Toxic Effects And Mechanisms Studies Of Lead-Loaded Ultrafine Carbon Black On Red Blood Cells,Nephrocytes And Related Biological Macromolecules

Fine particulate matter(PM2.5)could cause serious harm to human health.This has become a major atmospheric environment and health issue in China.Carbonaceous aerosol is an important part of PM2.5.The incomplete combustion of biomass and fossil fuels produces a large amount of carbon black,which accounts for 20%-50%of the mass concentration of PM2.5.In this study,ultrafine carbon black was selected as the research object.In the process of atmospheric transportation,carbon black is easy to adsorb pollutants,such as heavy metals,organics,etc.,due to its porous,small particle size,and large specific surface area.In this paper,lead,a representative heavy metal pollutant,was adsorbed onto the modified ultrafine carbon black(UFCB).Lead-loaded carbon black(UFCB-Pb)is a typical atmospheric composite particulate matter model.PM2.5 easily enters the blood through alveolar epithelial cells,and then enters the cardiovascular system and the renal system with the systemic circulation.Red blood cells are very susceptible to the influence of exogenous poisons and are important targets for pollutants to enter the body to produce toxicity.There is an urgent need for in-depth study of the interaction between UFCB-Pb and red blood cells.PM2.5 can cause early kidney damage by inducing oxidative stress.The generation of reactive oxygen species(ROS)is the most common mechanism of toxic effects related to ultrafine particles(UFPs),and its mechanism needs further study.In this paper,we investigated the effects of UFCB-Pb on red blood cells and nephrocyte s,and studied the mechanism of the effects of SOD and DNA on UFCB-Pb at the molecular level.In order to comprehensively and in-depth explore the biological toxicity effects of compound pollutants at the cellular and molecular levels in the body.The article is divided into the following seven parts:In the first part,we discussed the health hazards and sources of atmospheric fine particles.It also mentioned the research progress of the toxic effects of ultrafine particles and their compound pollutants.This part also included the interaction with biological macromolecules and the toxic effects and mechanisms on cells and organisms.It clarified the relationship between toxic effects induced by ultrafine particles and oxidative stress.This section summarized the research progress of the toxicity of UFCB-Pb,we put forward the urgent problems to be solved and determined the research plan of the thesis.In the second part,S160(20 nm)UFCB-Pb was selected as the carrier particles of the atmospheric composite particle model.We improved the dispersibility of UFCB in water through multiple hydrogen peroxide oxidation modification methods.The lead ion was loaded on the UFCB by ultrasonic method to prepare the UFCB-Pb model.The elemental analysis confirmed that the lead was effectively loaded on the surface and pores of the UFCB.The lead content measured by atomic spectrophotometry is 76.8μg/mg.The physical and chemical properties of the original CB,modified CB,and UFCB-Pb were determined by solubility experiment,Zeta potential detection,dynamic light scattering particle size analyzer,and scanning electron microscope.The results showed that the UFCB exists as agglomerates in the aqueous solution,and the particle size is much larger than 20nm.The system of UFCB-Pb was more stable than UFCB.The particle size of the aggregates of UFCB-Pb was smaller than UFCB.In the third part,we discussed the effects of UFCB-Pb on the structure of red blood cells,antioxidant capacity.The results showed that the UFCB-Pb has a weak ability to damage the cell membrane of red blood cells within the concentration range of 20 mg/L.It had a little effect on the morphology and structure of red blood cells.UFCB-Pb significantly inhibited the antioxidant capacity of red blood cells and causes lipid peroxidation and oxidative damage to red blood cells.The decline in the function of the red blood cell antioxidant defense system may affect cell metabolism and function.The results of joint exposure experiments showed that the combined exposure of ultrafine carbon black and lead ions caused more significant changes in the content of biomolecules in red blood cells.This indicates that the UFCB and lead ions have a synergistic effect on the antioxidant capacity of red blood cells.In the fourth part,we discussed the toxic effects of UFCB-Pb particles on nephrocytes.This provided reference and data support for evaluating the effect and mechanism of the atmospheric composite particulate UFCB-Pb on the nephrocytes toxicity.One mechanism for the combination of the two to induce the decrease of activity of nephrocytes may be that the particulate matter induces nephrocytes to produce excess reactive oxygen species,resulting in a decrease in the activity of SOD and an increase in the level of lipid peroxidation.This leads to the destruction of the antioxidant system and induces oxidative stress damage to nephrocytes.It also makes DNA oxidative damage and destroys the integrity of the membrane of nephrocytes,which in turn leads to a decline in cell viability.The greater the exposure concentration of pollutants,the more obvious the phenomenon.The oxidative damage of DNA induced by UFCB-Pb in nephrocytes mainly comes from the action of lead.In the fifth part,we studied the interaction mechanism of UFCB-Pb with calf thymus ctDNA.UFCB and UFCB-Pb are directly combined with ctDNA to form UFCB/UFCB-Pb-ctDNA polymer.This affectded the structure of ctDNA and causes fluorescence quenching of the ctDNA system.And it shows a more obvious dose-effect relationship.The combination is an electrostatic interaction combination mode.The presence of lead ions strengthens the electrostatic binding effect,which makes UFCB-Pb cause a greater degree of ctDNA conformation change than UFCB.Due to the combination of static electricity,the phosphate backbone of ctDNA and nitrogen-containing bases have changed.Exposure of particulate matter lead to a decrease in the interaction between bases in ctDNA,and therefore hydrogen bonds and hydrophobic interactions are weakened.In the sixth part,we studied the interaction mechanism of UFCB-Pb with SOD.The results showed that the UFCB-Pb made SOD conformational changes.This made the backbone of the SOD enzyme first compact and then lose,but the overall effect is to shrink.α-helix increased in the secondary structure of SOD.UFCB-Pb changes the microenvironment of the SOD chromophore.Tyr residues will be exposed to make the surrounding environment more hydrophobic.The lead ions in the high-concentration UFCB-Pb and SOD become stronger,and the fluorescence sensitization phenomenon is suppressed.The lead ions released in the high-concentration UFCB-Pb and SOD become stronger,and the fluorescence sensitization phenomenon of the system is inhibited.Part of the lead acetate released by the UFCB-Pb will combine with SOD.Lead binds with SOD and further act on the enzyme activity center,resulting in the enzyme activity decline.In the seventh part,we summarized the main conclusions of the thesis and puts forward the innovative points of the research.In this study,red blood cells,nephrocytes,antioxidant proteins SOD,and DNA were used as objects to systematically study the toxic effects of UFCB-Pb at the molecular and cellular levels.We summarized the structural effects and oxidative stress effects of UFCB-Pb on red blood cells after entering the blood.Our work has found that UFCB-Pb enters into mouse primary nephrocytes and induces cell oxidative damage and gene damage.We clarified the potential molecular mechanisms of DNA and SOD exposure to UFCB-Pb and provided basic data to evaluate their effects at the cellular level.The research results provide new insights for understanding the toxic effects and pathogenic cellular mechanisms of atmospheric composite particles,and may also provide basic data for the prevention and treatment of diseases related to UFCB and its metal ion complexes.