【摘要】 目的 研究表面修饰的超顺磁性四氧化三铁纳米颗粒（以Fe₃O₄NPs@PVP表示）的生物安全性，为磁纳米颗粒的临床应用提供依据。方法 将Fe₃O₄NPs@PVP以0.25、0.5、0.75和1 mg/ml的终浓度加入含有大鼠成纤维细胞的培养基，使用Alarma blue法检测细胞活力；将直径130 nm的Fe₃O₄NPs@PVP配置等体积，浓度分别为2、5、10和25 mg/kg的混悬液备用，将30只大鼠随机分为对照组、低剂量组、中剂量组、高剂量组、极高剂量组，每组6只，对照组经大鼠尾静脉注射0.2 ml生理盐水，其他组经大鼠尾静脉分别注射0.2 ml浓度为2、5、10和25 mg/kg的Fe₃O₄NPs@PVP混悬液，注射完毕后连续观察14天。实验周期结束后，取大鼠腹主动脉血行酶联免疫吸附试验（enzyme-linked immunosorbent assay, ELISA）评估免疫球蛋白IgG水平，并测定丙氨酸氨基转移酶（alanine aminotransferase, ALT）、天门冬氨酸氨基转移酶（aspartate aminotransferase, AST）、总胆红素（total bilirubin, TBIL）、白蛋白（albumin, ALB）、血尿素氮（blood urea nitrogen, BUN）、肌酐（creatinine, Cr）、肌酸激酶（creatine kinase, CK）、乳酸脱氢酶（lactate dehydrogenase, LDH）含量。对大鼠的心脏、肝脏、脾脏、肺脏、肾脏实质器官进行病理学检查。结果 将Fe₃O₄NPs@PVP分别以不同终浓度作用于大鼠成纤维细胞，随浓度的增加，细胞存活率呈正比下降；当Fe₃O₄NPs@PVP终浓度为1 mg/ml时，大鼠成纤维细胞活力值仍有81%。与对照组比较，高剂量组大鼠体内AST水平，极高剂量组大鼠体内AST、BUN、LDH水平均显著升高，组间比较差异均具有统计学意义（P均<0.05）;与极高剂量组比较，高剂量组大鼠的ALT水平，中剂量组大鼠的LDH水平较低，组间比较差异均具有统计学意义（P均<0.05）。与对照组比较，中剂量组大鼠的IgG水平较低（P<0.05）。病理结果显示：对照组大鼠心脏、肝脏、脾脏、肺脏、肾脏各实质器官均保持良好完整性，无明显病变。其他组心肌细胞无肥大、坏死及纤维化；肝小叶结构正常，肝血管无狭窄及堵塞；中剂量组的部分大鼠脾脏切片发现脾脏红髓增生，低剂量组、高剂量组、极高剂量组脾细胞无坏死、炎症、纤维化，脾小结结构正常；肺泡壁无炎症、纤维化等异常情况，肺泡结构正常，肺间质未见弥漫性水肿；肾小球无纤维化，肾小管的结构正常。结论 Fe₃O₄NPs@PVP经静脉注射进入大鼠体内后，大鼠在14天内正常存活，各器官均无明显病变，在极高剂量下仍未产生对大鼠致命性的毒性。该结果表明Fe₃O₄NPs@PVP具有良好的生物安全性，为后续临床实验研究奠定基础。更多还原

【Abstract】 Objective To study the biological safety of superparamagnetic Fe₃O₄ nanoparticles（Fe₃O₄NPs@PVP）, and to provide a basis for the clinical application of magnetic nanoparticles.Methods Fe₃O₄NPs@PVP was added to the culture medium containing rat fibroblast cells at final concentrations of 0.25, 0.5, 0.75 and 1 mg/ml, and cell viability was measured by the Alarma blue assay; Fe₃O₄NPs@PVP with a diameter of 130 nm was prepared in equal volumes at concentrations of 2, 5, 10 and 25 mg/kg as suspensions for later use, thirty rats were randomly divided into control group, low-dose group, medium-dose group, high-dose group and extremely high-dose group, with 6 rats in each, the control group received an intravenous injection of 0.2 ml saline into the tail vein, while the other dose groups received an intravenous injection of 0.2 ml of Fe₃O₄NPs@PVP suspension at concentrations of 2, 5, 10 and 25 mg/kg respectively, and the rats were observed continuously for 14 days after the injections. At the end of the experimental period, blood samples from the abdominal aorta were taken from the rats by enzyme-linked immunosorbent assay（ELISA） to assess immune globulin IgG levels and to measure the contents of alanine aminotransferase（ALT）, aspartate aminotransferase（AST）, total bilirubin（TBIL）, albumin（ALB）, blood urea nitrogen（BUN）, creatinine（Cr）, creatine kinase（CK） and lactate dehydrogenase（LDH）. Pathological examinations were conducted on the heart, liver, spleen, lung and kidney of the rats.Results When Fe₃O₄NPs@PVP was applied to rat fibroblasts at different final concentrations, the cell survival rate decreased directly with the increase of the concentration; when the final concentration of Fe₃O₄NPs@PVP was 1 mg/ml, the viability value of rat fibroblasts was still 81%. Compared with the control group, the levels of AST in rats in the high-dose group and the levels of AST, BUN and LDH in rats in the extremely high-dose group significantly increased, the differences between the two groups were statistically significant（all P<0.05）; compared with the extremely high-dose group, the level of ALT in rats in the high-dose group and the level of LDH in rats in the medium-dose group were lower, the differences between the two groups were statistically significant（all P<0.05）. Compared with contral group, the level of IgG in rats in midium-dose group was lower（P<0.05）. Pathological results showed that the heart, liver, spleen, lung and kidney of rats in the control group maintained good integrity without obvious lesions. Myocardial cells in other groups showed no hypertrophy, necrosis or fibrosis; the structure of hepatic lobules was normal, and there was no narrowing or blockage of hepatic blood vessels; some rats in the medium-dose group showed red pulp hyperplasia in their spleen slices, while splenic cells in low-dose group, medium-dose group, high-dose group and extremely high-dose group showed no necrosis, inflammation and fibrosis, and the structure of splenic nodules was normal; there was no abnormal inflammation or fibrosis in the alveolar walls of the lungs, and the structure of the alveoli was normal, no diffuse edema was observed in the interstitium of the lungs; the glomeruli showed no fibrosis, and the structure of renal tubules was normal.Conclusion After intravenous injection of Fe₃O₄NPs@PVP into rats, the rats survives normally within 14 days, with no significant pathological changes observe in any organs; even at extremely high-doses, Fe₃O₄NPs@PVP does not produce fatal toxicity to the rats. These results indicates that Fe₃O₄NPs@PVP has good biosafety and lays a foundation for subsequent clinical experimental research.更多还原