Caco-2 Cells To Self-assembly Of The Intake Of Sodium Alginate Nanoparticles

Objective To investigate the uptake of self-assembled sodium alginate(SA) nanoparticles labeled by FITC(sSAN-FITC) and Vitamin D₃ imbed by self-assembled sodium alginate nanoparticles labeled by FITC(SSAN-VD3-FITC),and their influential factors with Caco-2 cells.And to study the transmembrane transport of sSAN-FITC and SSAN-VD3-FITC with Caco-2 cells absorption model.Methods 1.In the uptake study,multifunction continuous spectrum enzyme-linked immunosorbent assay system was used to detect the fluorescence intensity and quantitative assay the effect of concentration,time and pH on the uptake of sSAN-FITC in Caco-2 cells.2.Fluorescence microscope and laser scanning confocal microscope(LSCM) were utilized to exam the uptake of sSAN-FITC and sSAN-VD₃-FITC absorbed by Caco-2 cells.And analyzed the effect of drug incubation time on the uptake.3.Caco-2 cells absorption model was used to study the transmembrane transport of sSAN-FITC and sSAN-VD₃-FITC from the apical side to basolateral side (AP-BL).Calculated the apparent permeability coefficient(Papp) and analyzed the effect of drug incubation time on the transmembrane transport.Multifunction continuous spectrum enzyme-linked immunosorbent assay system was used to detect the fluorescence intensity and quantitative assay the influence of time on the transmembrane transport of sSAN-FITC and sSAN-VD₃-FITC.Results 1.Multifunction continuous spectrum enzyme-linked immunosorbent assay system detection results showed that the fluorescence intensity in the cells was higher after sSAN-FITC was added(p<0.01),and the uptake was noted in a concentration and time dependent manner(p<0.01).①Compared with the control group,the fluorescence intensity in the cells was increased on different levels after different concentration(50,100,200,400μg/ml)of sSAN-FITC had interacted with Caco-2 cells for 2 hours.The fluorescence intensity of 200μg/ml and 400μg/ml groups were signifficantly higher(p<0.01),the difference was appararent compared with other groups(p<0.05,p<0.01).The one of 400μg/ml group was the highest(p<0.001).②The fluorescence intensity in the cells became higher on different levels after sSAN-FITC had ineracted with Caco-2 cells for different periods of time(1h,2h,4h)(p<0.01, p<0.001).It was the highest after having been incubated for 4h(p<0.001).③The pH of culture solution could influence the uptake of sSAN-FITC by Caco-2 cells.And the fluorescence intensity was higher in cells cultured in pH either higher or lower than 7.0(6.0,6.5,7.4) culture solution(p<0.05,p<0.01).It was the highest when pH was 6.0(p<0.01).2.Pictures of Caco-2 cells taken both by fluorescence microscope and laser scanning confocal microscope showed that sSAN-FITC and SSAN-VD3-FITC could be uptake by Caco-2 cells.After sSAN-FITC ineracted with Caco-2 cells for 1h,a lot of sSAN-FITC sticked to the membrane of Caco-2 cells,and formed a green fluorescence ring.Lots of fluorescence particle diameter appeared within Caco-2 cells after 4h.But it took only 1h for the appearance of sSAN-VD₃-FITC fluorescence particle diameter in Caco-2 cells.It was brighter when incubated for 4h.3.The results of transmembrane transport showed:First,from AP to BL,after having been incubated for different periods of time(0.5h,1h,2h,4h),the fluorescence intensity and accumulated transmembrane transport amount was enhanced at different levels compared with the control group(p<0.05,p<0.01).Both the fluorescence intensity and accumulated transmembrane transport quantity were the highest at 4h(compared with 0.5h,1h,2h,p<0.01).Papp was bigger while the time is longer.No saturation manifest had been found in this study. Second,as to the transmembrane transport of sSAN-VD3-FITC,the fluorescence intensity and the accum transport quantity were higher than the one of the control group(p<0.01), and they were higher while the time was longer and had statistics significance(p<0.05, p<0.01).Papp was fairly stabilized in the AP-BL direction,but it was smaller while the time is longer.Thirdly,the comparison of the transmembrane transport between sSAN-FITC and sSAN-VD₃-FITC showed:the fluorescence intensity of sSAN-VD₃-FITC was brighter than sSAN-FITC at 0.5h(p<0.05).Concision sSAN and sSAN-VD₃ can be uptake by Caco-2 cells,and the absorption of sSAN is dependent on the concentration and incubation time,it is also influenced by the pH of the culture solution.Both sSAN and sSAN-VD₃ can do transmembrane transport with Caco-2 cells absorption model,it is noted in a time-dependent manner.It indicates that sSAN and sSAN-VD₃ can be absorbed into blood via gastrointestinal tract pathway.