Hydroalcoholic Extract of Scrophularia Striata Attenuates Hypertrophic Scar, Suppresses Collagen Synthesis, and Stimulates MMP2 and 9 Gene Expression in Rabbit Ear Model

Abstract

Objectives: Hypertrophic scars (HSs) are caused by abnormal wound healing. To date, no standard treatment has been made available for HSs. Scrophularia striata has been reported to accelerate wound healing and has the potential to prevent HS formation. In this study, we investigated the anti-scarring effects of S. striata extract (SSE) in a rabbit ear model of scarring.

Methods: In this study, New Zealand white rabbit (weight 2.3-2.5 kg) were used. In the prevention phase of the study, three test groups received 5%, 10%, and 15% ointments of SSE in the Eucerin base, the fourth group received Eucerin, and the fifth group received no treatment. The samples were obtained on day 35 after wounding. In the treatment phase of the study, the test groups received an intralesional injection of SSE (5%, 10%, and 15%), the fourth group received an intralesional injection of triamcinolone, the fifth group received a solvent (injection vehicle), and the sixth group received no treatment. To evaluate the anti-scarring effects of SSE, the scar elevation index (SEI), epidermis thickness index (ETI), collagen deposition, and MMP2 and MMP9 gene expression were evaluated.

Results: A significant reduction in SEI, ETI, and collagen deposition was noted in animals treated with SSE compared with the control groups. In addition, topical SSE stimulated MMP2 and MMP9 gene expression.

Conclusion: The findings of this study demonstrate the potential for SSE in the prevention and treatment of HS. SSE could be prepared as an appropriate formulation to treat wounds and prevent abnormal scarring.

Keywords: epidermal thickness index; hypertrophic scar; mmp2; mmp9; scar elevation index; scrophularia striata.

Figure 1

Macroscopic view of scar samples obtained from studied groups (in the prevention phase of the study) on day 1, 21 and 35 post injury. SSE, Scrophularia striata extract.

Figure 2

Macroscopic view of scar samples obtained from studied groups (in treatment phase of study) on day 1, 42 (immediately before treatment initiation) and 100 (30 days after treatment completion). SSE, Scrophularia striata extract; vehicle, NS + ethanol.

Figure 3

Effect of topical SSE on scar elevation index (a) Micrographs of H&E stained scar tissue sample (100X) obtained from (A) Eucerin (B) control, (C) SSE 5% ointment, (D) SSE 10% ointment, (E) SSE 15% ointment groups, (F) In this image the scar elevation index calculation method is shown. (b) Scar elevation index. Data are shown as mean ± SD, ***(p < 0.001) indicates significant differences from Eucerin and control groups. p value < 0.05 was consider as statistical significant. n = 4, SSE, Scrophularia striata extract.

Figure 4

Effect of intra-lesional injection of SSE on SEI. (a) H&E staining of tissue samples was performed to evaluation of microscopic evaluation of hypertrophic scar (100X). (A) Control, (B) vehicle (solvent), (C) Triamcinolone, (D) SSE 5%, (E) SSE 10%, (F) SSE 15%. (b) Quantification and statistical analysis of SEI. ***(p < 0.001) indicates differences from control and vehicle (solvent) groups, +++(p < 0.001) indicates differences from Triamcinolone group. Data are shown as mean ± SD. n = 4, SSE, Scrophularia striata extract.

Figure 5

Effect of topical SSE on ETI. Data are expressed as mean ± SD, ***(p < 0.001) indicates significant differences from control and Eucerin groups, p < 0.05 was considered as statistical significant. n = 5, SSE, Scrophularia striata extract.

Figure 6

Effect of intra-lesional injection of SSE on ETI. Data are expressed as mean ± SD, ***(p < 0.001) indicates significant differences from control and vehicle (solvent) groups, p < 0.05 was considered as statistical significant. n = 5, SSE, Scrophularia striata extract.

Figure 7

Effect of topical SSE on collagen deposition. (a) Evaluation of collagen deposition using Mason’s Trichrom staining on day 35 post injury (100X). The collagen fibers of SSE treated groups are more similar to those of normal skin tissue. (b) Quantification of collagen deposition and statistical analysis. *(p < 0.05) indicates the significant reduction in collagen deposition comparing to control and Eucerin groups. Data are shown as mean ± SD, n = 4, SSE, Scrophularia striata extract.

Figure 8

Effect of intra-lesional injection of SSE on collagen deposition. (a) Mason’s Trichrome staining was used to evaluation of collagen deposition in scar tissue samples (100X). The collagen fibers are more regular in Triamcinolone, SSE 10 and 15% treated groups comparing to those of control and vehicle (solvent) groups. The more cellularity (brown to red spots) are seen in tissue samples obtained from control and vehicle groups. (b) Quantification of collagen deposition. Data are shown as mean ± SD, *(p < 0.05) indicates significant differences from vehicle and control groups. p value < 0.05 was consider as statistical significant. n = 4, SSE, Scrophularia striata extract.

Figure 9

Effect of topical SSE on MMP2 (a) and MMP9 (b) gene expression. Data are shown as mean ± SD. **(p < 0.01) and ***(p < 0.001) indicate the significant differences from control and Eucerin groups. n = 6, SSE, Scrophularia striata extract.

Figure 10

. Effect of intra-lesional injection of SSE on MMP2 (a) and MMP9 (b) gene expression. Data are shown as mean ± SD, n = 6, SSE, Scrophularia striata extract.