纳米活性炭,纳米二氧化硅和纳米二氧化钛对人胃肿瘤BGC-823细胞的毒性作用

Cytotoxic effects of activated carbon nanoparticles,silicon dioxide nanoparticles and titanium dioxide nanoparticles on human gastric carcinoma cell line BGC-823

目的探讨不同化学组成的纳米颗粒对人胃癌BGC-823细胞的毒性作用及其机制。方法分别以纳米活性炭(ACNP)、纳米二氧化硅(SiO2)和纳米二氧化钛(TiO2)100,200,400,800和1600mg·L-1悬液作用BGC-823细胞24,48和72h,MTT法检测细胞增殖,比色法检测乳酸脱氢酶(LDH)漏出量。ACNP100mg·L-1,纳米SiO2200mg·L-1,纳米TiO2200mg·L-1作用BGC-823细胞24h,透射电镜观察细胞形态及超微结构的影响。纳米SiO2和纳米TiO2100,200,400mg·L-1作用细胞24h后,AnnexinⅤ-FITC/PI双染法检测细胞凋亡。ACNP、纳米SiO2和纳米TiO2100,200mg·L-1作用细胞48h后,用PI染色法检测细胞周期。结果 ACNP,纳米SiO2和纳米TiO2均能明显抑制BGC-823细胞的增殖,作用72h后的IC50分别为874.2,676.2和883.5mg·L-1。与正常对照组相比,纳米SiO2100～800mg·L-1组LDH漏出量均显著升高,并呈浓度依赖性(r=0.9751,P<0.01),而纳米TiO2100mg·L-1作用细胞24h,LDH漏出量与对照组相比没有显著差异,但随着作用浓度增加和作用时间延长,各组LDH漏出量明显高于对照组(P<0.05)。ACNP100mg·L-1作用24h后,细胞出现细胞质浓缩、细胞核固缩和裂解。纳米SiO2200mg·L-1和纳米TiO2200mg·L-1作用24h后均出现细胞坏死。纳米颗粒ACNP,SiO2和TiO2作用组均可见纳米颗粒进入细胞及线粒体损伤。纳米SiO2100mg·L-1和纳米TiO2100mg·L-1作用24h,细胞坏死率与正常对照组(4.59±1.20)%相比显著升高(P<0.01),分别为(39.40±1.72)%和(14.12±0.90)%(P<0.05);细胞凋亡率与对照组相比没有显著差异。ACNP,纳米SiO2和纳米TiO2100和200mg·L-1作用细胞48h后,S期细胞增多,G0/G1期细胞减少,细胞碎片增多;ACNP组亚二倍体细胞增多。结论 ACNP、纳米SiO2和纳米TiO2能够抑制BGC-823细胞的增殖。ACNP可诱导细胞凋亡。纳米SiO2和纳米TiO2能损伤细胞膜,造成以细胞坏死为主的毒性损伤。

OBJECTIVE To explore cytotoxic effects of activated carbon nanoparticles(ACNP),silicon dioxide nanoparticles(nano-SiO2) and titanium dioxide nanoparticles(nano-TiO2) on the human gastric carcinoma cell line BGC-823.METHODS BGC-823 cells were exposed to ACNP,nano-SiO2 and nano-TiO2 100,200,400,800 and 1600 mg·L-1 for 24,48 and 72 h.The inhibitory rate was observed by MTT assay.The lactate dehydrogenase(LDH) leakages in the culture medium were determined.The morphological changes were observed by transmission electron microscopy after cells were treated with ACNP 100 mg·L-1,nano-SiO2 200 mg·L-1 or nano-TiO2 200 mg·L-1 for 24 h.The apoptotic rate was detected by Annexin Ⅴ-FITC/PI staining with flow cytometry after cells were treated with nano-SiO2 or nano-TiO2 100,200,400 mg·L-1 for 24 h.The cell cycle was detected by PI staining after cells treated with ACNP or nano-SiO2 or nano-TiO2 100 and 200 mg·L-1 for 48 h were collected.RESULTS ACNP,nano-SiO2 and nano-TiO2 significantly inhibited the proliferation of BGC-823 cells.The 50% inhibitory concentrations of ACNP,nano-SiO2 and nano-TiO2 after 72 h were 874.2,676.2 and 883.5 mg·L-1,respectively.The LDH leakages in nano-SiO2 100-800 mg·L-1 groups significantly increased in a concentration-dependent manner(r=0.9751,P<0.01).The LDH leakage of nano-TiO2 100 mg·L-1 was not higher than that in normal control group.The LDH leakage in nano-TiO2 groups increased with the concentration and time.Condensation of the cytoplasm,condensation and fragmentation of the nuclear chromatin were observed in ACNP 100 mg·L-1 for 24 h.The cells treated with nano-SiO2 or nano-TiO2 200 mg·L-1 for 24 h exhibited necrosis.Mitochondrial damage was observed in the cells treated with ACNP,nano-SiO2 or nano-TiO2.The necrotic rate of nano-SiO2 and nano-TiO2 100 mg·L-1 was(39.40±1.72)% and(14.12±0.90)%,respectively,which were significantly higher than that of the control group(4.59±1.20)%(P<0.05).The percentage of cells at S phase of ACNP,nano-SiO2 and nano-TiO2 100 and 200 mg·L-1 groups was higher than that in control group,while the percentage of G0/G1-phase cells decreased.The percentage of 'sub-G1' cells significantly increased in ACNP groups.CONCLUSION ACNP,nano-SiO2 and nano-TiO2 could inhibit proliferation of BGC-823 cells in vitro.ACNP could induce apoptosis of BGC-823 cells.Nano-SiO2 and nano-TiO2 could directly induce necrosis of BGC-823 cells by disintegrating plasma membrane.