Biological Effects Of Metal Oxide Nanomaterials And Carbon Nanotubes To Cells

Along with the rapid development of nanotechnology, more and more nanomaterials have played important roles in various fields due to their unique physicochemical properties. Metal oxide and carbon nanomaterials are the most widely-used two kinds in the nanomaterial family. The metal oxide nanoparticles(NPs) studied in this thesis, including titanium dioxide nanoparticles(Ti O2 NPs), aluminum oxide(Al2O3) NPs and zinc oxide(Zn O) NPs, are widely used in food, medicine, chemical industry and so on; the typical carbon nanomaterial, carbon nanotubes(CNTs), holds great promises in electronic, energy, composite material, information storage and biomedical areas. The production and applications of these metal oxide NPs and CNTs draw great concerns for their bio-safety issue. In this thesis, a systematic investigation of the in vitro behavior of these metal oxide NPs and CNTs was performed to provide meaningful findings for the biosafety assessment of these materials.Firstly, Caco-2 cell and Caco-2 monolayer, which closely relate with the intestine, were chosen as models to study the cytotoxicity, cellular uptake and translocation of metal oxide NPs(Ti O2, Al2O3, Zn O), focusing on the impact of the real situation and physicochemical properties of these particles. Secondly, we investigated the influence of agglomeration on the biological effect of CNTs. Variable sized agglomerates of oxidized multi-walled carbon nanotubes(O-MWCNTs) were prepared by using Ca2+, and the cellular uptake and cytotoxicity of agglomerates in Hela cells were studied. The major achievements of these experiments are presented below:1) Simulating the real situation after the NPs entered the gastrointestinal tract, the cytotoxicity, the cytotoxicity and translocation of two native Ti O2 NPs, and these two Ti O2 NPs pretreated with the digestion simulation fluid or BSA were investigated in undifferentiated Caco-2 cells, differentiated Caco-2 cells and Caco-2 monolayer. The results indicated that Ti O2 NPs did not induce any toxicity in differentiated Caco-2 cells and monolayers, however, Ti O2 NPs pretreated with digestion simulation fluids inhibited the growth of undifferentiated Caco-2 cells at a higher concentration. The cellular uptake of the NPs was influenced by cell differentiation and surface pretreatment. Differentiated cells possessed much lower uptake ability of the pretreated Ti O2 NPs. The cellular uptake of NPs was not the main reason of the cytotoxicity. In addition, the traverse of Ti O2 NPs through the Caco-2 monolayer was negligible. Our findings indicate that the cytotoxicity of Ti O2 NPs and the possibility of Ti O2 NPs traversing through the intestine are quite low.2) For the first time, the bioeffects of Al2O3 NPs to the gastrointestinal in vitro model were studied. Focusing on the influence of size, dose and crystalline phase, we evaluated the cytotoxicity and translocation of four Al2O3 particles(two nano-sizes and the other two submicro-sizes) with different crystalline phases(? and ? phase) on the intestinal cell Caco-2 and the Caco-2 monolayer. We found that the toxicity of particles mainly depended on the dose, but was not closely related with the size and crystalline phase. The ROS generation might be a reason for the toxicity of Al2O3 particles. Slight cytotoxicity could be found at the concentration of 50 μg/m L; with the increasing of doses, Al2O3 particles induced the cell cycle arrest at G1 phase, mitochondrial membrane potential(MMP) decreases, and apoptosis. For the Caco-2 monolayer, the integrity and γ-catenin retained after Al2O3 particle exposures, though cell death was observed. The traverse of the Al2O3 particles through the Caco-2 monolayer was pretty difficult, and the exposure time, doses, crystalline phase and size were not influencing factors. Our results imply that the Al2O3 particles was unlikely absorbed after the oral exposure.3) The cytotoxicity and translocation of Zn O NPs on the in vitro intestinal model Caco-2 monolayer were studied for the first time. Using two Zn O particles(one nano-size and the other submicro-size), we found that the toxicity of Zn O particles mainly depended on the dose, but was not closely related with the size. The toxicity mainly came from the dissolved Zn. The Zn O particles at 18 μg/m L began to induce the cell viability decrease, cell membrane damage, MMP decrease, apoptosis and interleukin-8(IL-8) generation. For the Caco-2 monolayer, the integrity retained after particle exposures, though the Zn O particles induced the slight and clustered cell death. The traverse of both Zn O particles through the Caco-2 monolayer under different conditions was negligible. The results indicate that Zn O particles are difficult to pass through the intestine in vivo.4) The physicochemical properties of CNTs play crucial roles in determining their biological effects. Agglomeration of CNTs is a common phenomenon, however, how agglomeration affects the biological consequence of CNTs has not been well investigated because of its complexity. We prepared variable sized agglomerates of O-MWCNTs by using Ca2+, and studied their cellular uptake and cytotoxicity in Hela cells. We found O-MWCNTs agglomerates could be controlled and adjusted by adding Ca2+. Agglomeration remarkably facilitated the cellular uptake of O-MWCNTs, due to the easy contact of agglomerates with cells. But agglomeration did not induce evident cytotoxicity, which supported by various assays, including cell proliferation, membrane integrity, apoptosis and ROS generation.