Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

Effect of topical interferon-γ gene therapy using gemini nanoparticles on pathophysiological markers of cutaneous scleroderma in Tsk/+ mice

Abstract

Scleroderma is a chronic disorder manifested by excessive synthesis and deposition of collagen in skin and connective tissue, vascular abnormalities, and autoimmunity. Using microarray and real-time PCR data, we show that intradermally expressed interferon γ (IFN-γ), generated after intradermal injection of IFN-γ-coding plasmid, and non-invasive topical nanoparticle (TNP) treatment with IFN-γ-coding plasmid, decreased collagen synthesis (via the Jak/Stat 1 pathway), upregulated Th1 cytokine levels, and downregulated the profibrotic cytokine Transforming growth factor β and the Smad pathways in the Tsk/+ (tight-skin scleroderma) mouse model. The TNP gene delivery system was constructed from gemini surfactant 16-3-16 and IFN-γ-coding plasmid. Topical administration of IFN-γ-coding plasmid in TNPs was effective in expressing IFN-γ levels after a 20-day treatment regimen without increased TLR4, CCL2, CCL11 and CCR2 mRNA levels that were observed in injected animals, signs considered to be innate responses to injury. The more uniform transgene IFN-γ expression caused significant (70–72%) collagen reduction, as assessed by reverse transcription real-time PCR. These results demonstrate efficient in vivo transfection using a gemini surfactant-based TNP delivery system able to modulate excessive collagen synthesis in scleroderma-affected skin.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Moschella S, Hurley H. . Connective tissue diseases. In: Moschella S, Hurley H (eds) Dermatology, 3rd ed edn. W.B. Saunders Company: Philadelphia, 1992, pp 1233–1245.

    Google Scholar 

  2. Roumm AD, Whiteside TL, Medsger Jr TA, Rodnan GP . Lymphocytes in the skin of patients with progressive systemic sclerosis. Quantification, subtyping, and clinical correlations. Arthritis Rheum 1984; 27: 645–653.

    Article  CAS  Google Scholar 

  3. Sapadin AN, Esser AC, Fleischmajer R . Immunopathogenesis of scleroderma--evolving concepts. Mt Sinai J Med 2001; 68: 233–242.

    CAS  PubMed  Google Scholar 

  4. Agarwal SK, Reveille JD . The genetics of scleroderma (systemic sclerosis). Curr Opin Rheumatol 2010; 22: 133–138.

    Article  CAS  Google Scholar 

  5. Seibold JR, Korn JH, Simms R, Clements PJ, Moreland LW, Mayes MD et al. Recombinant human relaxin in the treatment of scleroderma. A randomized, double-blind, placebo-controlled trial. Ann Intern Med 2000; 132: 871–879.

    Article  CAS  Google Scholar 

  6. Tsuji-Yamada J, Nakazawa M, Takahashi K, Iijima K, Hattori S, Okuda K et al. Effect of IL-12 encoding plasmid administration on tight-skin mouse. Biochem Biophys Res Commun 2001; 280: 707–712.

    Article  CAS  Google Scholar 

  7. Ihn H, Yamane K, Kubo M, Tamaki K . Blockade of endogenous transforming growth factor beta signaling prevents up-regulated collagen synthesis in scleroderma fibroblasts: association with increased expression of transforming growth factor beta receptors. Arthritis Rheum 2001; 44: 474–480.

    Article  CAS  Google Scholar 

  8. Zhou L, Askew D, Wu C, Gilliam AC . Cutaneous gene expression by DNA microarray in murine sclerodermatous graft-versus-host disease, a model for human scleroderma. J Invest Dermatol 2007; 127: 281–292.

    Article  CAS  Google Scholar 

  9. Varga J . Recombinant cytokine treatment for scleroderma: can the antifibrotic potential of interferon-gamma be realized clinically? Arch Dermatol 1997; 133: 637–642.

    Article  CAS  Google Scholar 

  10. Tovey MG, Lallemand C . Safety, tolerability and immunogenicity of interferons. Pharmaceuticals 2010; 3: 1162–1186.

    Article  CAS  Google Scholar 

  11. Ishida W, Mori Y, Lakos G, Sun L, Shan F, Bowes S et al. Intracellular TGF-beta receptor blockade abrogates Smad-dependent fibroblast activation in vitro and in vivo. J Invest Dermatol 2006; 126: 1733–1744.

    Article  CAS  Google Scholar 

  12. Ghosh AK, Yuan W, Mori Y, Chen S, Varga J . Antagonistic regulation of type I collagen gene expression by interferon-gamma and transforming growth factor-beta. Integration at the level of p300/CBP transcriptional coactivators. J Biol Chem 2001; 276: 11041–11048.

    Article  CAS  Google Scholar 

  13. Higashi K, Inagaki Y, Fujimori K, Nakao A, Kaneko H, Nakatsuka I . Interferon-gamma interferes with transforming growth factor-beta signaling through direct interaction of YB-1 with Smad3. J Biol Chem 2003; 278: 43470–43479.

    Article  CAS  Google Scholar 

  14. Smith GP, Chan ES . Molecular pathogenesis of skin fibrosis: insight from animal models. Curr Rheumatol Rep 2010; 12: 26–33.

    Article  CAS  Google Scholar 

  15. Christner PJ, Peters J, Hawkins D, Siracusa LD, Jimenez SA . The tight skin 2 mouse. An animal model of scleroderma displaying cutaneous fibrosis and mononuclear cell infiltration. Arthritis Rheum 1995; 38: 1791–1798.

    Article  CAS  Google Scholar 

  16. Badea I, Verrall R, Baca-Estrada M, Tikoo S, Rosenberg A, Kumar P et al. In vivo cutaneous interferon-gamma gene delivery using novel dicationic (gemini) surfactant-plasmid complexes. J Gene Med 2005; 7: 1200–1214.

    Article  CAS  Google Scholar 

  17. Badea I, Wettig S, Verrall R, Foldvari M . Topical non-invasive gene delivery using gemini nanoparticles in interferon-gamma-deficient mice. Eur J Pharm Biopharm 2007; 65: 414–422.

    Article  CAS  Google Scholar 

  18. Tang BL . ADAMTS: a novel family of extracellular matrix proteases. Int J Biochem Cell Biol 2001; 33: 33–44.

    Article  CAS  Google Scholar 

  19. Fantuzzi G . Adiponectin and inflammation: consensus and controversy. J Allergy Clin Immunol 2007; 121: 326–330.

    Article  Google Scholar 

  20. Snow JL, Su WP . Lipomembranous (membranocystic) fat necrosis. Clinicopathologic correlation of 38 cases. Am J Dermatopathol 1996; 18: 151–155.

    Article  CAS  Google Scholar 

  21. Siracusa LD, McGrath R, Ma Q, Moskow JJ, Manne J, Christner PJ et al. A tandem duplication within the fibrillin 1 gene is associated with the mouse tight skin mutation. Genome Res 1996; 6: 300–313.

    Article  CAS  Google Scholar 

  22. Baxter RM, Crowell TP, McCrann ME, Frew EM, Gardner H . Analysis of the tight skin (Tsk1/+) mouse as a model for testing antifibrotic agents. Lab Invest 2005; 85: 1199–1209.

    Article  CAS  Google Scholar 

  23. Jimenez SA, Feldman G, Bashey RI, Bienkowski R, Rosenbloom J . Co-ordinate increase in the expression of type I and type III collagen genes in progressive systemic sclerosis fibroblasts. Biochem J 1986; 237: 837–843.

    Article  CAS  Google Scholar 

  24. Jimenez SA, Millan A, Bashey RI . Scleroderma-like alterations in collagen metabolism occurring in the TSK (tight skin) mouse. Arthritis and Rheum 1984; 27: 180–185.

    Article  CAS  Google Scholar 

  25. Menton DN, Hess RA . The ultrastructure of collagen in the dermis of tight-skin (Tsk) mutant mice. J Invest Dermatol 1980; 74: 139–147.

    Article  CAS  Google Scholar 

  26. Pines M, Domb A, Ohana M, Inbar J, Genina O, Alexiev R et al. Reduction in dermal fibrosis in the tight-skin (Tsk) mouse after local application of halofuginone. Biochem Pharmacol 2001; 62: 1221–1227.

    Article  CAS  Google Scholar 

  27. Jimenez SA, Freundlich B, Rosenbloom J . Selective inhibition of human diploid fibroblast collagen synthesis by interferons. J Clin Invest 1984; 74: 1112–1116.

    Article  CAS  Google Scholar 

  28. Miller CH, Maher SG, Young HA . Clinical use of interferon-gamma. Ann N Y Acad Sci 2009; 1182: 69–79.

    Article  CAS  Google Scholar 

  29. Vlachoyiannopoulos PG, Tsifetaki N, Dimitriou I, Galaris D, Papiris SA, Moutsopoulos HM . Safety and efficacy of recombinant gamma interferon in the treatment of systemic sclerosis. Ann Rheum Dis 1996; 55: 761–768.

    Article  CAS  Google Scholar 

  30. Lortat-Jacob H, Baltzer F, Grimaud JA . Heparin decreases the blood clearance of interferon-gamma and increases its activity by limiting the processing of its carboxyl-terminal sequence. J Biol Chem 1996; 271: 16139–16143.

    Article  CAS  Google Scholar 

  31. Hunzelmann N, Anders S, Fierlbeck G, Hein R, Herrmann K, Albrecht M et al. Double-blind, placebo-controlled study of intralesional interferon gamma for the treatment of localized scleroderma. J Am Acad Dermatol 1997; 36 (3 Pt 1): 433–435.

    Article  CAS  Google Scholar 

  32. Pines M, Nagler A . Halofuginone: a novel antifibrotic therapy. Gen Pharmacol 1998; 30: 445–450.

    Article  CAS  Google Scholar 

  33. Hasegawa T, Nakao A, Sumiyoshi K, Tsuboi R, Ogawa H . IFN-gamma fails to antagonize fibrotic effect of TGF-beta on keloid-derived dermal fibroblasts. J Dermatol Sci 2003; 32: 19–24.

    Article  CAS  Google Scholar 

  34. Kirk TZ, Mark ME, Chua CC, Chua BH, Mayes MD . Myofibroblasts from scleroderma skin synthesize elevated levels of collagen and tissue inhibitor of metalloproteinase (TIMP-1) with two forms of TIMP-1. J Biol Chem 1995; 270: 3423–3428.

    Article  CAS  Google Scholar 

  35. Greenbaum D, Colangelo C, Williams K, Gerstein M . Comparing protein abundance and mRNA expression levels on a genomic scale. Genome Biol 2003; 4: 117.

    Article  Google Scholar 

  36. Pascal LE, True LD, Campbell DS, Deutsch EW, Risk M, Coleman IM et al. Correlation of mRNA and protein levels: cell type-specific gene expression of cluster designation antigens in the prostate. BMC Genomics 2008; 9: 246.

    Article  Google Scholar 

  37. Barnes PJ . Nuclear factor-kappa B. Int J Biochem Cell Biol 1997; 29: 867–870.

    Article  CAS  Google Scholar 

  38. Fehniger TA, Caligiuri MA . Interleukin 15: biology and relevance to human disease. Blood 2001; 97: 14–32.

    Article  CAS  Google Scholar 

  39. Menton DN, Hess RA, Lichtenstein JR, Eisen A . The structure and tensile properties of the skin of tight-skin (Tsk) mutant mice. J Invest Dermatol 1978; 70: 4–10.

    Article  CAS  Google Scholar 

  40. McGaha TL, Phelps RG, Spiera H, Bona C . Halofuginone, an inhibitor of type-I collagen synthesis and skin sclerosis, blocks transforming-growth-factor-beta-mediated Smad3 activation in fibroblasts. J Invest Dermatol 2002; 118: 461–470.

    Article  CAS  Google Scholar 

  41. Ciotti SN, Weiner N . Follicular liposomal delivery systems. J Liposome Res 2002; 12: 143–148.

    Article  CAS  Google Scholar 

  42. Rakhmilevich AL, Turner J, Ford MJ, McCabe D, Sun WH, Sondel PM et al. Gene gun-mediated skin transfection with interleukin 12 gene results in regression of established primary and metastatic murine tumors. Proc Natl Acad Sci USA 1996; 93: 6291–6296.

    Article  CAS  Google Scholar 

  43. Wu MH, Yokozeki H, Takagawa S, Yamamoto T, Satoh T, Kaneda Y et al. Hepatocyte growth factor both prevents and ameliorates the symptoms of dermal sclerosis in a mouse model of scleroderma. Gene Ther 2004; 11: 170–180.

    Article  CAS  Google Scholar 

  44. Bolstad BM, Irizarry RA, Astrand M, Speed TP . A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 2003; 19: 185–193.

    Article  CAS  Google Scholar 

  45. Asyali MH, Shoukri MM, Demirkaya O, Khabar KS . Assessment of reliability of microarray data and estimation of signal thresholds using mixture modeling. Nucleic Acids Res 2004; 32: 2323–2335.

    Article  CAS  Google Scholar 

  46. Pfaffl MW . A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 2001; 29: e45.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

Confocal microscopy images were recorded at the Saskatchewan Structural Sciences Centre, with the help of Dr Sophie Brunet. We thank Joe Petrik for preparing the illustrations and for editing the manuscript. This work was funded by the Canadian Institutes of Health Research (CIHR) and the Natural Sciences and Engineering Research Council of Canada (NSERC). We thank Dr Shawn Wettig for his assistance with some technical aspects of the animal experiments.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to M Foldvari.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Badea, I., Virtanen, C., Verrall, R. et al. Effect of topical interferon-γ gene therapy using gemini nanoparticles on pathophysiological markers of cutaneous scleroderma in Tsk/+ mice. Gene Ther 19, 978–987 (2012). https://doi.org/10.1038/gt.2011.159

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/gt.2011.159

Keywords

This article is cited by

Search

Quick links