Molecular Mechanisms Of Fe₃O₄ Nanoparticles With Different Sizes Into Cancer Cells

Superparamagnetic magnetic iron oxide nanoparticles ?SPIONs? possess some special characteristics, such as unique magnetic response, low toxicity and side effects, easily to be coupled with antibody/receptor ligands and drug. Based on these excellent performances, superparamagnetic magnetic iron oxide nanoparticles ?SPIONs? are expected to be widely employed to transport and controllable release of drugs, immunoassay, diagnostic imaging, etc. Howerer, the size of SPIONs is close to that of biological macromolecules, which can regulate the metabolism of key proteins. The interactions between them often leads to the great limitation of SPIONs in biomedical applications. Some research has showed that the pathway of SPIONs into cells and the mechanisms of its metabolic regulation is the key to the interaction between them. Furthermore, the biological characteristics and applications of SPIONs strongly depends on their size and surface modification of SPIONs. Therefore, it is very necessary to explore the molecular mechanisms of different sizes of Fe₃O₄ nanoparticles into cancer cells from the level of cell, protein and gene.three different sizes of Fe₃O₄ nanoparticles ?7 nm,11 nm and 14 nm? were prepared in the organic phase under high temperature decomposition. The ligand exchange methods were used to modify the surface of the prepared Fe₃O₄ nanoparticles, and poly ?ethylene glycol? ?PEG? was used as the ligand. And then the hydrodynamic diameter of the three Fe₃O₄ nanoparticles is increased to 15.3 nm,22.9 nm and 30.7 nm in water, corresponding. The human breast cancer cell, namely MCF7 cell, was utilized to establish the cell model between SPIONs and cells. The cytotoxicity of Fe₃O₄ nanoparticles was investigated by MTT method. In addition, inductively coupled plasma-atomic emission spectrometry ?ICP-AES? was used to deeply analyze the uptake behaviors of Fe₃O₄ nanoparticles into the MCF cells. Finally, the endocytosis pathways of Fe₃O₄ nanoparticles into the MCF cells were explored by means of transmission electron microscopy technology, real-time quantitative PCR detecting system and protein-perturbation analysis. The results of this study were as follows:?a? MTT assay showed that polyethylene glycol modified Fe₃O₄ nanoparticles scarcely have cytotoxicity to MCF cells in the concentration range of 20-160 ug Fe/mL; ?b? The uptake amount of the Fe₃O₄ nanoparticles by MCF7 cells was closely associated with particles'size, co-cultivation time and the dosage of SPIONs; ?c? TEM pictures of cells displayed that Fe₃O₄ nanoparticles were mainly gathered in the lysosomes after entering into cells, and 7nm of particles could enter into the nucleus, while llnm and 14nm of particles could not; ?d? qPCR assay showed that SPIONs enter into MCF7 cells mainly through three internalization pathways, including clathrin-mediated endocytosis, caveolin-mediated endocytosis and macropinocytosis; ?e? Protein inhibition assay further verified the results of the qPCR. And the flux of every internalization pathway of Fe₃O₄ nanoparticles of the three sizes was quantitatively determined.