目的 构建pH敏感/叶酸修饰白藜芦醇脂质体(pH-sensitive/folic acid-modified resveratrol liposomes,pH/FA-Res-Lips),并进行理化性质和抗肿瘤评价。方法 单因素法考察pH/FA-Res-Lips主要影响因素,使用Box-Behnken设计-效应面法(Box-Behnken design-response surface method,BBD-RSM)优化pH/FA-Res-Lips处方工艺。采用紫外-可见光吸收光谱(ultraviolet-visible absorption spectroscopy,UV)、傅里叶变换红外光谱法(Fourier transform infrared spectroscopy,FT-IR)、透射电子显微镜(transmission electron microscope,TEM)进行表征,考察pH/FA-Res-Lips在不同pH值磷酸盐缓冲液(PBS)中的释药行为。采用MTT法和Annexin V/PI双染法考察pH/FA-Res-Lips对人子宫内膜癌Ishikawa细胞的生长抑制及促凋亡作用。采用Ishikawa细胞建立子宫内膜癌荷瘤小鼠模型,分为模型对照组(生理盐水)、阳性对照组(顺铂,2.0 mg/kg)、白藜芦醇组(15 mg/kg)、白藜芦醇脂质体组(Res-Lips,15 mg/kg)、pH-Res-Lips组(15 mg/kg)、FA-Res-Lips组(15 mg/kg)、pH/FA-Res-Lips低剂量组(10 mg/kg)和pH/FA-Res-Lips高剂量组(15 mg/kg),比较pH/FA-Res-Lips体内抗肿瘤效果。通过血液生化指标和小鼠主要脏器HE染色来评价pH/FA-Res-Lips安全性。结果 pH/FA-Res-Lips最佳处方为磷脂与白藜芦醇用量比为5.80∶1,磷脂与胆固醇用量比为5.00∶1,磷脂与DSPE-PEOz-FA用量比为5.30∶1。pH/FA-Res-Lips平均包封率、载药量、平均粒径和ζ电位分别为(85.74±1.09)%、(8.77±0.11)%、(192.25±5.38)nm和(-28.12±1.20)mV。pH/FA-Res-Lips未影响白藜芦醇化学结构,白藜芦醇与脂质材料之间存在氢键络合作用。pH/FA-Res-Lips外貌呈囊泡状。pH/FA-Res-Lips在pH 5.5、6.0 PBS中累积释放度较高,具有明显的pH敏感性。MTT法结果显示,白藜芦醇和pH/FA-Res-Lips对Ishikawa细胞半数抑制浓度(IC₅₀)分别为98.14、48.43 μg/mL,并有效诱导细胞凋亡。pH/FA-Res-Lips显著抑制了子宫内膜癌荷瘤小鼠肿瘤生长,体质量未见显著性下降,pH/FA-Res-Lips(15 mg/kg)将白藜芦醇(15 mg/kg)抑瘤率由17.35%提高至66.67%。pH/FA-Res-Lips未对荷瘤小鼠各个脏器产生影响,使用安全性较高。结论 pH/FA-Res-Lips显著性提高了白藜芦醇抗肿瘤作用,为进一步开发成抗肿瘤药物提供了实验依据。

Objective The pH-sensitive/folic acid-modified resveratrol liposomes (pH/FA-Res-Lips) was prepared, its physicochemical properties and antitumor effects were also studied. Methods The main influencing factors of pH/FA-Res-Lips were investigated by single factor experiments, Box-Behnken response-surface design method (BBD-RSM) was employed to optimize prescriptions of pH/FA-Res-Lips. pH/FA-Res-Lips was characterized by ultraviolet-visible absorption spectroscopy (UV), Fourier transform infrared spectroscopy (FT-IR) and transmission electron microscopy (TEM). Drug release behavior of pH/FA-Res-Lips in phosphate buffer saline solution (PBS) with different pH values was investigated. MTT method and Annexin V/PI double staining method were used to investigate the inhibitory and proapoptotic effects of pH/FA-Res-Lips on Ishikawa cells, respectively. Ishikawa cells were used to establish the mouse model of endometrial cancer, and divided into model control group (normal saline), positive control group (cisplatin, 2.0 mg/kg), resveratrol group (15 mg/kg), resveratrol liposomes (Res-Lips, 15 mg/kg) group, pH-sensitive resveratrol liposomes (pH-Res-Lips, 15 mg/kg) group, FA-modified resveratrol liposomes (FA-Res-Lips, 15 mg/kg) group, low-dose pH/FA-Res-Lips (10 mg/kg) group and high-dose pH/FA-Res-Lips (15 mg/kg) group, the in vivo antitumor effects of pH/FA-Res-Lips were compared. Safety evaluation of pH/FA-Res-Lips was performed by blood biochemical indexes and HE staining of the main organs of mice. Results Optimal formulation of pH/FA-Res-Lips nanoparticles as follows: phospholipids to resveratrol ratio was 5.80:1, phospholipids to cholesterol ratio was 5.00:1, and phospholipids to DSPE-PEOz-FA ratio was 5.30:1. Average envelopment efficiency, drug loading, particle size and ζ potential were (85.74 ± 1.13)%, (8.77 ± 0.16)%, (192.25 ± 7.61) nm and (-28.12 ± 1.20) mV, respectively. pH/FA-Res-Lips did not affect the structure of resveratrol, and there was a hydrogen bond complexation between resveratrol and lipid materials. The appearance of pH/FA-Res-Lips was vesicular. pH/FA-Res-Lips existed higher cumulative release in PBS of pH 5.5 and 6.0, which demonstrated that pH/FA-Res-Lips had obvious pH-sensitivity. MTT method results showed that IC50 of resveratrol and pH/FA-Res-Lips on Ishikawa cells were 98.14, 48.43 μg/mL, respectively, and effectively induced apoptosis of Ishikawa cells. pH/FA-Res-Lips significantly inhibited the tumor growth of endometrial cancer-bearing mice, and there was no significant decrease in body weight. pH/FA-Res-Lips (15 mg/kg) increased the tumor inhibition rate of resveratrol (15 mg/kg) from 17.35% to 66.67%. pH/FA-Res-Lips did not affect the organs of tumor-bearing mice, which showed that the safety was relatively high. Conclusion pH/FA-Res-Lips significantly enhanced the anti-tumor efficacy of resveratrol, which had the potential to be developed into an antitumor drug.

R283.6

2025年度河南省科技攻关项目（252102310468）；河南省高等学校重点科研项目计划（23B320013）；河南应用技术职业学院2023年度校级课题（2023-K.J-54）；河南省医学科技攻关计划（SBGJ202102153）