Role Of Gold Nanoparticles In Inducing Osteoclast Differentiation In Vitro

Objective:The combination of nanotechnology and biomedicine is no longer a new topic.Among them,gold nanomaterials have the characteristics of good physical and chemical and easy surface modification properties,so they are heatedly debated in many medical fields.However,the cytotoxicity of nanoparticles also plagues its application in medicine,and its biosafety is the basis for the use of gold nanoparticles(GNPs) for medical purposes.At the same time,the role of gold nanomaterials in bone tissue regeneration has been concerned,and its biological role in osteoclast differentiation will be helpful for its research in the field of dental implant coating and osseointegration.In this experiment,gold nanoparticles were characterized and the gold nanoparticles with different concentrations and different particle sizes were used as variable to culture mouse bone marrow stromal cells in vitro,and their biocompatibility and osteoclasts were analyzed.Inhibition of differentiation.This experiment started with nuclear factor-activated T cell 1(Nfatc 1),c-fos and Tartrate-resistent Acid Phosphatase(TRAP)genes to study the expression changes of these genes in osteoclast differentiation induced by GNPs in vitro.Methods:The gold nanoparticle solution was detected by scanning electron microscopy and transmission electron microscopy.The zeta potential of the gold nanoparticles was determined,and the physical and chemical properties were analyzed.The mouse bone marrow cells were cultured in the medium of macrophage colony stimulating factor(M-CSF).High-purity bone marrow stromal cells were incubated and screened.The CCK-8 cytotoxicity was tested for the biocompatibility of GNPs with different particle sizes and concentrations.Bone marrow stromal stem cells were induced by adding receptor activator of nuclear factor-κB Ligand(RANKL)as a positive control group,and gold nanoparticles with different particle sizes and three equal concentrations were used as experimental groups.The differentiation state of the cells was observed at different time points,and the TRAP staining count was performed to compare the number and differentiation degree of osteoclasts induced by different concentrations of gold nanoparticles.Gold nanoparticles with excellent biocompatibility and inhibition of osteoclast differentiation were selected,and the concentration and particle size were confirmed.RT-PCR was used to detect the difference of Nfatc 1,c-fos and TRAP gene levels.Results:Electron microscopy showed that the gold nanoparticles showed oval shape,uniform dispersion,no obvious agglomeration;Zeta potential was-5.68 mV;At the same time,under the microscope,the morphology and size of the bone marrow stromal cells were consistent,and the distribution was uniform and wide.The cytotoxic activity of CCK8 showed that in the same particle size,the cytotoxicity of the gold nanoparticles in the 50 ng/ml group was significant,while the 30 ng/ml group and the 10 ng/ml group showed no difference compared with the control group.The cell morphology was mostly polygonal,long fusiform,oval,and the cell volume was uniform.The results of TRAP staining showed that under the same induction time and concentration,the cells in the experimental group were smaller than the positive control group,the number of TRAP-positive multinucleated cells was smaller,and the cell morphology was different.The cells in the 20 nm test group were polygonal,and the number of TRAP-positive multinucleated cells was higher than that in the 40 nm test group with statistical difference.Under the same induction time and concentration,the 10 ng/ml experimental group has the least proportion of TRAP-positive multinucleated cells compared with the other two experimental groups,the volume is small,and the cells are polygonal.RT-PCR results of 10 ng/ml and 20nm diameter spherical gold nanoparticles were obtained.The results showed that Nfatc 1,c-fos and TRAP were similar.The gene expression was positive control group>experimental group>control group.The difference was statistically significant(p<0.05).Conclusion:The spherical gold nanoparticles used in this experiment have uniform particle size,uniform dispersion and negative charge,and exhibit excellent physical and chemical properties.The mouse bone marrow cells were obtained as high-purified bone marrow stromal stem cells.Gold nanoparticles of a specific particle size can be fully endocytosed into the cytoplasm by cells,and have good biocompatibility.GNPs have the ability to inhibit osteoclast differentiation and exhibit concentration-dependent and size-dependent.Its inhibition may be achieved by inhibiting the osteoclast-related genes Nfatc 1,c-fos,and TRAP.