杯状细胞靶向壳聚糖纳米粒的制备及吸收

Preparation and absorption of goblet cell targeted chitosan nanoparticles

目的将多肽CSKSSDYQC(CSK)化学结合到三甲基壳聚糖(trimethyl chitosan chloride,TMC)上,构建杯状细胞靶向口服给药系统,以期获得口服生物利用度较高的给药系统。方法通过化学合成的方法将CSK连接到TMC上,并用异硫氰酸荧光素(fluorescein isothiocyanate,FITC)进行荧光标记。以粒径和成球率为指标,采用单因素实验得到纳米粒的制备处方。用乳酸脱氢酶细胞毒性测试(lactate dehydrogenase,LDH)法分别考察载体材料和纳米粒对Caco-2和HT29-MTX细胞存活率的影响。建立Caco-2/HT29-MTX共培养细胞单分子层,进行纳米粒的跨膜转运实验。利用免疫荧光法考察纳米粒在体内吸收情况。结果按照最优处方制得的FITC-TMC纳米粒和FITC-TMC-CSK纳米粒的粒径分别为214.5 nm和234.3 nm;电位分别为15.81 mV和8.96 mV;成球率分别为98.14%和91.58%。FITC-TMC-CSK的细胞毒性低于FITC-TMC,纳米粒的细胞毒性低于载体材料。FITC-TMC-CSK纳米粒的累积透过量高于FITC-TMC纳米粒,其表观渗透系数(Papp)值是后者的2.17倍。FITC-TMC-CSK纳米粒在正常大鼠不同肠段吸收顺序为回肠>空肠>十二指肠。结论将CSK键合到TMC上构建杯状细胞靶向纳米粒,能有效提高纳米粒的吸收,在药物口服吸收的研究中具有广阔的应用前景。

Objective To construct goblet cell targeted oral delivery system through the chemical combination of peptide CSKSSDYQC(CSK)with trimethyl chitosan chloride(TMC).Methods In this study,CSK was attached to TMC through chemical synthesis and was further labeled with fluorescein isothiocyanate(FITC).The prescription of nanoparticles was obtained by single factor experiment with the particle size and precipitation efficacy as the indicators.And lactate dehydrogenase(LDH)method was used to study the effects of polymers and nanoparticles on the survival rates of Caco-2 and HT29-MTX cells.The study also established the monolayer of Caco-2/HT29-MTX co-cultured cells to evaluate the transport of nanoparticles and applied immunofluorescence to investigate the absorption of nanoparticles in vivo.Results The size of FITC-TMC nanoparticles and FITC-TMC-CSK nanoparticles prepared according to the optimum formulation were 214.5 nm and 234.3 nm,respectively;the potentials were 15.81 mV and 8.96 mV,respectively;and the precipitation efficacy was 98.14%and 91.58%,respectively.The cytotoxicity of FITC-TMC-CSK was lower than that of FITC-TMC,and the cytotoxicity of nanoparticles was lower than that of polymers.The accumulative permeability of FITC-TMC-CSK nanoparticles was higher than that of FITC-TMC nanoparticles and the apparent permeability coefficient(Papp)of FITC-TMC nanoparticles was 2.17 times higher than that of FITC-TMC nanoparticles.The maximum absorption of FITC-TMC-CSK nanoparticles was observed in ileum,where existed more amount of goblet cells compared with jejunum and duodenum in normal rats.Conclusion The construction of goblet cell targeted nanoparticles by attaching CSK to TMC can effectively improve the absorption of nanoparticles,which has a broad application prospect in the study of oral delivery system.