ABSTRACT
Abstract A cationic liposomal gene delivery system comprising DOTAP, DOPE, and cholesterol was prepared and optimized. The results showed that the liposome/DNA (LP/DNA) system had spherical morphology, with a particle size of around 150 nm and zeta potential of approximately 30 mV. Cytotoxicity experiments showed that cells treated with all of the liposome carriers- with the exception of LP1-had more than 80% viability even at a weight ratio of 30. The in vitro transfection efficiency was measured using a Promega™ Luciferase Assay System. Of the tested lipoplexes, LP2/DNA showed the highest cell transfection efficiency (at a weight ratio of 10)-which was similar to or slightly lower than that of Lipofectamine® 2000 in HeLa, A549, and SPC-A1 cell lines. After freeze-drying, the cell transfection efficiency decreased slightly (P>0.05). The cell uptake mechanism study showed that LP/DNA lipoplexes mainly entered cells via clathrin-mediated and caveolin-mediated endocytic pathways. The results confirmed that LP2 has potential for use as an effective gene carrier, and provides experimental evidence to support its further development as a safe and effective gene delivery system.
ABSTRACT
Objective To investigate the uptake mechanism of HepG2.2.15 cells to the nanoparticles co-loaded with syringopicroside and hydroxytyrosol (SH-NPs). Methods The nanoparticles were prepared by using a nanoprecipitation method with mPEG-PLGA as nano-carrier co-loaded with syringopicroside and hydroxytyrosol. The uptake mechanism of HepG2.2.15 cells to SH-NPs was studied by fluorescence microscopy and flow cytometry using fluoresceineisothiocyanate (FITC) as a fluorescent marker. Results With colchicine as the inhibitor, the incubation time ranged from 0.5 to 24 h, the percentage of positive cells increased from 1.9% to 56.4%; When the drug concentration was 125, 250 μg/mL and 500 μg/mL, the positive cell percentages were 4.9%, 3.4% and 3.9%. With chloroquine as the inhibitor; the incubation time ranged from 0.5 to 24 h, the percentage of positive cells increased from 7.4% to 55.4%; When the drug concentration was 125, 250 and 500 μg/mL, the percentage of positive cells was 19.5%, 22.5% and 27.6%. Conclusion Colchicine and chloroquine have an inhibitory effect on HepG2.2.15 cells uptake, and the uptake of SH-NPs in HepG2.2.15 cells was positively correlated with drug concentration and incubation time. It can be concluded that the uptake mechanism of HepG2.2.15 cells to SH-NPs was nonspecific adsorption endocytosis.
ABSTRACT
OBJECTIVE: To prepare amphiphilic polycaprolactone-poly (arginine polymer) (PCL-R15)/siRNA Nanopexes, and two kinds of nanoparticles with different particle size were prepared by different process. After encapsulated siRNA with electrostatic interaction, both of two nanoplexes (NR60/siRNA and NR160/siRNA) were used to compare the effects in vitro cell levels. METHODS: The particle size and Zeta potential, siRNA loading and protection ability, cytotoxicity, cellular uptake mechanism and gene silencing efficiency of the two nanoplexes were investigated. RESULTS: The results show that the two nanoplexes have similar siRNA protection ability and cytotoxicity, but the difference between the two sizes is about 100 nm and the potential difference is about 20 mV. Moreover, NR160/siRNA complexes have higher cell uptake efficiency, more complex uptake pathways, and show greater gene silencing efficiency. CONCLUSION: These nanoplexes with different particle sizes can cause different transfection efficiency for siRNA delivery in cells.
ABSTRACT
With the development of nanotechnology,nanomaterials have great application potential in drug delivery system.Understanding the cellular uptake mechanism of nanoparticles has important scientific significance and application value in understanding life processes in cellular level,mechanisms of drug action and gene therapy,which provides the basis for developing safer and more effective nanosized drug carrier.This review summarizes the latest advancement about cellular uptake mechanisms of nanopatricles.Based on a brief introduction of the endocytic pathway of nanoparticles,influencing factors of endocytosis pathway are discussed,and the common methods used to study nanoparticle endocytosis pathways are also introduced in detail.
ABSTRACT
OBJECTIVE: To elucidate biological characteristics and uptake mechanism of Quercetin-nanostructured lipid carriers (Q-S) in vivo and in vitro. METHODS: Quercetin suspension (Q-X), quercetin-phospholipid complex (Q-L) and nanostructured lipid carriers loaded with quercetin (Q-S) were prepared. Then the intestinal permeability of these three formulations were studied using in situ rat intestinal perfusion, followed by in vivo relative oral bioavailability studies. In addition, in vitro cellular uptake of quercetin from Q-S in a time and concentration-dependent were carried out. To elucidate the uptake mechanism(s) from Q-S, Q-L and Q-X contribution of NPCILI carrier-mediated uptake, endocytosis and passive permeability were investigated. RESULTS: In situ studies demonstrated Q-S has 14-fold higher permeability than Q-X and 3-fold than Q-L. In vivo studies showed Q-S relative oral bioavailability is increased dramatically (Fr=4.6), meanwhile, no significant increased for Q-L compared with Q-X. Consistent with in situ results, in vitro time and concentration-dependent studies revealed quercetin uptake from Q-S was obviously higher than Q-X and Q-L. In vitro mechanistic characterization showed that the reduced contribution of NPCILI to the transport and passive diffusion enhancement of quercetin are primary explanations for its enhanced uptake from Q-S. CONCLUSION: Quercetin can enhance oral bioavailability attributed to enhance passive permeability and reduce transport of NPCILI formed nanostructured lipid carriers.
ABSTRACT
Some basic nuclear localization signals(NLS) have the capability of penetrating cell membrane and deliverying recombinant proteins, DNA, oligonucleotide into living cells. NLS based delivery system prohaps can satisfy most of major requirement in cargoing exogenous macromolecules or charged compounds. Different internalization mechanisms were found for different NLSs. Endocytosis or electrostatic binding of the anionic membranes and cationic NLSs probably induce celluar uptake.