氧化铁纳米颗粒引起肝血窦内皮细胞损伤及其机制研究

研究二巯基丁二酸修饰的Fe₃O₄纳米颗粒(DMSA-Fe₃O₄)对人原代肝血窦内皮细胞(HHSECs)的作用及对肝脏的影响。利用透射电镜和纳米颗粒追踪分析仪表征纳米颗粒尺寸和表面性质。采用实时无标记细胞分析技术、流式细胞术、实时荧光定量PCR技术和普鲁士蓝染色法等方法,检测0～200 μg/mL浓度范围内DMSA-Fe₃O₄对HHSECs的作用。将DMSA-Fe₃O₄(剂量为1 mg/kg)通过尾静脉注射到小鼠体内,分析肝脏的损伤情况。细胞实验重复数为3,动物实验每组4只动物。结果表明,HHSECs对DMSA-Fe₃O₄的摄取具有浓度和时间依赖性,细胞活性最低降到对照组的37.3%。DMSA-Fe₃O₄引起细胞ROS升高,是对照组的1.41倍;促进HO-1基因表达升高,是对照组的20.8倍;引起HIF-1α基因表达升高,是对照组的2.01倍;促进VEGF基因的表达升高,是对照组的4.2倍。4次静脉注射DMSA-Fe₃O₄后的第2 d,可观察到小鼠肝脏内皮细胞和库普弗细胞有DMSA-Fe₃O₄蓄积,并伴有肝组织染色变浅、少量肝细胞坏死;第158 d时,肝脏中蓄积的DMSA-Fe₃O₄明显减少,肝脏组织恢复正常形态。以上结果显示,HHSECs可大量摄取氧化铁纳米颗粒,引起细胞活性降低和氧化应激损伤。体内多次静脉注射的DMSA-Fe₃O₄可蓄积在肝脏中,伴有肝损伤。随着时间延长,纳米颗粒从肝脏中排出,肝脏损伤可以恢复。

Iron Oxide Nanoparticles Induced Hepatic Sinusoid Endothelial Cells Damage and the Mechanism Research

This work is aimed to investigate the effect of dimercaptosuccinic acid-magnetite nanoparticles (DMSA-Fe₃O₄) on hepatic sinusoid endothelial cells (HHSECs) in vitro and in vivo. Transmission electron microscopy and nanoparticle tracking analysis were applied to characterize the particle size and surface property. Real-time cellular analysis assay, flow cytometry, quantitative real-time PCR (q-PCR) and Prussian blue staining were applied to analyze the effect of DMSA-Fe₃O₄ to HHSECs with the concentration from 0 to 200 μg/mL. DMSA-Fe₃O₄ with the dose of 1 mg/kg was injected into mice through the tail vein to analyze liver damage. For cell experiments, n=3; For animal experiments, n=4. HHSECs engulfed DMSA-Fe₃O₄ in a dose and time dependent manner, with cell viability decreasing to 37.3% of that for control group. DMSA-Fe₃O₄ engulfment increased the intracellular ROS of HHSECs to 1.41 folds. The expression of HO-1 of the cells was significantly increased to 20.8 folds of the control group, the expression of HIF-1α was increased to 2.01 folds of the control group,the expression of VEGF was increased to 4.2 folds of the control group. DMSA-Fe₃O₄ were observed to accumulate in endothelial and Kupffer cells and induced liver damage on the 2nd day after the fourth injection. The amount of DMSA-Fe₃O₄ in the liver decreased significantly on day 158 and the hepatic damage was recovered. The DMSA-Fe₃O₄ nanoparticles was up-taken by HHSECs and induced cell oxidation related injury.In conclusion, multiple intravenous administrations of DMSA-Fe₃O₄ induced nanoparticle accumulation in the liver and its injury, which was recovered as DMSA-Fe₃O₄ being expelled from the liver over time.