Study On The Cytotoxic Effect Of Phytosterol Oxidation Products And Nanosized MRI Contrast Agent

Exogenous compoundsis very common in our life and has a close connection with the the environment. It might cause occurrence of these exogenous compounds related to the role to produce various bioactive substances, which have great research value. When these exogenous compounds into the body, they may produce harmful effect to destroy other enzymes that are important for the body’s normal physiological function or is better for health. Phytosterol oxidation products（POPs） and cholesterol oxidation products（COPs） are constituents of the human diet. There is evidence that cholesterol oxidation products（COPs） may be linked to the initiation and progression of several chronic diseases. However, Thus biological and safety aspects of POPs remain relatively unclear.With the development of nanotechnology in biomedicine, multifunction contrast agent is an important research field in nanotechnology and has extensive applications in medicine. The biological safety of nano-materialshas attracted the attention over the academic circles, however, research on biological activities of nano-contrast agent has been more limited, and the results in some cases have been contradictory to better understand and define exogenous compounds on human health. It is valuable and urgent to confirm biological and safety aspects of different exogenous compounds. The objective of the present study was to systematically compare and evaluate the effects of two types of phytosterol oxides（7-keto- and 7β-OH-phytosterol oxides）and two types of nanoscale contrast agents（Fe₃O₄@HSiO₂ and Gd-FA-Si） towarddifferent mammalian cell types. The main results are as follows:1. For comparisons, the cytotoxic effects of corresponding 7-ketocholesterol（7K-CH） were also assessed. All tested compounds were found to reduce cell viability in a significant and dose-time dependent way, particularly 7K-SI and 7K-CA showed highactivities.The order of 7-ketophytosterol oxide toxicity was 7K-SI > 7K-CA > 7K-BR > 7K-ST > 7K-MIX, 7β-OH- phytosterols oxides was 7β-OH-SI > 7β-OH-CA > 7β-OH-MIX > 7β-OH-ST≈7β-OH–Br in 120μM. In the present study, HIC cells were more sensitive to the 7-ketosterol oxides than7β-OH- phytosterols oxides. Meanwhile, 7K-SI and 7K-CA not only increased early apoptotic cells but caused cell cycle arrest in an accumulation in S phase and a simultaneous decrease of cells engaged in the G1 phase. However, cell death did not appear to be necessarily dependent on the generation of 7K-BR, 7K-ST and 7K-MIX. Furthermore, 7K-SI, 7K-CA and 7K-BR induced apoptosis by caspase-3 activity and the modulatory effects of Bcl-2. However, 7K-ST and 7K-MIX did not involve caspase-3 and Bcl-2.2. Cultured A549 cells exposed to 7-keto phytosterols oxides and 7β-OHphytosterols oxides showed a significant（p<0.05） reduce in the cell activity compartment versus control dose-time dependent way, at all concentration tested after 24 h of incubation, The order of 7-ketophytosterol oxide toxicity was 7K-SI > 7K-CA > 7K-BR ≈ 7K-ST ≈ 7K-MIX, 7β-OH-phytosterols oxides was 7β-OH-SI ≈ 7β-OH-CA > 7β-OH-MIX >7β-OH-ST ≈ 7β-OH-BR in 120μM. ContrarytoHICcells, A549 cells were more sensitive to the7β-OH- phytosterols oxides than 7-ketosterol oxides. Meanwhile, 7β-OH- phytosterols oxides not only increased late apoptotic cells but caused cell cycle in an accumulation in sub-G1 phase. However, cell death did not appear to be necessarily dependent on the chemical structure of phytosterols oxides. Furthermore, 7β-OH–CH, 7β-OH-SI, 7β-OH-CA, 7K-CH increased ROS to induce apoptosis by caspase-3 activity and the modulatory effects of Bcl-2.3. Cultured HepG2 cells exposed to 7-keto phytosterols oxides and 7β-OH-phytosterols oxides showed a significant（p<0.05） reduce in the cell activity versus control dose-time dependent way, at all concentration tested after 24 h of incubation, The order of 7-ketophytosterol oxide toxicity was 7K-SI > 7K-CA > 7K-BR > 7K-MIX > 7K-ST, 7β-OH- phytosterols oxides was 7β-OH-CA > 7β-OH-SI > 7β-OH-BR > 7β-OH-MIX > 7β-OH –ST in 120μM. Contrary to HICcells and A549 cells, administration of the oxysterols mixture yielded results that were contrary to the effects seen by components when administered individually, 7K-MIX were found to reduce cell viability in a significant way. 7β-OH–CA and 7K-CA not only reduce the cell activity but induced cell apoptosis. Furthermore, 7β-OH–CH,7β-OH-SI,7β-OH-CA increasedthe activity of SOD and CAT.Although a significant increase in ROS was assessed after treatment with all tested oxides MDA levels were insignificantly enhanced by 7β-OH–CA and 7K-CA. However, a link between cell death and oxidative stress did not seem to existfor all tested compounds. The different levels of oxide uptake in the cells seem to take a responsible part for the different toxic effects.4. Magnetic iron oxide coated in hydrogenation silica（Fe₃O₄@HSiO₂） is constructedas both a tumor drug carrier and a magnetic resonance（MR） contrastagent. The release performance of COLC can be controlled bypH, as the porous HSiO₂ shell can partially shed at pH below 3.0 to facilitatethe release of COLC. MR imaging（MRI） tests prove that Fe₃O₄@HSiO₂ at pH3.0（H+-Fe₃O₄@HSiO₂） shows a stronger MR contrast enhancement than Fe₃O₄. Cytotoxicity experiment indicates that Fe₃O₄@HSiO₂ has excellent biocompatibility and magnetic targeting performance. Additionally, COLC-loaded Fe₃O₄@HSiO₂（Fe₃O₄ @HSiO₂–COLC） displays a higher inhibition effect on tumor cells under a magnetic field than free COLC. The visibility upon MRI, high targeting, and pH-controlled release characteristics of Fe₃O₄@HSiO₂-COLC are favorableto achieve the aim of reducing side effects to normal tissues, making Fe₃O₄@HSiO₂-COLC an attractive drug delivery system for nanomedicine.5. A pH-responsive nanoplatform, hydroxylated mesoporous nanosilica（HMNS） coated by polyethyleneimine（PEI） coupled with gadolinium and folic acid（FA）（Gd-FA-Si）, was designed to deliver anticancer drug targeting and to promote contrast effect for tumor cells using magnetic resonance（MR） spectrometer. Doxorubicin（DOX） was chosen as the anticancer drug and loaded into nanopores of HMNS, then its release in simulated body fluid could be controlled through adjusting the pH. This nanoplatform could significantly enhance the MR contrast effect. The entire system possessed a high targeting performance to Hela and MDA-MB-231 cells because the FA located in the system could specifically bind to the folate-receptor sites on the surface of cell. Compared with free DOX, the nanoplatform presented a higher cell inhibition effect on the basis of cell assay. Therefore, this nanoplatform could be potentially applied as a tumor-targeted T1 MR contrast agent and pH-sensitive drug carrier system.