Abstract
Purpose. To delineate the characteristics and mechanisms of up- take of biodegradable poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles in primary cultured rabbit conjunctival epithelial cells (RCECs).
Methods. Poly(d,l-lactide-co-glycolide) nanoparticles (PLGA 50:50, 100 nm in diameter) containing 6-coumarin (as a fluorescent marker) were used. The effect of size was studied using various particle sizes (100 nm, 800 nm, and 10 μm). The effect of cytochalasin D, nocodazole, and metabolic inhibitors on nanoparticle uptake was investigated. The capability of nanoparticles to enhance the uptake of an encapsulated protein, BSA bound to Texas red (TR-BSA), was evaluated.
Results. Maximal uptake of nanoparticles at 37°C occurred at 2 h, and 100-nm particles had the highest uptake in RCECs in comparison with 800-nm and 10-μm particles. Nanoparticle uptake was saturable over the 0.1-4 mg/ml concentration range. Nanoparticle uptake was confirmed by confocal microscopy and was inhibited significantly by coumarin-free nanoparticles (of similar size), by lower incubation temperature, and by the presence of metabolic inhibitors and cytochalasin D. The uptake of encapsulated TR-BSA in RCECs at 4 h was 28% higher than free BSA application.
Conclusion. Our findings suggest that PLGA nanoparticle uptake in primary cultured rabbit conjunctival epithelial cells occurs most likely by adsorptive-type endocytosis.
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References
C. L. Bourlais, L. Acar, H. Zia, P. A. Sado, T. Needham, and R. Leverge. Ophthalmic drug delivery systems—recent advances. Prog. Retin. Eye Res. 17:33-58 (1998).
R. Wood, V. Li, J. Kreuter, and J. Robinson. Ocular disposition of poly-hexyl-2-cyano [3-14C] acrylate nanoparticles in the albino rabbit. Int. J. Pharm. 23:175-183 (1985).
A. Zimmer, P. Chetoni, M. Saettone, and H. Zerbe, and J. Kreuter. Evaluation of pilocarpine-loaded albumin nanoparticles as controlled drug delivery systems for the eye. II co-administration with bioadhesive and viscous polymers. J. Controll. Rel. 33:31-46 (1995).
A. Zimmer, J. Kreuter, and J. R. Robinson. Studies on the transport pathway of PBCA nanoparticles in ocular tissues. J. Microencapsul. 8:497-504 (1991).
L. Marchal-Heussler, D. Sirbat, M. Hoffman, and P. Maincent. Poly(epsilon-caprolactone) nanocapsules in carteolol ophthalmic delivery. Pharm. Res. 10:386-390 (1993).
M. Hashizoe, Y. Ogura, T. Takanashi, N. Kunou, Y. Honda, and Y. Ikada. Biodegradable polymeric device for sustained intravitreal release of ganciclovir in rabbits. Curr. Eye Res. 16:633-639 (1997).
R. Diepold, J. Kreuter, J. Himber, R. Gurny, V. H. Lee, J. R. Robinson, M. F. Saettone, and O. E. Schnaudigel. Comparison of different models for the testing of pilocarpine eyedrops using conventional eyedrops and a novel depot formulation (nanoparticles). Graefes Arch. Clin. Exp. Ophthalmol. 227:188-193 (1989).
D. K. Gilding and A. M. Reed. Biodegradable polymers for use in surgery: poly(glycolic) / poly(lactic acid) homo-and copolymers. Polymer 20:1459-1464 (1979).
P. N. Dilly. Contribution of the epithelium to the stability of the tear film. Trans. Ophthalmol. Soc. U. K. 104:381-389 (1985).
K. M. Hamalainen, K. Kananen, S. Auriola, K. Kontturi, and A. Urtti. Characterization of paracellular and aqueous penetration routes in cornea, conjunctiva, and sclera. Invest. Ophthalmol. Vis. Sci. 38:627-634 (1997).
P. Saha, T. Uchiyama, K. J. Kim, and V. H. Lee. Permeability characteristics of primary cultured rabbit conjunctival epithelial cells to low molecular weight drugs. Curr. Eye Res. 15:1170-1174 (1996).
J. Davda and V. Labhasetwar. Characterization of nanoparticle uptake by endothelial cells. Int. J. Pharm. 233:51-59 (2002).
U. B. Kompella, K. J. Kim, and V. H. Lee. Active chloride transport in the pigmented rabbit conjunctiva. Curr. Eye Res. 12:1041-1048 (1993).
P. Saha, K. J. Kim, and V. H. Lee. A primary culture model of rabbit conjunctival epithelial cells exhibiting tight barrier properties. Curr. Eye Res. 15:1163-1169 (1996).
J. J. Yang, H. Ueda, K. Kim, and V. H. Lee. Meeting future challenges in topical ocular drug delivery: development of an air-interfaced primary culture of rabbit conjunctival epithelial cells on a permeable support for drug transport studies. J Controll. Rel. 65:1-11 (2000).
M. P. Desai, V. Labhasetwar, E. Walter, R. J. Levy, and G. L. Amidon. The mechanism of uptake of biodegradable microparticles in Caco-2 cells is size dependent. Pharm. Res. 14:1568-1573 (1997).
P. Calvo, M. J. Alonso, J. L. Vila-Jato, and J. R. Robinson. Improved ocular bioavailability of indomethacin by novel ocular drug carriers. J. Pharm. Pharmacol. 48:1147-1152 (1996).
G. Wang, I. G. Tucker, M. S. Roberts, and L. W. Hirst. In vitro and in vivo evaluation in rabbits of a controlled release 5-fluorouracil subconjunctival implant based on poly(D,L-lactide-coglycolide). Pharm. Res. 13:1059-1064 (1996).
M. K. Pratten and J. B. Lloyd. Uptake of microparticles by rat visceral yolk sac. Placenta 18:547-552 (1997).
W. P. Jollie. Ultrastructural studies of protein transfer across rodent yolk sac. Placenta 7:263-281 (1986).
T. A. Gottlieb, I. E. Ivanov, M. Adesnik, and D. D. Sabatini. Actin microfilaments play a critical role in endocytosis at the apical but not the basolateral surface of polarized epithelial cells. J. Cell Biol. 120:695-710 (1993).
L. Manil, J. C. Davin, C. Duchenne, C. Kubiak, J. Foidart, P. Couvreur, and P. Mahieu. Uptake of nanoparticles by rat glomerular mesangial cells in vivo and in vitro. Pharm. Res. 11:1160-1165 (1994).
J. A. Oka, M. D. Christensen, and P. H. Weigel. Hyperosmolarity inhibits galactosyl receptor-mediated but not fluid phase endocytosis in isolated rat hepatocytes. J. Biol. Chem. 264:12016-12024 (1989).
C. Witschi and R. J. Mrsny. In vitro evaluation of microparticles and polymer gels for use as nasal platforms for protein delivery. Pharm. Res. 16:382-390 (1999).
E. Sander and C. Ashworth. A study of particulate intestinal absorption and hepatocellular uptake. Exp. Cell Res. 22:137-145 (1961).
Y. Horibe, K. Hosoya, K. J. Kim, T. Ogiso, and V. H. Lee. Polar solute transport across the pigmented rabbit conjunctiva: size dependence and the influence of 8-bromo cyclic adenosine monophosphate. Pharm. Res. 14:1246-1251 (1997).
P. Steuhl and J. W. Rohen. Absorption of horseradish peroxidase by the conjunctival epithelium of monkeys and rabbits 1. Graefes Arch. Clin. Exp. Ophthalmol. 220:13-18 (1983).
S. Latkovic and S. E. Nilsson. Phagocytosis of latex microspheres by the epithelial cells of the guinea pig conjunctiva. Acta Ophthalmol. (Copenh.) 57:582-590 (1979).
J. L. Goldstein, M. S. Brown, R. G. Anderson, D. W. Russell, and W. J. Schneider. Receptor-mediated endocytosis: concepts emerging from the LDL receptor system. Annu. Rev. Cell Biol. 1:1-39 (1985).
Y. Sai, M. Kajita, I. Tamai, J. Wakama, T. Wakamiya, and A. Tsuji. Adsorptive-mediated endocytosis of a basic peptide in enterocyte-like Caco-2 cells. Am. J. Physiol. 275:G514-G520 (1998).
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Qaddoumi, M.G., Ueda, H., Yang, J. et al. The Characteristics and Mechanisms of Uptake of PLGA Nanoparticles in Rabbit Conjunctival Epithelial Cell Layers. Pharm Res 21, 641–648 (2004). https://doi.org/10.1023/B:PHAM.0000022411.47059.76
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DOI: https://doi.org/10.1023/B:PHAM.0000022411.47059.76