Dissertation
Dissertation > Medicine, health > Pharmacy > Pharmacy > Pharmaceutics

Preliminary Study of Nanotoxicity about Amphiphilic Graft Polyphosphazenes

Author Chen
Tutor QiuLiZuo
School Zhejiang University
Course Pharmaceutical preparations
Keywords amphiphilic graft polyphosphazenes poly(ethylene glycol) biocompatibility nanotoxicology apoptosis ROS generation
CLC R943
Type Master's thesis
Year 2012
Downloads 9
Quotes 0
Download Dissertation

In recent years, amphiphilic graft polyphosphazenes have been proved to be promising delivery vehicles for drug targeting and anti-tumor therapy. However, the data of nano-biocompatibility and nanotoxicity assessment for these nanomaterials still remain unclear. We synthesized via thermal ring-opening polymerization and nucleophilic substitution reaction of methoxy-poly(ethylene glycol) (PEG), N,N-diisopropylethylenediamine (DPA) and serine methyl ester (SME). FTIR and 1H NMR studies have confirmed that the copolymers were synthesized successfully. The copolymers were able to be self-assembled in aqueous solution to form nanoparticles and the averaged particle size of spherical polymeric nanoparticles was 130-150 nm with a narrow distribution.In order to evaluate their biocompatibility in bloodstream, the plasma protein adsorption and phagocytic uptake behavior in human THP-1 macrophages were performed. The results suggested only a minor percentage of the nanoparticles were involved in BSA binding and phagocytic uptake as the result of PEGylation on the particulate surface. The copolymer revealed strong capabilities of anti-opsonophagocytosis. The results of comparing the plasma protein adsorption behavior of two different polymeric nanoparticles- PDS-BOC and PDS-NH2 indicated that the positively charged nanoparticles had relatively more protein adsorption and phagocytic uptake rate. The FITC labeled PDS was applied to determine the biodistribution in plasma and different tissues of MCF-7 bearing mice using spectrofluorophotometer. The results showed a clear cumulative effect of copolymer in tumor site, a prolonged circulation time in blood and high amounts of retention in liver and kidney.To determine the nanotoxicity as well as the molecular mechanism of cell damage on human normal hepatic cell line L-02, we took two kind of polymeric nanoparticles-PDS-NH2 and PDS-BOC for comparison, the former was with -NH2 exposure and the later was linked with tertbutyloxycarbonyl (BOC) groups to protect and hide -NH2 group. We measured cell viability, apoptosis and necrosis, reactive oxygen species (ROS) generation, the loss of mitochondrial membrane potential and the levels of the apoptotic signaling proteins after the cells being exposed to polymeric nanoparticles with different concentrations (0.1,0.2,0.5 mg/ml) for 24 h. Our data indicated that the two nanoparticles induced cytotoxicity in a dose dependent manner; only at high dosage level (0.5 mg/ml) the distinct cytotoxicity was observed. In addition, the increased necrotic rate of PDS-NH2 with high dosage indicated the NH2 group not only induced cell apoptosis but also increased cell necrosis to a certain extent, which was a possible explanation for the greater cytotoxicity caused by PDS-NH2 when compared with PDS-BOC. The increased generation of reactive oxygen species (ROS), the loss of mitochondrial membrane potential, the decreased ATP level and the increased activities of caspase-3 and caspase-9, all suggested that the polymeric nanoparticles triggered cellular apoptosis through mitochondria-dependent pathways in L-02 cells. The experiments presented here could provide an in vitro nanotoxical evaluation platform for other promising polymeric nanomaterials.Again, we took PDS-NH2 and PDS-BOC to determine the nanotoxicity and the cell damage mechanism on human proximal tubular (HK-2) cells, we measured cell viability, apoptosis and necrosis, reactive oxygen species (ROS) generation, the loss of mitochondrial membrane potential and the levels of the apoptotic signaling proteins in HK-2 cells after the cells being exposed to nanoparticles of different concentrations (0.1,0.2,0.5mg/ml) for 24 h. Our data indicated the two nanoparticles induced cytotoxicity in a dose dependent manner; PDS-NH2 caused more cytotoxicity than PDS-BOC. The polymeric nanoparticles triggered cellular apoptosis through the extrinsic pathways and intrinsic pathways.

Related Dissertations
More Dissertations