C60 衍生物在自微乳药物传递系统中对雷公藤内酯醇诱导的体外细胞毒性的细胞保护作用。

The Cytoprotective Effect of C60 Derivatives in the Self-Microemulsifying Drug Delivery System against Triptolide-Induced Cytotoxicity In Vitro.

机构信息

College of Pharmaceutical Sciences, Zhejiang Chinese Medical University, Hangzhou 311400, China.

出版信息

Molecules. 2024 Aug 28;29(17):4073. doi: 10.3390/molecules29174073.

DOI:10.3390/molecules29174073

PMID:39274920

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11396586/

Abstract

OBJECTIVE

The aim of this study was to optimize the formulation of a C60-modified self-microemulsifying drug delivery system loaded with triptolide (C60-SMEDDS/TP) and evaluate the cytoprotective effect of the C60-SMEDDS/TP on normal human cells.

RESULTS

The C60-SMEDDS/TP exhibited rapid emulsification, an optimal particle size distribution of 50 ± 0.19 nm (PDI 0.211 ± 0.049), and a near-neutral zeta potential of -1.60 mV. The release kinetics of TP from the C60-SMEDDS/TP exhibited a sustained release profile and followed pseudo-first-order release kinetics. Cellular proliferation and apoptosis analysis indicated that the C60-SMEDDS/TP (with a mass ratio of TP: DSPE-PEG-C60 = 1:10) exhibited lower toxicity towards L02 and GES-1 cells. This was demonstrated by a higher IC50 (40.88 nM on L02 cells and 17.22 nM on GES-1 cells) compared to free TP (21.3 nM and 11.1 nM), and a lower apoptosis rate (20.8% on L02 cells and 26.3% on GES-1 cells, respectively) compared to free TP (50.5% and 47.0%) at a concentration of 50 nM. In comparison to the free TP group, L02 cells and GES-1 cells exposed to the C60-SMEDDS/TP exhibited a significant decrease in intracellular ROS and an increase in mitochondrial membrane potential (ΔψM). On the other hand, the C60-SMEDDS/TP demonstrated a similar inhibitory effect on BEL-7402 cells (IC50 = 28.9 nM) and HepG2 cells (IC50 = 107.6 nM), comparable to that of the free TP (27.2 nM and 90.4 nM). The C60-SMEDDS/TP group also exhibited a similar intracellular level of ROS and mitochondrial membrane potential compared to the SMEDDS/TP and free TP groups.

METHOD

Fullerenol-Grafted Distearoyl Phosphatidylethanolamine-Polyethylene Glycol (DSPE-PEG-C60) was synthesized and applied in the self-microemulsifying drug delivery system. The C60-SMEDDS/TP was formulated using Cremophor EL, medium-chain triglycerides (MCT), PEG-400, and DSPE-PEG-C60, and loaded with triptolide (TP). The toxicity and bioactivity of the C60-SMEDDS/TP were assessed using normal human liver cell lines (L02 cells), normal human gastric mucosal epithelial cell lines (GES-1 cells), and liver cancer cell lines (BEL-7402 cells and HepG2 cells). The production of reactive oxygen species (ROS) after the C60-SMEDDS/TP treatment was assessed using 2',7'-dichlorofluorescein diacetate (DCFDA) staining. The alterations in mitochondrial membrane potential (ΔψM) were assessed by measuring JC-1 fluorescence.

CONCLUSIONS

The cytoprotection provided by the C60-SMEDDS/TP favored normal cells (L02 and GES-1) over tumor cells (BEL-7402 and HepG2 cells) in vitro. This suggests a promising approach for the safe and effective treatment of TP.

摘要

目的

本研究旨在优化载有雷公藤内酯醇（C60-SMEDDS/TP）的 C60 修饰自微乳给药系统的配方，并评价 C60-SMEDDS/TP 对正常人类细胞的细胞保护作用。

结果

C60-SMEDDS/TP 表现出快速乳化作用，最佳粒径分布为 50±0.19nm（PDI 0.211±0.049），接近中性的 Zeta 电位为-1.60mV。C60-SMEDDS/TP 中雷公藤内酯醇的释放动力学呈现出持续释放的特征，并遵循拟一级释放动力学。细胞增殖和凋亡分析表明，C60-SMEDDS/TP（TP：DSPE-PEG-C60 的质量比为 1:10）对 L02 和 GES-1 细胞的毒性较低。与游离 TP（L02 细胞为 21.3 nM，GES-1 细胞为 11.1 nM）相比，C60-SMEDDS/TP 的 IC50 更高（L02 细胞为 40.88 nM，GES-1 细胞为 17.22 nM），与游离 TP（L02 细胞为 50.5%，GES-1 细胞为 47.0%）相比，细胞凋亡率更低（L02 细胞为 20.8%，GES-1 细胞为 26.3%）。与游离 TP 组相比，暴露于 C60-SMEDDS/TP 的 L02 和 GES-1 细胞的细胞内 ROS 显著减少，线粒体膜电位（ΔψM）增加。另一方面，C60-SMEDDS/TP 对 BEL-7402 细胞（IC50=28.9 nM）和 HepG2 细胞（IC50=107.6 nM）的抑制作用与游离 TP 相似（27.2 nM 和 90.4 nM）。C60-SMEDDS/TP 组与 SMEDDS/TP 组和游离 TP 组相比，细胞内 ROS 和线粒体膜电位水平也相似。

方法

合成了全氟醇接枝二硬脂酰基磷脂酰乙醇胺-聚乙二醇（DSPE-PEG-C60），并将其应用于自微乳给药系统中。采用 Cremophor EL、中链甘油三酯（MCT）、PEG-400 和 DSPE-PEG-C60 制备 C60-SMEDDS/TP 并负载雷公藤内酯醇（TP）。采用正常人类肝细胞系（L02 细胞）、正常人类胃黏膜上皮细胞系（GES-1 细胞）和肝癌细胞系（BEL-7402 细胞和 HepG2 细胞）评估 C60-SMEDDS/TP 的毒性和生物活性。采用 2'，7'-二氯荧光素二乙酸酯（DCFDA）染色法评估 C60-SMEDDS/TP 处理后活性氧（ROS）的产生。通过测量 JC-1 荧光评估线粒体膜电位（ΔψM）的变化。