Inhibition of Smad3 Expression in Radiation-Induced Fibrosis Using a Novel Method for Topical Transcutaneous Gene Therapy

• Published in: Archives of otolaryngology--head & neck surgery Vol. 136; no. 7; pp. 714 - 719
• Main Authors: Lee, Judy W, Tutela, John P, Zoumalan, Richard A, Thanik, Vishal D, Nguyen, Phuong D, Varjabedian, Leon, Warren, Stephen M, Saadeh, Pierre B
• Format: Journal Article
• Language: English
• Published: Chicago, IL American Medical Association 01.07.2010
• Subjects: Administration, Cutaneous; Animals; Biological and medical sciences; Disease Models, Animal; Epidermis - growth & development; Epidermis - pathology; Fibrosis - pathology; Gels; Gene expression; Gene Expression Regulation; Gene therapy; Genetic Therapy - methods; Genotype & phenotype; Immunohistochemistry; Medical sciences; Mice; Mice, Inbred C57BL; Mice, Knockout; Otolaryngology; Otorhinolaryngology. Stomatology; Radiation Injuries - genetics; Radiation Injuries - pathology; Radiation Injuries - therapy; Random Allocation; RNA, Small Interfering - pharmacology; Rodents; Sensitivity and Specificity; Skin; Skin - pathology; Skin - radiation effects; Smad3 Protein - antagonists & inhibitors; Smad3 Protein - genetics; Smad3 Protein - metabolism; Treatment Outcome; Wound Healing - genetics; Wound Healing - physiology; Treatment; Inhibition; Gene therapy; Fibrosis
• ISSN: 0886-4470; 2168-6181; 1538-361X; 2168-619X
• DOI: 10.1001/archoto.2010.107

OBJECTIVE To attempt to mitigate the effects of irradiation on murine skin after high-dose radiation using a novel transcutaneous topical delivery system to locally inhibit gene expression with small interfering RNA (siRNA) against Smad3. DESIGN Laboratory investigation. SETTING University laboratory. SUBJECTS Twenty-five wild-type C57 mice. INTERVENTION In an isolated skin irradiation model, the dorsal skin of C57 wild-type mice was irradiated (45 Gy). Just before irradiation, Smad3 and nonsense siRNA were applied to 2 separate dorsal skin areas and then reapplied weekly. Skin was harvested after 1 and 4 weeks. Smad3 expression were assessed by immunohistochemistry, and collagen deposition and architecture was examined using picrosirius red collagen staining. MAIN OUTCOME MEASURES Epidermal thickness was measured semiquantitatively at 4 weeks. Radiation-induced fibrosis was measured quantitatively via tensiometry. The Young modulus, a measure of cutaneous rigidity inversely related to elasticity, was determined, with normal irradiated skin serving as a control specimen. RESULTS Murine skin treated with topical Smad3 siRNA demonstrated effective Smad3 inhibition at 1 week and persistent suppression at 4 weeks. Collagen deposition and epidermal thickness were significantly decreased in skin treated with Smad3 siRNA compared with control irradiated skin. Tensiometry demonstrated decreased tension in Smad3 siRNA–treated skin, with a Young modulus of 9.29 MPa (nonirradiated normal skin, 7.78 MPa) compared with nonsense (control) siRNA–treated skin (14.68 MPa). CONCLUSIONS Smad3 expression can be effectively silenced in vivo using a novel topical delivery system. Moreover, cutaneous Smad3 inhibition mitigates radiation-induced changes in tissue elasticity, restoring a near-normal phenotype.Arch Otolaryngol Head Neck Surg. 2010;136(7):714-719-->