m处理组细胞体积缩小、变圆,细胞生长受抑制,活力下降;P-5c处理组与对照组相比无差异。结论合成肽P-5、P-5n、P-5m抑制MDA-MB-231细胞增殖的作用是通过诱导细胞凋亡实现的,组-甘-赖氨酸序列可能是其发挥活性作用的核心序列。

Objective To investigate the effect of domain 5 （D5） of cleaved high molecular weight kininogen （HKa） on proliferation and apoptosis of tumor cells, and the function domain for its activity. Methods Recombinant HK domain 5 and synthetic peptides P-5, P-5n, P- 5m and P-5c were prepared. The effect of synthetic peptides derived from HKa D5 on proliferation and apoptosis of MDA-MB-231 cells were detected by WST colorimetrical assay and Annexin V-FTTC/PI double label method. Results Compared with the control group, the cell number decreased significantly with the increase of r-HRD ,P-5 ,P-5n and P-5m,which induced apoptosis in MDA-MB-231 cells, and P-5c did not show inhibitory activity. The morphological changes of MDA-MB-231 cells, such as shrinking and turning round,were observed under the inverted phase contrast microscope after treated with P-5n and P-5m for 24 hours;P-5c did not show inhibitory activity. Conclusion HK r-HRD and peptides derived from HKa D5 can inhibit MDA-MB-231 cell proliferation by inducing apoptosis and the amino acid sequence His-Gly-Lys （HGK） in D5 is the core motif for inhibitory activity.