| Background:Due to their particular surface plasmon resonance,physical and chemical properties,gold nanoparticles have extensive potential applications in the field of in-situ imaging and targeting drug delivery.Nanomedicines based on AuNPs are generally designed to be administered through intravenous injection to increase their bioavailability,it is necessary to investigate the blood compatibility of AuNPs.Our previous studies found that PEG@AuNPs accelerated the aggregation of band3 protein in erythrocyte membrane,the oxygen-carrying capacity and deformability of erythrocytes was decreased consequently,however,the mechanism of oxidative damage induced by PEG@AuNPs remains unclear.In this research project,we analyzed the deoxygenation effect of PEG@AuNPs on hemoglobin,thus to investigate the influence of PEG@AuNPs on the metabolism of erythrocytes.And our work will provide the basis for comprehensive hemocompatibility evaluation of PEG@AuNPs.Objective:To elucidate the mechanism by which three diameter PEG@AuNPs caused oxidative damage to erythrocytes,we analyzed the deoxygenation effect of PEG@AuNPs on hemoglobin,and the influence of PEG@AuNPs on the metabolism of erythrocytes were also investigated.We also explored the effect of AuNPs on the coagulation system and the functions of leukocytes,platelets and erythrocytes,thus providing more comprehensive hemocompatibility evaluation of PEG@AuNPs.Methods:?Three different diameter PEG@AuNPs were synthesized by our research group,their physical and chemical properties were characterized.UV-Vis absorption spectrum,Far-UV circular dichroism,surface plasmon resonance and raman spectroscopy were employed to monitor the interaction between different diameter PEG@AuNPs and hemoglobin/albumin.?ICP-MS and four kinds of endocytosis inhibitors were employed to analyze the endocytosis mechanism of three diameter PEG@AuNPs.?Based on the erythrocytes function evaluation platform established by our research group and metabolomics technology,the influence of three different diameter PEG@AuNPs on central carbon metabolism,as well as the erythrocytes deformability,oxygen-carrying capacity were analyzed.?Percoll discontinuous density gradient centrifugation and ICP-MS were used to analyze the distribution of 13nm PEG@AuNPs and CT-AuNPs in blood cell components after incubation with whole blood under 37?,as well as the effect of two AuNPs on the coagulation system and the functions of blood cells were also analyzed.Results:?UV-Vis spectroscopy and dynamic light scattering revealed the formation of protein corona on the surface of PEG@AuNPs,and the results of Far-UV circular dichroism and raman spectroscopy showed that PEG@AuNPs lead to the deoxygenation of hemoglobin.?According to the ICP-MS results,the endocytosis efficiency of three diameter PEG@AuNPs were quite different.The larger the particle size,the higher endocytosis efficiency of erythrocytes,and low temperature could reduce the efficiency of endocytosis.ICP-MS revealed that 4.5nm,13nm PEG@AuNPs entered erythrocytes mainly by caveolae-mediated endocytosis,and clathrin mediated endocytosis also played an important role in endocytosis process of 30nm PEG@AuNPs.?PEG@AuNPs promoted hemoglobin deoxygenation and binding to the band3 protein,thus PEG@AuNPs induced oxdative damage to erythrocytes by suppress pentose phosphate pathway.Moreover,three different diameter PEG@AuNPs had effect on the erythrocytes deformability and oxygen-carrying capacity.?Whole blood was separated into plasma,leukocytes,platelets and erythrocytes by Percoll discontinuous density gradient centrifugation after incubation with PEG@AuNPs or CT-AuNPs.ICP-MS were used to analyze the distribution of PEG@AuNPs and CT-AuNPs in those four blood components,and we found that all three kinds of blood cells uptake PEG@AuNPs and CT-AuNPs,surprisingly,large amounts of AuNPs accumulated in platelets.We found that PEG@AuNPs and CT-AuNPs did not have great effect on hemolysis and erythrocytes deformability,neither PEG@AuNPs or CT-AuNPs stimulates monocytes to release IL-1?.And PEG@AuNPs didn't have great effect on platelet activation and aggregation function,while high concentration of CT-AuNPs activated platelets and impaired platelet aggregation functionConclusion:PEG@AuNPs have deoxygenation effect on hemoglobin,however,the largest size of PEG@AuNPs had the most significant deoxygenation effect Deoxyhemoglobin can bind to the band3 protein to regulate metabolic flux of erythrocytes.In denucleated mature erythrocytes,pentose phosphate pathway was the main source of NADPH generation,thus PEG@AuNPs caused an increase in NADP+/NADPH ratio and generated oxdative damage to erythrocytes.This is a new evidence of gold nanoparticles interfere with metabolic flux of erythrocytes and induce oxidative damage.In addition,large amounts of AuNPs accumulated in platelets during incubation with whole blood deserves further attention. |
PDF Full Text Request