载血卟啉单甲醚高分子纳米粒用于光声显像引导下声动力治疗:实验研究

Experimental study of hematoporphyrinmonomethyl ether based PLGA nanoparticles for photoacoustic imaging-guided sonodynamic therapy

目的制备一种载血卟啉单甲醚(HMME)的多功能分子探针,评估其体外光声成像及声动力治疗(SDT)效果。方法采用双乳化法制备载HMME的聚乳酸-乙醇酸(PLGA)纳米粒(HMME@PLGA),观察纳米粒的基本表征和体外增强光声显像情况,通过流式细胞仪检测人乳腺癌MDA-MB-231细胞与HMME@PLGA共孵育后在超声辐照下产生活性氧的能力,并在细胞水平验证其SDT效果。结果所制备的HMME@PLGA纳米粒大小均一,形态规则,平均粒径为(333.67±17.50)nm,表面电位为(-10.57±1.98)mV,包封率为75.62%,载药量为2.90%,浓度为1mg/ml的纳米粒与人乳腺癌MDA-MB-231细胞共孵育24h的细胞存活率为87.21%。随浓度增加,纳米粒光声信号逐渐增强。经超声辐照,HMME@PLGA纳米粒能使人乳腺癌MDA-MB-231细胞产生大量活性氧,并产生明显细胞毒性作用,使细胞大量死亡,Calcein-AM/PI染色呈红色荧光;通过吖啶橙染色观测到细胞溶酶体结构消失。结论本研究成功制备了包裹HMME的高分子纳米粒,可实现体外光声成像引导下的SDT。

Objective To prepare hematoporphyrinmonomethyl ether(HMME)based poly(lactic-co-glycolic acid)(PLGA)nanoparticles,and to investigate their enhancement effect on photoacoustic(PA)imaging and sonodynamic therapy(SDT)efficacy in vitro.Methods HMME-based PLGA(HMME@PLGA)nanoparticles were synthesized using a facile double emulsion strategy.The characteristics and enhanced PA imaging capability were observed in vitro.Upon ultrasound irradiation,the reactive oxygen species(ROS)generated with MDA-MB-231 cells up-taken HMME@PLGA was investigated using flow cytometry.SDT efficacy on cellular level was further investigated.Results The fabricated HMME@PLGA nanoparticles were demonstrated with homogenized size and distribution,and the average diameter was(333.67±17.50)nm.The Zeta potential was(-10.57±1.98)mV.The encapsulation efficiency of HMME in HMME@PLGA was75.62%,and the drug loading was 2.90%.When incubation with HMME@PLGA nanoparticles(1 mg/ml)for 24 h,the viability of MDA-MB-231 cells was 87.21%.PA signal intensities increased with the increase of HMME@PLGA concentration in vitro.Upon ultrasound irradiation,ROS was produced in MDA-MB-231 cells with the assistance of HMME@ PLGA nanoparticles,further leading to cytotoxic effects and cellular death which showed obvious red fluorescence stained with CalceinAM/PI.Meanwhile,the lysosomes structures of the dead cells disappeared when stained by acridine orange.Conclusion HMME@PLGA nanoparticles are successfully fabricated and can achieve PA imaging-guided SDT.