Skip to main content
SpringerLink
Log in

Imaging Functional Nucleic Acid Delivery to Skin

  • Protocol
RNA Imaging

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1372))

Abstract

Monogenic skin diseases arise from well-defined single gene mutations, and in some cases a single point mutation. As the target cells are superficial, these diseases are ideally suited for treatment by nucleic acid-based therapies as well as monitoring through a variety of noninvasive imaging technologies. Despite the accessibility of the skin, there remain formidable barriers for functional delivery of nucleic acids to the target cells within the dermis and epidermis. These barriers include the stratum corneum and the layered structure of the skin, as well as more locally, the cellular, endosomal and nuclear membranes. A wide range of technologies for traversing these barriers has been described and moderate success has been reported for several approaches. The lessons learned from these studies include the need for combinations of approaches to facilitate nucleic acid delivery across these skin barriers and then functional delivery across the cellular and nuclear membranes for expression (e.g., reporter genes, DNA oligonucleotides or shRNA) or into the cytoplasm for regulation (e.g., siRNA, miRNA, antisense oligos). The tools for topical delivery that have been evaluated include chemical, physical and electrical methods, and the development and testing of each of these approaches has been greatly enabled by imaging tools. These techniques allow delivery and real time monitoring of reporter genes, therapeutic nucleic acids and also triplex nucleic acids for gene editing. Optical imaging is comprised of a number of modalities based on properties of light-tissue interaction (e.g., scattering, autofluorescence, and reflectance), the interaction of light with specific molecules (e.g., absorbtion, fluorescence), or enzymatic reactions that produce light (bioluminescence). Optical imaging technologies operate over a range of scales from macroscopic to microscopic and if necessary, nanoscopic, and thus can be used to assess nucleic acid delivery to organs, regions, cells and even subcellular structures. Here we describe the animal models, reporter genes, imaging approaches and general strategies for delivery of nucleic acids to cells in the skin for local expression (e.g., plasmid DNA) or gene silencing (e.g., siRNA) with the intent of developing nucleic acid-based therapies to treat diseases of the skin.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Lorenzer C et al (2015) Going beyond the liver: progress and challenges of targeted delivery of siRNA therapeutics. J Control Release 203:1–15

    Article  CAS  PubMed  Google Scholar 

  2. Hickerson RP et al (2011) Use of self-delivery siRNAs to inhibit gene expression in an organotypic pachyonychia congenita model. J Invest Dermatol 131:1037–1044, In press

    Article  CAS  PubMed  Google Scholar 

  3. Leachman SA et al (2008) Therapeutic siRNAs for dominant genetic skin disorders including pachyonychia congenita. J Dermatol Sci 51(3):151–157

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Leachman SA et al (2010) First-in-human mutation-targeted siRNA phase Ib trial of an inherited skin disorder. Mol Ther 18(2):442–446

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  5. Leslie Pedrioli DM et al (2012) Generic and personalized RNAi-based therapeutics for a dominant-negative epidermal fragility disorder. J Invest Dermatol 132(6):1627–1635

    Article  CAS  PubMed  Google Scholar 

  6. Préat V, Dujardin N (2001) Topical delivery of nucleic acids in the skin. STP Pharma Sci 1:57–68

    Google Scholar 

  7. Hickerson RP et al (2008) Single-nucleotide-specific siRNA targeting in a dominant-negative skin model. J Invest Dermatol 128(3):594–605

    CAS  PubMed  Google Scholar 

  8. Wong P, Domergue R, Coulombe PA (2005) Overcoming functional redundancy to elicit pachyonychia congenita-like nail lesions in transgenic mice. Mol Cell Biol 25(1):197–205

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  9. Smith FJ et al (2008) Development of therapeutic siRNAs for pachyonychia congenita. J Invest Dermatol 128(1):50–58

    Article  CAS  PubMed  Google Scholar 

  10. Chen J, Roop DR (2005) Mouse models in preclinical studies for pachyonychia congenita. J Investig Dermatol Symp Proc 10(1):37–46

    Article  CAS  PubMed  Google Scholar 

  11. Cao T et al (2001) An inducible mouse model for epidermolysis bullosa simplex: implications for gene therapy. J Cell Biol 152(3):651–656

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  12. Broderick KE, Humeau LM (2015) Electroporation-enhanced delivery of nucleic acid vaccines. Expert Rev Vaccines 14(2):195–204

    Article  CAS  PubMed  Google Scholar 

  13. Vogel FR, Sarver N (1995) Nucleic acid vaccines. Clin Microbiol Rev 8(3):406–410

    PubMed Central  CAS  PubMed  Google Scholar 

  14. Colluru VT et al (2013) Preclinical and clinical development of DNA vaccines for prostrate cancer. Urol Oncol. doi:10.1016/j.urolonc.2013.09.014, Pii: S1078-1439(13)00387-6

    Google Scholar 

  15. Gonzalez-Gonzalez E et al (2010) Increased interstitial pressure improves nucleic acid delivery to skin enabling a comparative analysis of constitutive promoters. Gene Ther 17(10):1270–1278

    Article  CAS  PubMed  Google Scholar 

  16. Hickerson RP et al (2015) Non-invasive intravital imaging of siRNA-mediated mutant keratin gene repression in skin. Mol Imaging Biol. doi:10.1007/s11307-015-0875-z

    Google Scholar 

  17. McLean WH et al (2011) The phenotypic and molecular genetic features of pachyonychia congenita. J Invest Dermatol 131(5):1015–1017

    Article  CAS  PubMed  Google Scholar 

  18. Eliason MJ et al (2012) A review of the clinical phenotype of 254 patients with genetically confirmed pachyonychia congenita. J Am Acad Dermatol 67(4):680–686

    Article  PubMed  Google Scholar 

  19. Kaspar RL (2005) Challenges in developing therapies for rare diseases including pachyonychia congenita. J Investig Dermatol Symp Proc 10(1):62–66

    Article  PubMed  Google Scholar 

  20. Sonn GA et al (2009) Fibered confocal microscopy of bladder tumors: an ex vivo study. J Endourol 23(2):197–201

    Article  PubMed  Google Scholar 

  21. Ra H et al (2010) Assessing delivery and quantifying efficacy of small interfering ribonucleic acid therapeutics in the skin using a dual-axis confocal microscope. J Biomed Opt 15(3):036027

    Article  PubMed Central  PubMed  Google Scholar 

  22. Gonzalez-Gonzalez E et al (2011) Visualization of plasmid delivery to keratinocytes in mouse and human epidermis. Sci Rep 1:158

    PubMed Central  PubMed  Google Scholar 

  23. Rogers FA, Hu RH, Milstone LM (2013) Local delivery of gene-modifying triplex-forming molecules to the epidermis. J Invest Dermatol 133(3):685–691

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  24. Hirokawa D, Lee JB (2011) Dermatoscopy: an overview of subsurface morphology. Clin Dermatol 29(5):557–565

    Article  PubMed  Google Scholar 

  25. Prescher JA, Contag CH (2010) Guided by the light: visualizing biomolecular processes in living animals with bioluminescence. Curr Opin Chem Biol 14(1):80–89

    Article  CAS  PubMed  Google Scholar 

  26. Ra H et al (2011) In vivo imaging of human and mouse skin with a handheld dual-axis confocal fluorescence microscope. J Invest Dermatol 131(5):1061–1066

    Article  CAS  PubMed  Google Scholar 

  27. Ra H et al (2008) Three-dimensional in vivo imaging by a handheld dual-axes confocal microscope. Opt Express 16(10):7224–7232

    Article  PubMed Central  PubMed  Google Scholar 

  28. Wang TD et al (2004) Confocal fluorescence microscope with dual-axis architecture and biaxial postobjective scanning. J Biomed Opt 9(4):735–742

    Article  PubMed Central  PubMed  Google Scholar 

  29. Wang TD et al (2003) Dual-axis confocal microscope for high-resolution in vivo imaging. Opt Lett 28(6):414–416

    Article  PubMed Central  PubMed  Google Scholar 

  30. Ra H et al (2010) In vivo imaging of human and mouse skin with a handheld dual-axis confocal fluorescence microscope. J Invest Dermatol 131:1061–1066

    Article  PubMed  Google Scholar 

  31. Ra H et al (2007) Two-dimensional MEMS scanner for dual-axes confocal microscopy. J Microelectromech Syst 16:969–976

    Article  Google Scholar 

  32. Piyawattanametha W et al (2012) In vivo near-infrared dual-axis confocal microendoscopy in the human lower gastrointestinal tract. J Biomed Opt 17(2):021102

    Article  PubMed Central  PubMed  Google Scholar 

  33. Gonzalez-Gonzalez E et al (2009) siRNA silencing of keratinocyte-specific GFP expression in a transgenic mouse skin model. Gene Ther 16(8):963–972

    Article  CAS  PubMed  Google Scholar 

  34. Hickerson RP et al (2013) Gene silencing in skin after deposition of self-delivery siRNA with a motorized microneedle array device. Mol Ther Nucleic Acids 2:e129

    Article  PubMed Central  PubMed  Google Scholar 

  35. Schindelin J et al (2012) Fiji: an open-source platform for biological-image analysis. Nat Methods 9(7):676–682

    Article  CAS  PubMed  Google Scholar 

  36. Thevenaz P, Ruttimann UE, Unser M (1998) A pyramid approach to subpixel registration based on intensity. IEEE Trans Image Process 7(1):27–41

    Article  CAS  PubMed  Google Scholar 

  37. Sawicki JA et al (1998) A composite CMV-IE enhancer/beta-actin promoter is ubiquitously expressed in mouse cutaneous epithelium. Exp Cell Res 244(1):367–369

    Article  CAS  PubMed  Google Scholar 

  38. Gonzalez-Gonzalez E et al (2010) Silencing of reporter gene expression in skin using siRNAs and expression of plasmid DNA delivered by a soluble protrusion array device (PAD). Mol Ther 18(9):1667–1674

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  39. Chong RH et al (2013) Gene silencing following siRNA delivery to skin via coated steel microneedles: In vitro and in vivo proof-of-concept. J Control Release 166(3):211–219

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  40. Kaspar R, McLean W, Schwartz M (2009) Achieving successful delivery of nucleic acids to skin: 6th annual meeting of the international pachyonychia congenita consortium. J Invest Dermatol 129:2085–2087

    Article  CAS  PubMed  Google Scholar 

  41. Takanashi M et al (2009) Therapeutic silencing of an endogenous gene by siRNA cream in an arthritis model mouse. Gene Ther 16(8):982–989

    Article  CAS  PubMed  Google Scholar 

  42. Hegde V et al (2014) In vivo gene silencing following non-invasive siRNA delivery into the skin using a novel topical formulation. J Control Release 196:355–362

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  43. Yi X et al (2010) MITF-siRNA formulation is a safe and effective therapy for human melasma. Mol Ther 19:362–371

    Article  PubMed Central  PubMed  Google Scholar 

  44. Hsu T, Mitragotri S (2011) Delivery of siRNA and other macromolecules into skin and cells using a peptide enhancer. Proc Natl Acad Sci USA 108(38):15816–15821

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  45. Chen M et al (2014) Topical delivery of siRNA into skin using SPACE-peptide carriers. J Control Release 179:33–41

    Article  PubMed Central  PubMed  Google Scholar 

  46. Zheng D et al (2012) Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation. Proc Natl Acad Sci U S A 109(30):11975–11980

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  47. Wang Q et al (2007) Delivery and inhibition of reporter genes by small interfering RNAs in a mouse skin model. J Invest Dermatol 127(11):2577–2584

    Article  CAS  PubMed  Google Scholar 

  48. Inoue T et al (2007) Modulation of scratching behavior by silencing an endogenous cyclooxygenase-1 gene in the skin through the administration of siRNA. J Gene Med 9(11):994–1001

    Article  CAS  PubMed  Google Scholar 

  49. Ritprajak P, Hashiguchi M, Azuma M (2008) Topical application of cream-emulsified CD86 siRNA ameliorates allergic skin disease by targeting cutaneous dendritic cells. Mol Ther 16(7):1323–1330

    Article  CAS  PubMed  Google Scholar 

  50. Lara MF et al (2012) Inhibition of CD44 gene expression in human skin models, using self-delivery short interfering RNA administered by dissolvable microneedle arrays. Hum Gene Ther 23(8):816–823

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  51. Lin CM et al (2012) A simple, noninvasive and efficient method for transdermal delivery of siRNA. Arch Dermatol Res 304(2):139–144

    Article  CAS  PubMed  Google Scholar 

  52. Garcia M et al (2010) Development of skin-humanized mouse models of pachyonychia congenita. J Invest Dermatol 131:1053–1060

    Article  PubMed  Google Scholar 

  53. Contag CH, Bachmann MH (2002) Advances in in vivo bioluminescence imaging of gene expression. Annu Rev Biomed Eng 4:235–260

    Article  CAS  PubMed  Google Scholar 

  54. Rajadhyaksha M et al (1995) In vivo confocal scanning laser microscopy of human skin: melanin provides strong contrast. J Invest Dermatol 104(6):946–952

    Article  CAS  PubMed  Google Scholar 

  55. Hickerson RP et al (2008) Stability study of unmodified siRNA and relevance to clinical use. Oligonucleotides 18(4):345–354

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  56. Rogers FA et al (2012) Targeted gene modification of hematopoietic progenitor cells in mice following systemic administration of a PNA-peptide conjugate. Mol Ther 20(1):109–118

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  57. Liu JT et al (2013) Real-time pathology through in vivo microscopy. Stud Health Technol Inform 185:235–264

    PubMed  Google Scholar 

  58. Liu JT et al (2011) Point-of-care pathology with miniature microscopes. Anal Cell Pathol (Amst) 34(3):81–98

    Article  Google Scholar 

Download references

Acknowledgments

This work was funded in part by the NIH through a grant called the NIH NIAMS GO Delivery! consortium grant (to RLK, LMM and CHC; RC2AR058955), support from the Pachyonychia Congenita Project (RK and CHC), and a gift from the Chambers Family Foundation (CHC).

Author information

Authors and Affiliations

  1. TransDerm Inc., 2161 Delaware Ave, Santa Cruz, CA, 95060, USA

    Roger L. Kaspar, Manuel A. Flores & Tycho P. Speaker

  2. Centre for Dermatology and Genetic Medicine, University of Dundee, Dundee, UK

    Robyn P. Hickerson

  3. Canvax Biotech S.L., Technological Park, Cordoba, Spain

    Emilio González-González

  4. Department of Therapeutic Radiology, Yale University School of Medicine, New Haven, CT, USA

    Faye A. Rogers

  5. Department of Dermatology, Yale University School of Medicine, New Haven, CT, USA

    Leonard M. Milstone

  6. Molecular Imaging Program at Stanford (MIPS), E150 Clark Center, Stanford University School of Medicine, 318 Campus Drive, Stanford, CA, 94305, USA

    Christopher H. Contag

  7. Department of Pediatrics, E150 Clark Center, Stanford University School of Medicine, 318 Campus Drive, Stanford, CA, 94305, USA

    Christopher H. Contag

  8. Department of Radiology, E150 Clark Center, Stanford University School of Medicine, 318 Campus Drive, Stanford, CA, 94305, USA

    Christopher H. Contag

  9. Microbiology and Immunology, E150 Clark Center, Stanford University School of Medicine, 318 Campus Drive, Stanford, CA, 94305, USA

    Christopher H. Contag

Authors
  1. Roger L. Kaspar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Robyn P. Hickerson
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Emilio González-González
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Manuel A. Flores
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Tycho P. Speaker
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Faye A. Rogers
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Leonard M. Milstone
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Christopher H. Contag
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Roger L. Kaspar or Christopher H. Contag .

Editor information

Editors and Affiliations

  1. Massachusetts General Hospital, Charlestown, Massachusetts, USA

    Zdravka Medarova

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Kaspar, R.L. et al. (2016). Imaging Functional Nucleic Acid Delivery to Skin. In: Medarova, Z. (eds) RNA Imaging. Methods in Molecular Biology, vol 1372. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3148-4_1

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3148-4_1

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3147-7

  • Online ISBN: 978-1-4939-3148-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation