Experimental Study On Monodisperse Polystyrene Nanospheres As SGC-7901Cell Strain Non-viral Gene Carrier In Vitro

Objective:To research and make monodisperse polystyrene nanospheres nanoparticles,text a cell fluorescence imaging,cell reproductive capacity,cellcycle to evaluate the ability of fluorescence imaging andbiocompatibility.According to this study we can find the possibility ofmonodisperse polystyrene nanospheres as advanced stomach cancernon-viral gene carrier,and we suppose to cure a tumor through it.Methord:(1)Make monodisperse polystyrene nanospheres nano particles byformula uesing dispersion polymerization method.(2)Set monodispersepolystyrene nanospheres nano particles concentrations as25ppm、50ppm、100ppm、200ppm、400ppm and separately cultured them withGES-1,GES-1,Hela cells12hours then used fluorescence nanoscopyobserve cell imaging、 flow cytometry detected fluorescenceintensity;(3)Culture the various concentrations of monodispersepolystyrene nanospheres nano particles with GES-1、SGC7901、Hela cellsafter24hours、48hours and72hours and test MTT.(4)After cultueSGC7901cells with each concentrations of monodisperse polystyrenenanosphere,test the cell cycles with FCM.Control group with nomonodisperse polystyrene nanosphere particles.（5） Use SPSS21.0performed the statistical analysis。 Results:(1) Set initial monomer concentration at12%,we can get a nearly200nm diameter particles,observe the surface with SEM we can findsmooth particles with perfect monodispersity and sphericity.Increase theconcentration of initial monomer to20%with other condition stay thesame,we can find that particle size getting bigger,at most850nm,andmonodispersity and sphericity are good enough.But when oncentration ofinitial monome more than20%,particles getting coagulate heavily andsphericity getting broken.Increase initiator concentration from0.5%to3%,particle size gettingbigger and particle distribution getting smaller at first then gettingbigger.In our study initiator concentration at1%,reaction most fast lead tonanosphere particles crash second time to create smallerparticles,therefore particle distribution become wider.According to the manufacturing operation,we choose polyvinylpyrrolidone(PVP) as stabilizer. With other condition stay steady,turnsPVP concentration6%up to10%,particles amount getting more,particlesize getting smaller.When we set PVP concentration lower than6%,particles collision increasing,particle size and particle distributionincreasing.(2)Use fluorescence nanoscope observe cells at both highmagnification and low magnification,bright field shows cells growth welland with great quantities;Dark field shows scattered pink spot with clearimages,and spots brightness increases with monodisperse polystyrenenanospheres particles concentration; Use FCM wih excitation wavelength488nm and emission wavelength488-630nm to test fluorescenceintensity,the result shows each experimental group compared with the control group remained statistically significant(P<0.05);eachexperimental group remained statisticallysignificant(P<0.05);fluorescence intensity in direct proportion withmonodisperse polystyrene nanospheres particles’ concentration.(3) MTT testing result:Use MTT method test cytotoxicity,no significantdifference between the experimental group and controlgroup(P>0.05);25ppm、50ppm、100ppm experimental groups’ absorbanceunder490nm wave length was positively associated with theconcentration of monodisperse polystyrene nanospheresparticles;200ppm、400ppm group cell proliferation turned to godown.According to this study proliferation index with monodispersepolystyrene nanospheres particles went upgrade firstly than descendinglatter tendency,200ppm is the maximum proliferation concentration,eachexperimental group remained statistically significant(P<0.05)(4) Cell cycle analysis:According to flow cytometry(FCM) test:0ppm、25ppm、50ppm、100ppm experimental groups cells proportion in G1andS phase had a decreasing trend,G2phase had a increasingtrend;significant differences had shown between experimental groups andcontrol groups(P<0.05).200ppm、400ppm experimental groups G1and Sphase cells increased, G2phase cells decreased, compared with controlgroup remained statistically significant (P<0.05).Conclusion:(1)We have made a kind of good monodispersity and sphericitynanospheres with proper diameter which can focus on further experiment.(2)Monodisperse polystyrene nanospheres particles’ fluorescenceimaging is clearly and stable,shows monodisperse polystyrenenanospheres can insert into the cells and with stable fluorescence intensity which can apply to further experiments.(3) From our study monodisperse polystyrene nanosphere particles ina certain concentration rate will not reduce cell activity.Monodispersepolystyrene nanosphere particle can lead cells G1phase and S phase toG2phase,and this is how it works on cell proliferation.