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Abnormal scar identification with spherical-nucleic-acid technology

Nature Biomedical Engineeringvolume 2pages227238 (2018) | Download Citation


The accurate diagnosis of scar type and severity relies on histopathology of biopsied tissue, which is invasive and time-consuming, causes discomfort and may exacerbate scarring. Here, we show that imaging nanoprobes for the live-cell detection of intracellular messenger RNA (mRNA) (also known as NanoFlares) enable measurements of the expression of connective tissue growth factor (CTGF) as a visual indicator of hypertrophic scars and keloids. During cell culture, NanoFlares enabled the distinction of hypertrophic and keloidal fibroblasts from normal fibroblasts, and the detection of changes in CTGF expression resulting from the regulatory effects of transforming growth factor-β (TGF-β) agonists and TGF-β antagonists. We also applied the NanoFlares topically to the skin of live mice and rabbits, and to ex vivo human skin models. Transepidermal penetration of the NanoFlares enabled the visual and spectroscopic quantification of underlying abnormal fibroblasts on the basis of CTGF mRNA expression. Our proof-of-concept studies of topically applied NanoFlare technology as a means of biopsy-free scar diagnosis may eventually inform therapeutic decisions on the basis of the mRNA-expression patterns of skin disorders.

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  1. 1.

    Van de Vijver, M. J. et al. A gene-expression signature as a predictor of survival in breast cancer. N. Engl. J. Med. 347, 1999–2009 (2002).

  2. 2.

    Yao, M. et al. Gene expression analysis of renal carcinoma: adipose differentiation-related protein as a potential diagnostic and prognostic biomarker for clear-cell renal carcinoma. J. Pathol. 205, 377–387 (2005).

  3. 3.

    Zakrewsky, M., Kumar, S. & Mitragotri, S. Nucleic acid delivery into skin for the treatment of skin disease: proofs-of-concept, potential impact, and remaining challenges. J. Control. Release 219, 445–456 (2015).

  4. 4.

    Prigodich, A. E. et al. Multiplexed nanoflares: mRNA detection in live cells. Anal. Chem. 84, 2062–2066 (2012).

  5. 5.

    Halo, T. L. et al. NanoFlares for the detection, isolation, and culture of live tumor cells from human blood. Proc. Natl Acad. Sci. USA 111, 17104–17109 (2014).

  6. 6.

    Giljohann, D. A. et al. Gold nanoparticles for biology and medicine. Angew. Chem. Int. Ed. 49, 3280–3294 (2010).

  7. 7.

    Chinen, A. B. et al. Nanoparticle probes for the detection of cancer biomarkers, cells, and tissues by fluorescence. Chem. Rev. 115, 10530–10574 (2015).

  8. 8.

    Yang, Y. et al. FRET nanoflares for intracellular mRNA detection: avoiding false positive signals and minimizing effects of system fluctuations. J. Am. Chem. Soc. 137, 8340–8343 (2015).

  9. 9.

    Lahm, H. et al. Live fluorescent RNA-based detection of pluripotency gene expression in embryonic and induced pluripotent stem cells of different species. Stem Cells 33, 392–402 (2015).

  10. 10.

    Choi, C. H. J., Hao, L., Narayan, S. P., Auyeung, E. & Mirkin, C. A. Mechanism for the endocytosis of spherical nucleic acid nanoparticle conjugates. Proc. Natl Acad. Sci. USA 110, 7625–7630 (2013).

  11. 11.

    Wu, X. A., Choi, C. H. J., Zhang, C., Hao, L. & Mirkin, C. A. Intracellular fate of spherical nucleic acid nanoparticle conjugates. J. Am. Chem. Soc. 136, 7726–7733 (2014).

  12. 12.

    Jensen, S. A. et al. Spherical nucleic acid nanoparticle conjugates as an RNAi-based therapy for glioblastoma. Sci. Transl. Med. 5, 209ra152 (2013).

  13. 13.

    Randeria, P. S. et al. siRNA-based spherical nucleic acids reverse impaired wound healing in diabetic mice by ganglioside GM3 synthase knockdown. Proc. Natl Acad. Sci. USA 112, 5573–5578 (2015).

  14. 14.

    Sita, T. L. et al. Dual bioluminescence and near-infrared fluorescence monitoring to evaluate spherical nucleic acid nanoconjugate activity in vivo. Proc. Natl Acad. Sci. USA 114, 4129–4134 (2017).

  15. 15.

    Zheng, D. et al. Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation. Proc. Natl Acad. Sci. USA 109, 11975–11980 (2012).

  16. 16.

    Alster, T. S. & Tanzi, E. L. Hypertrophic scars and keloids: etiology and management. Am. J. Clin. Dermatol. 4, 235–243 (2003).

  17. 17.

    Gold, M. H. et al. Updated international clinical recommendations on scar management: part 2—algorithms for scar prevention and treatment. Dermatol. Surg. 40, 825–831 (2014).

  18. 18.

    Wolfram, D., Tzankov, A., Pülzl, P. & Piza-Katzer, H. Hypertrophic scars and keloids—a review of their pathophysiology, risk factors, and therapeutic management. Dermatol. Surg. 35, 171–181 (2009).

  19. 19.

    Sarrazy, V., Billet, F., Micallef, L., Coulomb, B. & Desmoulière, A. Mechanisms of pathological scarring: role of myofibroblasts and current developments. Wound Repair Regen. 19, s10–s15 (2011).

  20. 20.

    Van der Veer, W. M. et al. Potential cellular and molecular causes of hypertrophic scar formation. Burns 35, 15–29 (2009).

  21. 21.

    Grotendorst, G. R. Connective tissue growth factor: a mediator of TGF-beta action on fibroblasts. Cytokine Growth Factor Rev. 8, 171–179 (1997).

  22. 22.

    Leask, A. & Abraham, D. J. TGF-beta signaling and the fibrotic response. FASEB J. 18, 816–827 (2004).

  23. 23.

    Wang, Y. W. et al. siRNA-targeting transforming growth factor-beta type I receptor reduces wound scarring and extracellular matrix deposition of scar tissue. J. Investig. Dermatol. 134, 2016–2025 (2014).

  24. 24.

    Zhang, Z. et al. Recombinant human decorin inhibits TGF-β1-induced contraction of collagen lattice by hypertrophic scar fibroblasts. Burns 35, 527–537 (2009).

  25. 25.

    Wong, V. W., You, F., Januszyk, M., Gurtner, G. C. & Kuang, A. A. Transcriptional profiling of rapamycin-treated fibroblasts from hypertrophic and keloid scars. Ann. Plast. Surg. 72, 711–719 (2014).

  26. 26.

    Armendariz-Borunda, J. et al. A controlled clinical trial with pirfenidone in the treatment of pathological skin scarring caused by burns in pediatric patients. Ann. Plast. Surg. 68, 22–28 (2012).

  27. 27.

    Ichida, J. K. et al. A small-molecule inhibitor of TGF-beta signaling replaces sox2 in reprogramming by inducing nanog. Cell. Stem Cell. 5, 491–503 (2009).

  28. 28.

    Lin, T. et al. A chemical platform for improved induction of human iPSCs. Nat. Methods 6, 805–808 (2009).

  29. 29.

    Nestle, F. O., Di Meglio, P., Qin, J. Z. & Nickoloff, B. J. Skin immune sentinels in health and disease. Nat. Rev. Immunol. 9, 679–691 (2009).

  30. 30.

    Kua, E. H. J., Goh, C. Q., Ting, Y., Chua, A. & Song, C. Comparing the use of glycerol preserved and cryopreserved allogenic skin for the treatment of severe burns: differences in clinical outcomes and in vitro tissue viability. Cell Tissue Bank. 13, 269–279 (2012).

  31. 31.

    Pasyk, K. A., Argenta, L. C. & Hassett, C. Quantitative analysis of the thickness of human skin and subcutaneous tissue following controlled expansion with a silicone implant. Plast. Reconstr. Surg. 81, 516–523 (1988).

  32. 32.

    Kloeters, O., Tandara, A. & Mustoe, T. A. Hypertrophic scar model in the rabbit ear: a reproducible model for studying scar tissue behavior with new observations on silicone gel sheeting for scar reduction. Wound Repair Regen. 15, S40–S45 (2007).

  33. 33.

    Yeo, D. C., Balmayor, E. R., Schantz, J.-T. & Xu, C. Microneedle physical contact as a therapeutic for abnormal scars. Eur. J. Med. Res. 22, 28 (2017).

  34. 34.

    Sellheyer, K. & Bergfeld, W. F. A retrospective biopsy study of the clinical diagnostic accuracy of common skin diseases by different specialties compared with dermatology. J. Am. Acad. Dermatol. 52, 823–830 (2005).

  35. 35.

    Lewandowski, K. T. et al. Topically delivered tumor necrosis factor-α-targeted gene regulation for psoriasis. J. Investig. Dermatol. 137, 2027–2030 (2017).

  36. 36.

    Avior, Y., Sagi, I. & Benvenisty, N. Pluripotent stem cells in disease modelling and drug discovery. Nat. Rev. Mol. Cell Biol. 17, 170–182 (2016).

  37. 37.

    Choi, S. M. et al. Efficient drug screening and gene correction for treating liver disease using patient-specific stem cells. Hepatology 57, 2458–2468 (2013).

  38. 38.

    Gammon, S. T., Leevy, W. M., Gross, S., Gokel, G. W. & Piwnica-Worms, D. Spectral unmixing of multicolored bioluminescence emitted from heterogeneous biological sources. Anal. Chem. 78, 1520–1527 (2006).

  39. 39.

    Beziere, N. et al. Dynamic imaging of PEGylated indocyanine green (ICG) liposomes within the tumor microenvironment using multi-spectral optoacoustic tomography (MSOT). Biomaterials 37, 415–424 (2015).

  40. 40.

    Vujačić, A. et al. Fluorescence quenching of 5,5’-disulfopropyl-3,3’-dichlorothiacyanine dye adsorbed on gold nanoparticles. J. Phys. Chem. C 117, 6567–6577 (2013).

  41. 41.

    Hayes, A. J. et al. Wide versus narrow excision margins for high-risk, primary cutaneous melanomas: long-term follow-up of survival in a randomised trial. Lancet Oncol. 17, 184–192 (2016).

  42. 42.

    Anselmo, A. C. & Mitragotri, S. Nanoparticles in the clinic. Bioeng. Transl. Med. 1, 10–29 (2016).

  43. 43.

    Banga, R. J., Chernyak, N., Narayan, S. P., Nguyen, S. T. & Mirkin, C. A. Liposomal spherical nucleic acids. J. Am. Chem. Soc. 136, 9866–9869 (2014).

  44. 44.

    Banga, R. J. et al. Cross-linked micellar spherical nucleic acids from thermoresponsive templates. J. Am. Chem. Soc. 139, 4278–4281 (2017).

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This work was supported by the NTU-Northwestern Institute for Nanomedicine. We thank G. Yu (current location: Nanjing University of Posts and Telecommunications, China) for assistance with animal handling.

Author information


  1. School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore, Singapore

    • David C. Yeo
    • , Christian Wiraja
    •  & Chenjie Xu
  2. NTU-Northwestern Institute for Nanomedicine, Nanyang Technological University, Singapore, Singapore

    • Amy S. Paller
    • , Chad A. Mirkin
    •  & Chenjie Xu
  3. Department of Dermatology, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA

    • Amy S. Paller
  4. Department of Chemistry, Northwestern University, Evanston, IL, USA

    • Chad A. Mirkin
  5. International Institute for Nanotechnology, Northwestern University, Evanston, IL, USA

    • Chad A. Mirkin


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C.A.M., A.S.P., C.X. and D.C.Y. conceived and designed the experiments. D.C.Y. and C.W. performed the experiments. D.C.Y., C.W. and C.X. analysed and interpreted the data. D.C.Y., C.X., A.S.P. and C.A.M. wrote the manuscript. All authors read and approved the final manuscript.

Competing interests

A patent application based on the reported data has been filed. C.A.M. is a cofounder of Aurasense (the company that co-developed and licensed the NanoFlare technology to Merck–Millipore, which produced over 1,600 commercial versions of NanoFlares sold under the trade name SmartFlares).

Corresponding authors

Correspondence to Amy S. Paller or Chad A. Mirkin or Chenjie Xu.

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