纳米锰锌铁氧体颗粒对L-02细胞的氧化损伤作用

Oxidative damage of Mn_(0.5)Zn_(0.5)Fe_2O_4 nanoparticles to L-02 cells

目的探讨磁性锰锌铁氧体纳米颗粒(Mn0.5Zn0.5Fe2O4)对人肝细胞株L-02的毒性作用机制。方法 Mn0.5Zn0.5Fe2O4800 mg·L-1作用L-02细胞48 h,透射电镜观察细胞形态及超微结构的变化。Mn0.5Zn0.5Fe2O4200,400和800 mg·L-1作用48 h后,检测L-02细胞内丙二醛(MDA)的含量、超氧化物歧化酶(SOD)和还原型谷胱甘肽(GSH)的活性;荧光染色观察凋亡细胞形态;流式细胞术检测细胞周期及凋亡;荧光定量PCR仪检测胱天蛋白酶3 mRNA表达。结果 Mn0.5Zn0.5Fe2O4800 mg·L-1作用48 h后,纳米颗粒进入细胞内,细胞膜发生破损,细胞器消失,染色体异常聚集。与正常对照组比较,Mn0.5Zn0.5Fe2O4200～800 mg·L-1使细胞内MDA含量逐渐升高,SOD与GSH活性逐渐降低(P<0.05)。Mn0.5Zn0.5Fe2O4可使细胞周期发生改变,G0/G1期细胞百分率有降低的趋势,S期和G2/M期细胞百分率有升高的趋势。Hoechst33258显示明显的细胞凋亡形态。Mn0.5Zn0.5Fe2O4可引起L-02细胞发生剂量依赖性的细胞凋亡,Mn0.5Zn0.5Fe2O4800 mg·L-1作用48 h后,细胞凋亡率达到30.3%,是对照组细胞凋亡率(2.4%)的12.6倍。胱天蛋白酶3 mRNA表达量先增加后降低,但都明显高于正常对照组(P<0.05)。结论 Mn0.5Zn0.5Fe2O4可破坏细胞膜完整性并进入细胞内,诱导细胞发生氧化应激,改变细胞周期,引发细胞凋亡,产生细胞毒性。

OBJECTIVE To explore the toxic mechanisms of Mn0.5 Zn0.5 Fe2O4 nanoparticles on L-02 cells. METHODS Morphological changes were observed by transmission electron micros- copy after L-02 cells were treated with Mn0.5 Zn0.5 Fe2O4 nanoparticles 800 mg- L - 1 for 48 h. Malond- ialdehyde （MDA） content, superoxide dismutase （SOD） and glutathione （ GSH ） activities were determined after cells were exposed to Mn0.5 Zn0.5 Fe2O4 nanoparticles 200,400 and 800 mg. L- 1 for 48 h. Cell cycle and apoptosis were detected by flow cytometry. Morphologic changes were observed by Hoe fluorescence microscopes. Expression of caspase 3 mRNA was analyzed by real time PCR. RESULTS After L-02 cells were treated with Mn0.5 Zn0.5 Fe2O4 nanoparticles 800 mg·L-1 for 48 h, the uhrastructure of cells changed, cell organelles disappeared and the nucleus shrank in size, which served as evidence of apoptosis when nanoparticles went into L-02 cells. Compared with normal control group, MDA content in Mn0.5 Zn0.5 Fe2O4 nanoparticles 200 -800 mg·L- 1 groups significantly increased while GSH and SOD activities significantly decreased （ P 〈 0.05 ）. Compared with normal control group, the percentage in S phase and G2/M phase increased but decreased in G0/G1 phase in Mn0.sZn0.5 Fe204 treated cells. Mn0.5 Zn0.5 Fe2O4 nanoparticles could induce apoptosis in L-02 cells. After cells were exposed to Mn0.5 Zn0.5 Fe2O4 nanoparticles 800 mg. L - 1 for 48 h, the cell apoptosis rate was 30.3%, 12.6 times that in normal control group. Compared with normal control group, the expression of caspase 3 mRNA significantly increased in Mn0.5 Zn0.5 Fe2O4 200 - 800 mg. L - 1 groups （ P 〈 0.05 ）. CONCLUSION ultrastructure of cells, which results in apoptosis in stress. Mn0.5 Zn0.5 Fe2O4 nanoparticles can change the L-02 cells through cell cycles and oxidative stress.