载阿帕替尼金属有机框架纳米颗粒的制备及其体外抗乳腺癌的作用

Preparation of apatinib@zeolitic imidazolate framework⁃8 nanoparticles and its anti⁃breast cancer effect in vitro

目的构建载阿帕替尼的金属有机框架纳米颗粒(apatinib@zeolitic imidazolate framework‑8,A@ZIF‑8),并探讨其对小鼠乳腺癌4T1细胞增殖和凋亡的影响。方法采用物理搅拌法制备沸石咪唑酯骨架结构材料(zeolitic imidazolate framework‑8,ZIF‑8),用一步法将阿帕替尼载入ZIF‑8构建A@ZIF‑8。采用透射电子显微镜(TEM)和扫描电子显微镜(SEM)观察ZIF‑8和A@ZIF‑8的粒径大小及形态特征,激光粒度仪检测Zeta电位值。采用CCK‑8法和流式细胞术检测A@ZIF‑8对4T1细胞增殖及凋亡的影响。从13只Balb/c雌性小鼠中随机选取1只小鼠用于A@ZIF‑8体外溶血试验;其余小鼠随机分为对照组(n=6)和A@ZIF‑8组(n=6),并于首次尾静脉注射后14 d处死,收集血液检测血常规及血液生化指标,HE染色观察心、肝、脾、肺、肾等器官变化,评估A@ZIF‑8生物安全性。结果TEM和SEM均显示,与对照组比较,ZIF‑8载入阿帕替尼后粒径大小增加,Zeta电位明显升高(均P<0.001),表明成功构建A@ZIF‑8。CCK‑8法检测结果显示,A@ZIF‑8对4T1细胞的增殖抑制作用随浓度升高而增强,IC_(50)=27.69μg/mL。流式细胞术检测结果显示,A@ZIF‑8组细胞凋亡率较对照组高(P<0.001)。溶血试验显示,A@ZIF‑8在小鼠体内不易发生溶血。注射A@ZIF‑8溶液后的小鼠血常规、肝功能、肾功能等指标均在正常范围内,小鼠重要器官HE染色在镜下均未发现细胞结构损伤。结论本研究构建的A@ZIF‑8可抑制乳腺癌4T1细胞增殖并促进细胞凋亡,且生物安全性良好,有望用于乳腺癌的体内治疗。

Objective To construct the apatinib@zeolitic imidazolate framework‑8(A@ZIF‑8)nanoparticles,and to investigate the effect of A@ZIF‑8 on the proliferation and apoptosis of mouse breast cancer 4T1 cells.Methods The zeolitic imidazolate framework‑8(ZIF‑8)was prepared by a physical agitation method,the apatinib was loaded into ZIF‑8 by the one‑step method to construct the A@ZIF‑8.The particle size and morphological characteristics of ZIF‑8 and A@ZIF‑8 were observed by transmission electron microscope(TEM)and scanning electron microscope(SEM),the Zeta potentials were measured by laser particle size analyzer.CCK‑8 assay and flow cytometry were used to detect the effects of A@ZIF‑8 on the proliferation and apoptosis of 4T1 cells.One mouse was randomly selected from 13 Balb/c female mice for A@ZIF‑8 in vitro hemolysis experiment.The remaining mice were randomly divided into the control group(n=6)and the A@ZIF‑8 group(n=6),and sacrificed on the 14th day after the first tail vein injection.Blood was collected for blood routine and blood biochemical index detection,and HE staining was used to observe the changes of organs,including heart,liver,spleen,lung,kidney and others,to evaluate the biological safety of A@ZIF‑8.Results Both TEM and SEM showed that compared with the controlgroup,the particle size ofZIF‑8 increased afterloaded with apatinib and the Zeta potentialincreased significantly(allP<0.001),indicating that A@ZIF‑8 was successfully constructed.CCK‑8 assay showed that the inhibitory effect of A@ZIF‑8 on 4T1 cell proliferation increased with the increase of its concentration,IC_(50)=27.69μg/mL.Flow cytometry showed that the apoptosis rate of the A@ZIF‑8 group was higher than that of control group(P<0.001).Hemolysis test showed that the A@ZIF‑8 was not prone to hemolysis in mice.After injection of A@ZIF‑8 solution,the blood routine,liver function,kidney function and other indicators of mice were within the normal range,and no cell structure damage was found in HE staining of mouse vital organs under microscope.Conclusions The A@ZIF‑8 constructed in this study can inhibit the proliferation of breast cancer 4T1 cells and promote cell apoptosis,with good biosafety,which is expected to be used for the treatment of breast cancer in vivo.