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Seeing the gene therapy: application of gene gun technique to transfect and decolour pigmented rat skin with human agouti signalling protein cDNA

Gene Therapy volume 11, pages 1033–1039 (2004)Cite this article

Abstract

We developed a gene gun method for the transfer of human agouti signalling protein (ASP) cDNA to alter rat skin colour in vivo. Human ASP cDNA was cloned into a modified cytomegalovirus plasmid and delivered to the skin of Long–Evans rats by gene gun bombardment. Skin pigmentation, body weight and blood sugar of ASP cDNA-transfected rats were recorded against the control group, which were injected with plasmids encoding for green fluorescent protein. The treated skin showed lighter skin colour after 3 days of ASP gene transfection. This depigmentation effect was most prominent on day 14 and the skin gradually returned to its original pigmentation by day 28. Successful transfection of ASP gene in skin and hair follicles, as well as downregulation of melanocortin-1 receptor (MC1R) and tyrosinase expression upon treatment, was confirmed using immunohistochemistry and Western blot analysis. Body weight and blood sugar in the treated rats did not show statistically significant differences as compared to control groups. These observations demonstrate that gene transfer using the gene gun method can induce high cutaneous ASP production and facilitate a switch from dark to fair colour without systemic pleiotropic effects. Such a colour switch may be that ASP is acting in a paracrine fashion. In addition, this study verifies that ASP exerts its functions by acting as an independent ligand that downregulates the melanocyte MC1R and tyrosinase protein in an in vivo system. Our result offers new, interesting insights about the effect of ASP on pigmentation, providing a novel approach to study the molecular mechanisms underlying skin melanogenesis.

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Acknowledgements

This work was performed with the support of the Industrial Technology Research Institute of Taiwan and the National Science Council Grant NMRP 92-2314-B-182A-202 and Genome Project. We thank Dr VJ Hearing (National Cancer Institute, National Institutes of Health, MD, USA) for the polyclonal antibody (aPEP16) and Dr GS Barsh (Department of Pediatrics and Genetics, Stanford University School of Medicine, CA, USA) for the ASP plasmid.

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Authors and Affiliations

  1. Department of Dermatology, Chang Gung Memorial Hospital, Taipei, Taiwan

    C-H Yang, J C Lee & H-S Hong

  2. Department of Ophthalmology, Chang Gung Memorial Hospital, Taipei, Taiwan

    S-C Shen

  3. Gene Therapy Laboratory, E-DA Hospital, I-Shou University, Kaohsiung Hsien, Taiwan

    P-C Wu, C-Y Lu, C-T Meng & L-C Yang

  4. Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan

    S-F Hsueh

  5. Gene Therapy Laboratory, Tajen Institute of Technology, Pingtung Hsien, Taiwan

    L-C Yang

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  1. C-H Yang
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Yang, CH., Shen, SC., Lee, J. et al. Seeing the gene therapy: application of gene gun technique to transfect and decolour pigmented rat skin with human agouti signalling protein cDNA. Gene Ther 11, 1033–1039 (2004). https://doi.org/10.1038/sj.gt.3302264

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