1. The Majority of Keratinocytes Incorporate Intradermally Injected Plasmid DNA Regardless of Size but Only a Small Proportion of Cells Can Express the Gene Product 
Daisuke Sawamura, Kana Yasukawa, Kazuo Kodama, Koichi Yokota, Kazuko C. Sato-Matsumura, Hiroshi Shimizu 
Journal of Investigative Dermatology 
Volume 118, Issue 6, Pages (June 2002)
DOI: /j x
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Sequential change in the distribution of the IL-6-expressing keratinocytes after introduction of the gene. The phIL-6 plasmid expressing IL-6 was injected into the back skin of hairless rats. Skin biopsy specimens were taken from the treated sites at various times after gene introduction. IL-6 expression was examined with immunoreactivity to IL-6 antibody. (A) 4 h, (B) 8 h, (C) 12 h, (D) 18 h, (E) 24 h, (F) 48 h after gene transfer. Scale bar: 50 µm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
The lower epidermal layers incorporate and lose DNA earlier than the upper layers. We semiquantied a time course for the expression in the lower, middle, and upper layers of the epidermis. Skin samples were taken at 8 h, 12 h, 18 h, 24 h, and 48 h after gene introduction, and the IL-6 amounts were measured using ELISA method. Also, we observed 30 IL-6-positive areas form the corresponding skin sections. In each of the three layers (lower, middle, and upper), the ratio of IL-6 staining was scored out of a maximum of 10 (e.g. 3 for the lower layers, 4 for the middle layers, 3 for the upper layers). Finally, the value from the mean point of each layer multiplied by the IL-6 amount was expressed as the arbitrary transgene expression in each layer.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Expression of the type VII collagen gene in keratinocyte in vivo. The phCOL7 plasmid expressing the full-length cDNA of type VII collagen was injected into the back skin of hairless rats. Skin biopsy specimens were taken from the treated sites 24 h after gene introduction. The expression was examined with immunoreactivity to type VII collagen IL-6. (A) Control vector pCY4B, (B) phCOL7. Scale bar: 35 µm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
RT-PCR detects mRNA the type VII collagen gene. The phCOL7 plasmid was injected into the back skin of hairless rats. Skin biopsy specimens were taken from the treated sites 12 h, 24 h, 3 d, and a week after gene introduction. The mRNA expression was examined by RT-PCR. PCR amplified the 600 bp cDNA of type VII collagen gene. M, maker; lane 1, 12 h samples without reverse transcriptase; lane 2, without gene introduction; lane 3, 12 h; lane 4, 24 h; lane 5, 3 d; lane 6, a week samples.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Expression of GFP and presence of rhodamine after introduction of GFP expression plasmid labeled with rhodamine. The plasmid was injected into the back skin of hairless rats. Skin biopsy specimens were taken from the treated sites at various times after gene introduction. Red and green fluorescence was generated by rhodamine and GFP, respectively. (A) 0 h, (B) 4 h, (C); 8 h, (D) 12 h, (E) 24 h, after gene transfer. Arrows in (B) and (E) indicate the basement membrane zone. Scale bar: 40 µm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Dose of injected plasmid DNA and β-galactosidase activity. We took skin biopsied 24 h after introduction of pCMS-lacZ in several doses, and measured the β-galactosidase activity. β-Gal activity was expressed as relative light units (RLU) per µg protein.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2002 The Society for Investigative Dermatology, Inc Terms and Conditions