Wang Yujuan, Jiang Liangqian, Yin Qifeng, Liu Huihui, Liu Guoxiang, Zhu Guoteng, Li Bing
Department of genetics and cell biology, Basic medical college, 308 Ningxia Road, Qingdao University, Qingdao, China, 266071.
Affiliated Hospital of Qingdao University, Qingdao, China, 266000.
J Cancer. 2017 Aug 25;8(15):3001-3013. doi: 10.7150/jca.21059. eCollection 2017.
The novel C-PC/CMC-CD59sp-NPs were made by carbocymethyl chitosan (CMC) loading C-phycocyanin (C-PC) with the lead of CD59 specific ligand peptide (CD59sp) for targeting, and the characteristics and targeted anti-tumor mechanism were explored in order to realize the targeted therapy of C-PC on the growth of HeLa cells both and . The targeting nanoparticles were synthesized by ionic-gelation method, and the optimal condition was selected out by orthogonal analysis. The properties of nanoparticles were observed by laser particle analyzer and dynamic light scattering (DLS) and Fourier Transform Infrared Spectrometer (FTIR). The effects of nanoparticles on the proliferation of HeLa cells were assessed by MTT assay. The mice model with tumor was constructed by subcutaneous injection of HeLa cells into the left axilla of NU/NU mice. The weight of tumor and the spleen were tested. The expression quantities of cleaved caspase-3, Bcl-2 were determined by western blot and immunofluorescent staining. Results showed the morphology of the finally prepared nanoparticles was well distributed with a diameter distribution of 200±11.3 nm and zeta potential of -19.5±4.12mV. Under the guidance of CD59sp, the targeting nanoparticles could targetedly and efficiently arrive at the surface of HeLa cells, and had obvious inhibitory effect on HeLa cells proliferation both in vitro and vivo. Moreover, the nanoparticles could induce cell apoptosis by up-regulation of cleaved caspase-3 proteins expression, but down-regulation of Bcl-2 and cyclinD1 proteins. Our study provided a new idea for the research and development of marine drugs, and supplied a theoretical support for the target therapy of anticancer drug.
新型C-PC/CMC-CD59sp-NPs是通过羧甲基壳聚糖（CMC）负载藻蓝蛋白（C-PC），并以CD59特异性配体肽（CD59sp）为导向制备而成，旨在探索其特性和靶向抗肿瘤机制，以实现C-PC对HeLa细胞生长的体内外靶向治疗。通过离子凝胶法合成靶向纳米颗粒，并通过正交分析筛选出最佳条件。利用激光粒度分析仪、动态光散射（DLS）和傅里叶变换红外光谱仪（FTIR）观察纳米颗粒的性质。采用MTT法评估纳米颗粒对HeLa细胞增殖的影响。将HeLa细胞皮下注射到NU/NU小鼠的左腋窝构建荷瘤小鼠模型，检测肿瘤重量和脾脏重量。通过蛋白质免疫印迹法和免疫荧光染色法测定裂解的caspase-3、Bcl-2的表达量。结果显示，最终制备的纳米颗粒形态分布均匀，直径分布为200±11.3 nm，zeta电位为-19.5±4.12 mV。在CD59sp的引导下，靶向纳米颗粒能够靶向并有效地到达HeLa细胞表面，对HeLa细胞的体外和体内增殖均有明显的抑制作用。此外，纳米颗粒可通过上调裂解的caspase-3蛋白表达，下调Bcl-2和细胞周期蛋白D1蛋白诱导细胞凋亡。本研究为海洋药物的研发提供了新思路，为抗癌药物的靶向治疗提供了理论支持。
