| Caenorhabditis elegans(C.elegans)plays an important role in the development of life sciences as an animal model.Caenorhabditis elegans has been widely used in the study of nervous system,aging mechanism and programmed cell death due to its simple structure,short life cycle,convenient genetic manipulation,and transparency.Unlike higher organisms,C.elegans lacks adaptive immune pathways.Only the innate immune pathway plays an important role in resisting pathogens and oxidative stress.Transforming growth factor ?(TGF-?),programmed cell death(PCD),insulin/insulin-like growth factor(insulin/IGF-1),mitogen-activated protein kinase are in its body.These four immune-related signal transduction pathways(mitogen activated protein kinases,MAPK)play a major role in different environments.At the same time,the innate immune system of C.elegans has many conserved points in evolution,which provides new ideas for the study of immune mechanisms in higher organisms.Nanoenzymes are a class of artificial mimic enzymes that not only have the unique properties of nanomaterials,but also have the catalytic efficiency and enzymatic reaction kinetics similar to natural enzymes.The discovery of nanoenzymes reveals new biological properties of nanomaterials.Because of it's good economy,high catalytic activity,and stability of mimic enzyme,the research of nanoenzymes has quickly become a research hotspot.At present,nanoenzymes have different degrees of cross-over research in different fields such as medicine,chemistry,and physics.There are many types of nanoenzymes.According to the material composition,nanoenzymes can be divided into carbon-based nanoenzymes,metal oxide nanoenzymes,metal nanoenzymes,and composite nanoenzymes.In the field of medical treatment,nanoenzymes can kill tumor cells powerfully and effectively.The antibacterial effect of nanoenzymes is one of the research hotspots in recent years.However,the research on iron-sulfur nanozymes in the field of biomedicine is not yet mature,and the biosafety assessment of nanozymes is far behind the speed of its research and development.However,most of the experiments stay in vitro experiments and have certain limitations.Therefore,choosing a good animal model as the subject of in vivo experiments is a scientific research problem that needs to be solved urgently.The aims of this study is to prepare iron-sulfur nano-enzymes and study in vivo with C.elegans,explore the effects of nano-enzymes on nematode innate immunity.By the nematode mutant strains,further explore the mechanism of nanoenzymes affecting the nematode innate immunity,and understand how the nematode regulates autoimmunity to achieve the effect of self-protection.Part I:Preparation of iron-sulfur nanozymeThe iron-sulfur nanozyme was prepared by solvothermal synthesis and configured as an aqueous solution.The microstructure and size of the iron-sulfur nanozyme were observed by scanning electron microscope and the elemental analysis of the nanozyme.This experiment successfully synthesized the iron-sulfur nanozyme with a particle size of about 100 nm.Part II:Effect of iron-sulfur nanozyme on the immunophenotype of Caenorhabditis elegansBy mixing the synthesized iron-sulfur nanozyme with E.coli food and spreading it on NGM medium to cultivate the wild strain(N2)of C.elegans,it was found that C.elegans had slight avoidance behavior to nanozymes,but with the intake of food,enter the body and gradually excrete the nanoenzyme from the body over time.Even if a higher concentration(4mg/mL)of iron-sulfur nanozyme was used to treat C.elegans,it has little effect on its survival rate,indicating that it has good biocompatibility.After 48 hours of iron-sulfur nanozyme treatment of nematodes,it was found that nanozymes had an effect on the immune phenotype of C.elegans.The lifespan of C.elegans was prolonged,and the number of eggs laid and the development of body length was decreased.After treatment with different concentrations of nanozyme(0.1-2 mg/mL),the ROS in the nematodes was increased with the increase of the concentration.The ROS level of C.elegans was further continuously tested.The nematodes treated with the nanozyme at a concentration of 1.5 mg/mL and the control group.The ROS levels of the nematodes in the first 5 days of the initial period increased with time,but the ROS levels of the nematodes treated with iron-sulfur nanozymes were much lower than the control group after the 6th day.At the same time,the SOD level in C.elegans was detected,and it was found that the SOD level in C.elegans was increased after iron-sulfur nanozyme treatment.In addition,using Fe3O4 nanoparticles of the same particle size as an experimental control,it was found that the nanomaterial has little effect on the immune phenotypes of C.elegans.Part?:The mechanism of iron-sulfur nanozyme on the innate immunity of Caenorhabditis elegansAfter treating C.elegans with iron-sulfur nanozyme,the expression of the screened innate immunity-related genes was detected.The results showed that the expression of genes related to innate immunity of C.elegans treated with iron-sulfur nanozymes was changed significantly.Among them,the expression of acdh-1,daf-2,daf-16,daf-18 genes were decreased,while the expression of lys-7 and pdk-1 Gene expression were up-regulated.Nematode strains with mutations in genes such as daf-2(e1370),daf-16(zls356 IV),daf-18(e1375),andpdk-1(mg142)?lys-7(ok1384).acdh-1(ok1489)mutations,which are important factors in the insulin pathway,were further explored and the results showed that iron-sulfur nanozymes could affect the innate immunity of C.elegans.Caenorhabditis elegans may affect the metabolism of C.elegans through insulin pathway,regulation of intestinal immunity and synthesis of antimicrobial peptides,so as to resist the changes of external environment and improve its immunity.This study provides an experimental basis for the study of innate immunity of iron sulfide nanozyme in C.elegans. |
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