The Safety And Toxicology Experiment And Targeted Experiment In Vivo Of Ang2-siRNA Plasmid Chitosan Magnetic Nanoparticles By Intravenous Administration

ObjectiveInjecting the Ang2- si RNA plasmid chitosan magnetic nanoparticles into the vivo of mice and rats to observe the safety and toxicology of this nanoparticles, and to build the malignant melanoma nude mouse models to observe the targeting of the Ang2- si RNA plasmid chitosan magnetic nanoparticles, which lay the foundation for the further study of targeted inhibition in angiogenesis and tumor growth of malignant melanoma.Methods1、The acute toxicology experiment of the Ang2- si RNA plasmid chitosan magnetic nanoparticlesWe choose the KM mice as the subject in the acute toxicology experiment,by single injecting five concentrations(including 91.6 mg/kg, 152.8 mg/kg, 254.6 mg/kg,424.2 mg/kg, 707.0 mg/kg) of Ang2- si RNA plasmid chitosan magnetic nanoparticles.And the control group injected normal saline.In 14 days, we will observe deaths and the change of the diet and weight of the mice.After the mouse were killed,the colour and morphology of the mouse ’s organs could be observed.And the mouse’s cardiovascular, liver, spleen, lung, and kidney were made to pathological sections for HE stain and Prussian blue stain,which can observe the situation of important organs.2、The repeat-dose study of the Ang2- si RNA plasmid chitosan magnetic nanoparticlesWe choose the SD rats as the subject in the chronic experiment.The rats were divided into four groups,including low, medium and high dose group(35.35 mg/kg,70.70 mg/kg, 353.50 mg/kg)and control group.Control group injected normal saline.All groups would be injected 1ml Ang2- si RNA plasmid chitosan magnetic nanoparticles continuously in 14 days,and then stop drug on the 15 th day,and observe the rats until the 21 th days.During the time,record the usual manifestation and the weight changes of the rats.On the 21 th day,the rats were killed,and taking blood by removalling eyeball to blood routine examination and biochemical test of rats. After the rats were killed,the colour and morphology of the rats’ organs could be observed.And the rats’ cardiovascular, liver, spleen, lung, and kidney were made to pathological sections for HE stain and Prussian blue stain,which can observe the situation of important organs.3、The targeted study in vivo of the Ang2- si RNA plasmid chitosan magnetic nanoparticlesAfter malignant melanoma nude mouse transplantation tumor models were constructed,malignant melanoma cells which were in logarithmic phase would be made cell suspension to inoculate in the right axillary subcutaneous of nude mice.When the subcutaneous tumor grow up to about 6 mm × 6 mm,malignant melanoma nude mouse will be divided into 3 groups at random, including the targeted group,the untargeted group and the blank control group.The nude mice in targered group will be injected 0.4ml Ang2- si RNA plasmid chitosan magnetic nanoparticles and be in the 4000 GS magnetic field cling right axillary subcutaneous. After 60 min,nude mice will be killed.The nude mice in untargeted group will also be injected0.4ml Ang2- si RNA plasmid chitosan magnetic nanoparticles without magnetic field.The nude mice also be killed after 60 min.The nude mice in blank control group will be injected normal saline 0.4ml without magnetic,and the nude mice also be killed after 60 min.And then the tumor tissue will be made to pathological sections for HE stain and Prussian blue stain to test the particles distribution in the organization.ConclusionThe Ang2- si RNA plasmid chitosan magnetic nanoparticles could be the target organ of lungs.But the particles have no influence on the general performance、diet、weight and the fuction of liver and kidney. Its LD50>707.0mg/kg and its non-observed effect dose in a long term >35.35mg/kg.According to the coefficient of equivalent dose conversion method,the non-toxic reaction dosage in human > 222.71mg/kg. d x 14 d(namely 3117.87 mg/kg),which is far higher than the amount needed for clinical. It suggest that the particles are safe in a certain range.The targeted study in vivo of the Ang2- si RNA plasmid chitosan magnetic nanoparticles have confirmed that the particles can move to targeted tissues in an additional magnetic.It suggest that the particles have the characteristics of targeted drug delivery systerm,which can be a reliable and efficient targeted drug deliery carrier and have a great application in the tumor target therapy.At the same time,the particles also lay the foundation of the for the further study of targeted inhibition in angiogenesis and tumor growth of malignant melanoma.