法氏囊活性五肽BP5的抗肿瘤作用

Antitumor mechanism of Bursopentin (BP5)

摘要:

摘要:【目的】法氏囊活性五肽Bursopentin (BP5)是一种多功能生物学活性因子，不仅具有免疫调节和抗氧化功能，还具有抗肿瘤活性。目前，BP5发挥抗肿瘤生物功能的作用机制尚不清晰。【方法】本研究利用T7噬菌体展示技术构建鸡DT40细胞T7噬菌体cDNA文库，以BP5-BSA为靶分子对文库进行亲和淘选，通过序列测定和生物信息学分析，获得BP5的结合蛋白。通过分析BP5诱导的鸡DT40细胞mRNA基因表达谱和检测BP5对HCT116细胞中p53 Luciferase活性和p53信号通路中相关蛋白表达的影响来探讨BP5抗肿瘤的作用机制。【结果】构建的鸡DT40细胞T7噬菌体cDNA文库原始滴度为1.2×104 pfu/mL，重组率为91.7%，扩增后滴度为1.9×109 pfu/mL;通过生物淘选获得3条与BP5结合的鸡源蛋白血管内皮生长因子A(vascular endothelial growth factor A)、细胞周期蛋白E1(cyclin E1)和运输蛋白颗粒复合体2(trafficking protein particle complex 2)。基因表达谱分析结果显示，BP5参与调节多条与抗肿瘤功能有关的通路，其中包括p53信号通路，且p53信号通路涉及的5个差异表达基因均与细胞周期调控或抗肿瘤相关，其中SIAH1、 TP53(p53)、CIP1(p21)基因被上调，CHEK2、CCNE1基因被下调。RT-PCR方法检测10个表达水平有明显变化的差异基因，其结果与基因芯片分析结果相符;BP5对p53 Luciferase 活性的影响实验结果显示，BP5在0.2－20μg/mL 浓度下能够明显提高HCT116细胞中p53相对Luciferase活性，Western blotting结果证实，BP5能显著促进p53、p21蛋白的表达水平，而CCNE1和CHEK2蛋白的表达水平则明显降低。【结论】本研究结果表明BP5通过p53信号通路途径发挥其抗肿瘤生物学功能，为进一步研究BP5抗肿瘤具体的分子机制提供了参考依据。

Abstract:

Abstract:［Objective］Bursopentin (BP5) is a multi-functional bioactive peptide with functions of immunomodulatory，antioxidant and antitumor. However，the antitumor mechanism of BP5 is still unclear.［Methods］We constructed T7 phage cDNA library of DT40 cells，and the proteins interacted with BP5 were identified. Then，the expression profile of BP5-treated DT40 cells were analyzed using gene microarray，p53 Luciferase activity was detected.［Results］The results of the expression profiling revealed that BP5 regulated expression of 1078 genes，of which 537 were up-regulated and 541 were down-regulated．Differentially expressed genes involved in various pathways were identified，of which 25 pathways were associated with immune responses and tumorigenic processes，including the p53 signaling．Furththmore，BP5 significantly enhanced p53 luciferase activity and stimulated expression of p53 protein in HCT116 cells.［Conclusion］These results suggest that BP5 exerted antitumor activity through p53 signaling and that this study provides novel insights on the antitumor mechanism of BP5.