纳米银诱导的以免疫毒性为主的早期全身毒性研究

Abstract

目的：考察纳米银对大鼠免疫系统的影响以及其他早期全身毒性反应；同时考察纳米银在体外对淋巴细胞炎性因子和免疫因子释放的影响。通过体内外试验的综合研究考察纳米银的潜在免疫系统毒性。方法：SD大鼠经连续灌胃给入低剂量(1 mg/kg)和高剂量(10 mg/kg)纳米银溶液，生理盐水溶液(对照组)。于首次给入后2小时、连续给入7天、14天及停止给入14天后分别处死大鼠，然后进行大鼠体内各脏器银元素定量测定和病理学分析、血液学及血液生化检测、相关免疫因子的检测。体外试验中，在纳米银暴露或非暴露条件下，将淋巴细胞于含或不含脂多糖的培养液中培养24小时，分析IL-6、IL-4、IgG、IgE等的含量，以考察纳米银引起的免疫系统毒性。结果和讨论：纳米银暴露组显示纳米银剂量、时间依存性的在大鼠血液及脏器组织内蓄积。与对照组相比，经口给入高剂量纳米银(10 mg/Kg)引起了大鼠淋巴细胞IgE、IgG与IL-4的显著增加；高、低剂量组给入纳米银14天停止给入14天后，血清IL-6水平与B细胞CD45RA+水平均显著增加。虽然经纳米银灌胃后大鼠组织病理学检查未发现显著的病理改变，但血糖和碱性磷酸酶的升高提示有胰脏和肝脏的损伤。血常规显示血液内银在低浓度时刺激炎性细胞反应性增加；而高浓度时则诱导细胞毒性，抑制了淋巴细胞的正常生长。在体外不同浓度纳米银(0.5 μg/mL，5 μg/mL)作用下，LPS诱导的淋巴细胞分泌IgG和IL-6显著减少。综合体内外实验结果表明，本研究所用的纳米银主要引起了大鼠以Th2型免疫毒性为主的早期全身毒性。 Objective: To investigate the effects on immune system and other systemic toxicity of silver nano-particles (SNPs) in rats by oral exposure, and combined an in vitro study of SNPs on the release of inflammatory cytokines and immune factors in lymphocytes, to get a better understanding about potential immunotoxicity of silver nanoparticles. Methods: SD rats were orally administrated with low-dose (1 mg/kg) and high-dose (10 mg/kg) of SNPs and saline (control) respectively, and the organ and blood samples were harvested after 2 hrs (n = 9), 7 days (n = 18), 14 days (n = 18) ex-posure and14 days of post-exposure (n = 9) for determination of silver content, hematology, blood biochemistry, and immune factor assessment. Lymphocytes were incubated with or without lipo-polysaccharide in the presence or absence of silver nanoparticles for 24 hours in vitro. And IL-6, IL-4, IgG and IgE of cell culture supernatants were analyzed by ELISA. Results and Discussion: SNPs exposure group showed SNPs accumulated time-dose dependently in blood and organs of rats. The IgG (14 days), IgE (14 days) and IL-4 (7 days) of rats after 10 mg/kg SNPs exposure were signifi-cantly increased, and IL-6 and CD45RA+ (B cells) at 14 days post-exposure of SNPs were obviously increased, compared with that in control. But we did not find significant pathological changes. The rise of glucose and ALP suggested damage of pancreas and liver of rats after SNPs oral administra-tion. Low concentration silver in blood increased inflammatory cells in response to the stimuli by blood examination, while high concentration induced cytotoxicity which inhibited the normal growth of lymphocyte. Secretion of IgG and IL-6 induced by LPS of lymphocytes in low-dose and high-dose (0.5 μg/ml, 5 μg/ml) SNPs exposure was inhibited in vitro. In conclusion, our study showed that silver nanoparticles mainly caused a Th2-type immune toxicity-based early systemic toxicity.