Pretreated Macrophage Membrane-coated Gold-silver Nanocages For Treatment Of Bacterial Infections & Case Reports

Part 1.Pretreated macrophage membrane-coated gold nanocages for treatment of bacterial infections Objectives: Gold nanocage(gold-silver nanocage,GSNC)has photothermal effect under the irradiation of near infrared laser,gold nanocage can be used as an effective antibacterial agent.Because of the hollow inner structure and porous wall structure of the gold nanocage,the nanoparticles can be used as the carrier of antibacterial drugs.However,there are bacterial recognition receptors on the surface of macrophage,which interact with the pathogen-associated molecular patterns in bacteria.Thus,according to the above two kinds of material,we designed a kind of new targeted delivery system,explored after certain bacteria treatment,the changes of the corresponding recognition on the macrophage membrane receptor expression,and further explored the targeting effect of bacteria and antibacterial effect of the macrophage membrane-coated gold nanocages in vivo and in vitro.Experiment 1.haracterization of bacteria pretreated macrophages Methods: After S.aureus,E.coli and macrophages co-culture,with the untreated macrophages as control,the macrophages with bacteria treatment were proceeded TLR2 and TLR4 protein immunofluorescence staining and flow cytometry to detect receptor expression on macrophage cell membrane.RT-PCR detected the TLR2 and TLR4 and TLR6 in the expression of mRNA level.Results: Cell protein immunofluorescence staining and cell flow analysis showed increased expression of TLR2 and TLR4 on macrophage membrane,and RT-PCR results showed up-regulated expression of TLR2,TLR4 and TLR6 in the mRNA level.Experiment 2.Characterization,photothermal behavior,and release profile ofSa-M-GSNCs Methods: Gold nanocages were prepared by alcohol reduction and gold-silver exchange reactions.Macrophage membranes were obtained by ultrasonic crushing and gradient centrifugation.After the above two kinds of material were mixed,the macrophage membranes-coated-gold nanocages were prepared by mechanical extrusion.The nanoparticles were characterized by a series of morphological and photothermal characterization,and photocontrolled pulse releasing was carried out using simulated drugs.Results: The macrophage membranes covered the surface of the gold nanocages uniformly,and the macrophage membrane-coated-nanocages showed good structural stability and protein biological stability,as well as excellent photothermal conversio n ability and photothermal conversion stability.The drug-loaded-gold nanocages could release drugs in response to light pulses.Experiment 3.Antibacterial activity of Sa-M-GSNCs in vitro Methods: Different macrophage membrane-coated-gold nanocages and s.aureus bacteria incubated together,the flow cytometry tested macrophage membrane-coated-gold nanocages with bacteria specific combining ability.And the bacterial growth curve,plate colony count and live/dead staining could detect the targeting bacteria and the antibacterial properties under the irradiation of the near-infrared light.Results: The highest binding rate of the macrophage membrane-coated-gold nanocages with S.aureus was found in the macrophage membrane treated with S.aureus,followed by E.coli pretreated macrophage membrane.Under the irradiation of near-infrared light,the antibacterial effect of the S.aureus pretreated macrophage membrane-coated-gold nanocages was the best.Experiment 4.Antibacterial activity of Sa-M-GSNCs in vivo Methods:1.Blood drug concentration and organ metabolism at different time points after intravenous injection were detected.2.The fluorescent-loaded macrophage membrane-coated-gold nanocages were locally injected into the subcutaneous abscess site of mice,and the retention of nanoparticles in the infected site was detected by fluorescence imaging.The macrophage membrane-coated-gold nanocages were injected into the subcutaneous abscess,under the irradiation of near-infrared light,the antibacterial effect was detected by the temperature rising and skin lesions at the infected site and count of bacterial colonies in the infected tissue.3.The fluorescent-loaded macrophage membrane-coated-gold nanocages were intravenously injected into the model mice of osteomyelitis,and the aggregation of nanoparticles at the infected site was detected by fluorescence imaging.After the macrophage membrane-coated-gold nanocages' systemic injection,under the irradiation of near-infrared light,the antibacterial effects of the infected site were detected by count of the bacterial colonies of the infected tissue.Results:1.Macrophage membrane coating could give the nanoparticles a longer time blood circulation,and the main metabolic organs were liver and kidney.2.The S.aureus pretreated macrophage membrane-coated-gold nanocages had the greatest drug retention at the subcutaneous abscess infected by S.aureus.Under the irradiation of near-infrared light,the greatest antibacterial effect of the nanoparticles was the S.aureus pretreated macrophage membrane-coated-gold nanocages.3.The S.aureus pretreated macrophage membrane-coated-gold nanocages could specifically target the infected site of the osteomyelitis infected by S.aureus.Under the irradiation of the near-infrared light,the greatest antibacterial effect of the nanoparticles was the S.aureus pretreated macrophage membrane-coated-gold nanocages.Conclusion: This experiment developed a kind of S.aureus pretreated macrophage membrane-coated-gold nanocages,this nano-system had obvious bacterial adhesion,and showed prolonged blood circulation in the body and good biocompatibility,and after local or systemic injection,the nanoparticles could effectively target and reserve in the infection site,and showed great antibacterial effect.