Toxic Effect And Mechanism Of Ultrafine Carbon Black Loaded With Lead On CAT,SOD And Mouse Hepatocytes

In recent years,the haze pollution is more and more serious.As a major component of haze,the impact of ultrafine particulates(UFPs)on health has also attracted extensive attention from the society.UFPs are widely distributed and can be contacted with the human body through the respiratory tract.It can also enter the cell and affect the structure and functional expression of proteins,thus inducing acute pneumonia,cardiovascular and cerebrovascular damage,immune system damage ect.The composition of UFPs is complex,and its toxic effect is the result of the interaction of various components.Currently,there is a lack of discussion and research on the toxic effect mechanism of each component of UFPs.Ultrafine carbon black(UFCB)has similar physical and chemical properties with UFPs and can be used as an alternative model for studys of UFPs.Lead is easily adsorbed by UFPs,and the UFPs loaded with lead widely exist in the atmosphere,which brings new problems to human health.At present,toxicological studies on UFPs mainly focus on the effect of single UFCB particles on respiratory system cells.The mechanism of multi-component UFPs acting on functional proteins and organs to induce diseases is still lacking.Oxidative stress is the cause of many diseases.The liver shoulders the important task of the body's oxidative defense.Catalase(CAT)and superoxide dismutase(SOD)play important roles in protecting the body from oxidative stress.In this study,mouse primary hepatocytes,CAT and SOD were selected as targets,and UFCB was selected as the replacement model of UFPs.The effects and mechanism of UFCB and lead-loaded UFCB induced oxidative stress were compared at the cellular and molecular levels.This paper mainly includes the following parts:(1)The present situation of atmospheric particulate matter,UFPs,UFCB and lead pollution and the progress of toxicology research at the present stage were reviewed.The shortcomings of the evaluation of the toxicity mechanism of lead-loaded UFPs at this stage were clarified,and determined the purpose and research content of this study.(2)UFCB was modified by various methods.The zeta potential of the modified ultrafine carbon black(MCB)aqueous solution was-39.7 mV.which was much higher than-18.2 mV before modification,which proving that the dispersion stability of MCB was better than that before modification.MCB was applied to the adsorption experiment of lead,and finally a lead-loaded MCB(Pb-MCB)having a lead content of 0.235 g/g was obtained.(3)The effects of MCB and Pb-MCB on oxidative stress in mouse hepatocytes were compared.The results showed that there as a negative activity-dose relationship between hepatoeyte activity and MCB or Pb-MCB exposure concentration.The intracellular VIDA content increased with the increase of MCB or Pb-MCB exposure concentration.When the concentration of MCB and Pb-MCB was 50mg/L.the cell viability was 204.44%and 172.49%.respectively.High concentration of MDA breaks the intracellular redox balance inside cells,It also destroys the integrity of the cell membrane,leading to cell v itality reduction and even death.With the increase of exposure concentration of MCB and Pb-MCB,the relative viability of intracellular CAT increased first and then decreased.The exposure of MCB also led to the relative activitv change of intracellular SOD first rising and then falling.But with the increase of Pb-MCB exposure concentration,the relative activity of SOD increased graduallv.The effect of MCB and Pb-MCB on the oxidative stress index of mouse hepatocytes under the same exposure conditions showed that the cytotoxicity of MCB was stronger than that of Pb-MCB.because MCB had more negative charge than Pb-MCB.which is easier to combine with membrane proteins.CAT and SOD,affecting the vitality and functional expression of cells and enzymes.(4)A variety of spectroscopy methods were used to study the effects of MCB and Pb-MCB exposure on the structure and function of CAT and SOD molecules.The results showed that both MCB and Pb-MCB combined with CAT and SOD to form a protein film on the surface of carbon black particles,which changed the microenvironment of CAT and SOD chromophores.resulting in protein skeleton structure contraction and enhanced hydrophobicity.Both MCB and Pb-MCB exposures caused the activity of CAT to rise and then decrease and the activity of SOD continued to decrease.The changes in CAT and SOD activity indicate that structural changes affect the functional expression of them:on the other hand,the binding of CAT and SOD to MCB or Pb-MCB affects the microenvironment of the amino acid near its active site,thereby indirectly affecting the activity of them.Comparing the effects of MCB and Pb-MCB on CAT and SOD molecules under the same exposure conditions,it was found that the effect of MCB on the structure and activity of them was greater than Pb-MCB,because MCB had a large number of negative charges,which is more easily combined with CAT or SOD,and changes the structure of them to reduce its activity.The presence of trace Pb2+ in the Pb-MCB solution promotes the agglomeration of Pb-MCB particles,and the increase of particle size reduced the force and binding area between pb-MCB and protein.so MCB showed stronger molecular toxicity than Pb-MCB.This experiment explored and compared the toxic effects of MCB and Pb-MCB on the oxidative stress mechanism of the body from the cellular and molecular levels.and provided a theoretical and methodological reference for the evaluation of the toxic effects of multi-component atmospheric particles.