Pharmaceutical Research

, Volume 26, Issue 1, pp 72–81 | Cite as

Poly (lactide-co-glycolide)-Polymethacrylate Nanoparticles for Intramuscular Delivery of Plasmid Encoding Interleukin-10 to Prevent Autoimmune Diabetes in Mice

  • Ashwin Basarkar
  • Jagdish Singh
Research Paper



Determine the efficiency of cationic nanoparticles prepared by blending poly (lactide-co-glycolide; PLGA) and methacrylate copolymer (Eudragit® E100) to deliver a therapeutic gene encoding mouse interleukin-10, in vitro and in vivo.


Nanoparticles prepared with PLGA and E100 were evaluated for delivery of plasmid DNA encoding mouse interleukin-10 in vitro and in vivo in mice upon intramuscular injection. Blood-glucose, serum interferon-gamma levels and histology of pancreas were studied to determine therapeutic efficacy. Histological evaluation of skeletal muscle from the injection site was performed to assess the biocompatibility of nanoparticles.


PLGA/E100 nanoparticles showed endosomal escape evidenced by confocal microscopy and buffering ability. Transfecting HEK293 cells with plasmid-loaded PLGA/E100 nanoparticles resulted in significantly (p < 0.05) greater expression of interleukin-10 compared to PLGA nanoparticles. Mice treated with PLGA/E100 nanoparticles displayed higher serum levels of interleukin-10 and lower blood glucose levels compared to those treated with interleukin-10 plasmid alone or PLGA nanoparticles. High expression of interleukin-10 facilitated suppression of interferon-gamma levels and reduced islet infiltration. Histology of muscle showed that nanoparticles were biocompatible and did not cause chronic inflammatory response.


Nanoparticles prepared by blending PLGA with methacrylate can efficiently and safely deliver plasmid DNA encoding mouse interleukin-10 leading to prevention of autoimmune diabetes.


interferon-gamma (IFN-γ) interleukin-10 (IL-10) plasmid DNA poly (lactide-co-glycolide) (PLGA) polymethacrylate 



We acknowledge financial support to JS from NIH grant # HD 46483-01 and Fraternal Order of Eagles fund.


  1. 1.
    J. A. Wolff, R.W. Malone, P. Williams, W. Chong, G. Acsadi, A. Jani, and P. L. Felgner. Direct gene transfer into mouse muscle in vivo. Science 247:1465–1468 (1990). doi: 10.1126/science.1690918.PubMedCrossRefGoogle Scholar
  2. 2.
    S. K. Tripathy, E. C. Svensson, H. B. Black, E. Goldwasser, M. Margalith, P. M. Hobart, and J. M. Leiden. Long-term expression of erythropoietin in the systemic circulation of mice after intramuscular injection of a plasmid DNA vector. Proc. Natl. Acad. Sci. U. S. A. 93:10876–10880 (1996). doi: 10.1073/pnas.93.20.10876.PubMedCrossRefGoogle Scholar
  3. 3.
    Q. L. Lu, G. Bou-Gharios, and T. A. Partridge. Non-viral gene delivery in skeletal muscle: a protein factory. Gene. Ther. 10:131–142 (2003). doi: 10.1038/ Scholar
  4. 4.
    I. S. Kim, S. K. Lee, Y. M. Park, Y. B. Lee, S. C. Shin, K. C. Lee, and I. J. Oh. Physicochemical characterization of poly(l-lactic acid) and poly(d,l-lactide-co-glycolide) nanoparticles with polyethylenimine as gene delivery carrier. Int. J. Pharm. 298:255–262 (2005). doi: 10.1016/j.ijpharm.2005.04.017.PubMedCrossRefGoogle Scholar
  5. 5.
    P. Li, J. M. Zhu, P. Sunintaboon, and F. W. Harris. New route to amphiphilic core-shell polymer nanospheres: graft copolymerization of methyl methacrylate from water-soluble polymer chains containing amino groups. Langmuir 18:8641–8646 (2002). doi: 10.1021/la0261343.CrossRefGoogle Scholar
  6. 6.
    D. Y. Kwoh, C. C. Coffin, C. P. Lollo, J. Jovenal, M. G. Banaszczyk, P. Mullen, A. Phillips, A. Amini, J. Fabrycki, R. M. Bartholomew, S. W. Brostoff, and D. J. Carlo. Stabilization of poly-l-lysine/DNA polyplexes for in vivo gene delivery to the liver. Biochim. Biophys. Acta. 1444:171–190 (1999).PubMedGoogle Scholar
  7. 7.
    R. Moriguchi, K. Kogure, H. Akita, S. Futaki, M. Miyagishi, K. Taira, and H. Harashima. A multifunctional envelope-type nano device for novel gene delivery of siRNA plasmids. Int. J. Pharm. 301:277–285 (2005). doi: 10.1016/j.ijpharm.2005.05.021.PubMedCrossRefGoogle Scholar
  8. 8.
    A. Bozkir and O. M. Saka. Chitosan nanoparticles for plasmid DNA delivery: effect of chitosan molecular structure on formulation and release characteristics. Drug Deliv. 11:107–112 (2004). doi: 10.1080/10717540490280705.PubMedCrossRefGoogle Scholar
  9. 9.
    K. A. Howard, U. L. Rahbek, X. Liu, C. K. Damgaard, S. Z. Glud, M. Ø. Andersen, M. B. Hovgaard, A. Schmitz, J. R. Nyengaard, F. Besenbacher, and J. Kjems. RNA interference in vitro and in vivo using a novel chitosan/siRNA nanoparticle system. Mol. Ther. 14:476–484 (2006). doi: 10.1016/j.ymthe.2006.04.010.PubMedCrossRefGoogle Scholar
  10. 10.
    S. Wang, N. Ma, S. J. Gao, H. Yu, and K. W. Leong. Transgene expression in the brain stem effected by intramuscular injection of polyethylenimine/DNA complexes. Mol. Ther. 3:658–664 (2001). doi: 10.1006/mthe.2001.0324.PubMedCrossRefGoogle Scholar
  11. 11.
    J. H. Williams, S. R. Sirsi, D. R. Latta, and G. J. Lutz. Induction of dystrophin expression by exon skipping in mdx mice following intramuscular injection of antisense oligonucleotides complexed with PEG-PEI copolymers. Mol. Ther. 14:88–96 (2006). doi: 10.1016/j.ymthe.2005.11.025.PubMedCrossRefGoogle Scholar
  12. 12.
    J. P. Behr. The proton sponge—a trick to enter cells the viruses did not exploit. CHIMIA 51:34–36 (1997).Google Scholar
  13. 13.
    P. Chollet, M. C. Favrot, A. Hurbin, and J. L. Coll. Side-effects of a systemic injection of linear polyethylenimine–DNA complexes. J. Gene. Med. 4:84–91 (2002). doi: 10.1002/jgm.237.PubMedCrossRefGoogle Scholar
  14. 14.
    S. M. Moghimi, P. Symonds, J. C. Murray, A. C. Hunter, G. Debska, and A. Szewczyk. A two-stage poly(ethylenimine)-mediated cytotoxicity: implications for gene transfer/therapy. Mol. Ther. 11:990–995 (2005). doi: 10.1016/j.ymthe.2005.02.010.PubMedCrossRefGoogle Scholar
  15. 15.
    P. Dubruel, B. Christiaens, M. Rosseneu, J. Vandekerckhove, J. Grooten, V. Goossens, and E. Schacht. Buffering properties of cationic polymethacrylates are not the only key to successful gene delivery. Biomacromolecules 5:379–388 (2004). doi: 10.1021/bm034438d.PubMedCrossRefGoogle Scholar
  16. 16.
    P. Dubruel, B. Christiaens, B. Vanloo, K. Bracke, M. Rosseneu, J. Vandekerckhove, and E. Schacht. Physicochemical and biological evaluation of cationic polymethacrylates as vectors for gene delivery. Eur. J. Pharm. Sci. 18:211–220 (2003). doi: 10.1016/S0928-0987(02)00280-4.PubMedCrossRefGoogle Scholar
  17. 17.
    J. W. Yoon and H. S. Jun. Autoimmune destruction of pancreatic beta cells. Am. J. Ther. 12:580–591 (2005). doi: 10.1097/01.mjt.0000178767.67857.63.PubMedCrossRefGoogle Scholar
  18. 18.
    D. Mathis, L. Vence, and C. Benoist. Beta-Cell death during progression to diabetes. Nature 414:792–798 (2001). doi: 10.1038/414792a.PubMedCrossRefGoogle Scholar
  19. 19.
    C. J. Clarke, A. Hales, A. Hunt, and B. M. Foxwell. IL-10-mediated suppression of TNF-alpha production is independent of its ability to inhibit NF kappa B activity. Eur. J. Immunol. 28:1719–1726 (1998). doi: 10.1002/(SICI)1521-4141(199805)28:05<1719::AID-IMMU1719>3.0.CO;2-Q.PubMedCrossRefGoogle Scholar
  20. 20.
    T. A. Hamilton, Y. Ohmori, and J. Tebo. Regulation of chemokine expression by antiinflammatory cytokines. Immunol. Res. 25:229–245 (2002). doi: 10.1385/IR:25:3:229.PubMedCrossRefGoogle Scholar
  21. 21.
    Z. L. Zhang, S. X. Shen, B. Lin, L. Y. Yu, L. H. Zhu, W. P. Wang, F. H. Luo, and L. H. Guo. Intramuscular injection of interleukin-10 plasmid DNA prevented autoimmune diabetes in mice. Acta. Pharmacol. Sin. 24:751–756 (2003)Medline.PubMedGoogle Scholar
  22. 22.
    M. Lee, K. S. Ko, S. Oh, and S. W. Kim. Prevention of autoimmune insulitis by delivery of a chimeric plasmid encoding interleukin-4 and interleukin-10. J. Control Release 88:333–342 (2003). doi: 10.1016/S0168-3659(03)00031-2.PubMedCrossRefGoogle Scholar
  23. 23.
    J. J. Koh, K. S. Ko, M. Lee, S. Han, J. S. Park, and S. W. Kim. Degradable polymeric carrier for the delivery of IL-10 plasmid DNA to prevent autoimmune insulitis of NOD mice. Gene Ther. 7:2099–2104 (2000). doi: 10.1038/ Scholar
  24. 24.
    K. S. Ko, M. Lee, J. J. Koh, and S. W. Kim. Combined administration of plasmids encoding IL-4 and IL-10 prevents the development of autoimmune diabetes in nonobese diabetic mice. Mol. Ther. 4:313–316 (2001). doi: 10.1006/mthe.2001.0459.PubMedCrossRefGoogle Scholar
  25. 25.
    M. X. Tang and F. C. Szoka. The influence of polymer structure on the interactions of cationic polymers with DNA and morphology of the resulting complexes. Gene Ther. 4:823–832 (1997). doi: 10.1038/ Scholar
  26. 26.
    C. G. Oster, N. Kim, L. Grode, L. Barbu-Tudoran, A. K. Schaper, S. H. Kaufmann, and T. Kissel. Cationic microparticles consisting of poly(lactide-co-glycolide) and polyethylenimine as carriers systems for parental DNA vaccination. J. Control Release. 104:359–377 (2005).PubMedGoogle Scholar
  27. 27.
    M. Singh, M. Briones, G. Ott, and D. O’Hagan. Cationic microparticles: a potent delivery system for DNA vaccines. Proc. Natl. Acad. Sci. U. S. A. 97:811–816 (2000). doi: 10.1073/pnas.97.2.811.PubMedCrossRefGoogle Scholar
  28. 28.
    E. Kawasaki, N. Abiru, and K. Eguchi. Prevention of type 1 diabetes: from the view point of beta cell damage. Diabetes Res. Clin. Pract. 66(Suppl 1):S27–32 (2004). doi: 10.1016/j.diabres.2003.09.015.PubMedCrossRefGoogle Scholar
  29. 29.
    V. Deleuze, D. Scherman, and M. F. Bureau. Interleukin-10 expression after intramuscular DNA electrotransfer: kinetic studies. Biochem. Biophys. Res. Commun. 299:29–34 (2002). doi: 10.1016/S0006-291X(02)02580-9.PubMedCrossRefGoogle Scholar
  30. 30.
    B. Wang, I. André, A. Gonzalez, J. D. Katz, M. Aguet, C. Benoist, and D. Mathis. Interferon-gamma impacts at multiple points during the progression of autoimmune diabetes. Proc. Natl. Acad. Sci. U. S. A. 94:13844–13849 (1997). doi: 10.1073/pnas.94.25.13844.PubMedCrossRefGoogle Scholar
  31. 31.
    B. Hultgren, X. Huang, N. Dybdal, and T. A. Stewart. Genetic absence of gamma-interferon delays but does not prevent diabetes in NOD mice. Diabetes 45:812–817 (1996). doi: 10.2337/diabetes.45.6.812.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  1. 1.Department of Pharmaceutical Sciences, College of Pharmacy, Nursing, and Allied SciencesNorth Dakota State UniversityFargoUSA

Personalised recommendations