Mechanisms of ivermectin-induced wound healing

Abstract

Background: Wounds cause structural and functional discontinuity of an organ. Wound healing, therefore, seeks to re-establish the normal morphology and functionality through intertwined stages of hemostasis, inflammation, proliferation, and tissue remodelling. Ivermectin, a macrolide, has been used as an endectoparasiticide in human and veterinary medicine practice for decades. Here, we show that ivermectin exhibits wounding healing activity by mechanisms independent of its well-known antiparasitic activity. This study aimed to evaluate the wound healing property of ivermectin cream using histochemistry and enzyme-linked immunosorbent assay techniques.

Results: Non-irritant dose of ivermectin cream (0.03-1%) decreased wound macroscopic indices such as exudation, edge edema, hyperemia, and granulation tissue deposition by day 9 compared to day 13 for the vehicle-treated group. This corresponded with a statistically significant wound contraction rate, hydroxyproline deposition, and a decreased time to heal rate. The levels of growth factors TGF-β1 and VEGF were significantly elevated on day 7 but decreased on day 21. This corresponded with changes in cytokines (IL-1α, IL-4, IL-10, and TNF-α) and eicosanoids (LTB4, PGE₂, and PGD₂) levels on days 7 and 21._. Interestingly, low doses of ivermectin cream (0.03-0.1%) induced wound healing with minimal scarring compared to higher doses of the cream and the positive control, Silver Sulfadiazine.

Conclusion: Ivermectin promotes wound healing partly through modulation of the inflammatory process and the levels of Transforming Growth Factor-Beta 1 and Vascular Endothelial Growth Factor. Low doses of ivermectin cream have the potential to be used in treating wounds with minimal scar tissue formation.

Keywords: Cytokines; Eicosanoids; Growth factors; Hydroxyproline; Ivermectin; TGF-β 1; VEGF.

Fig. 1

Morphometric evaluation of the effects of ivermectin cream on cutaneous wound contraction. The time-course of the healing process (a) with the corresponding Areas Under Curve (AUC) (b); Wound Contraction rate (WC) expressed in percentages (c) with the corresponding AUCs (d) are as presented. Statistical analysis for each AUCs is by One-Way ANOVA. ** means p < 0.01 and *** means p < 0.001 when compared to the vehicle (naïve) control group

Fig. 2

Effects of ivermetin cream on wound collagen content. The wound tissues were taken and fixed in 4% phosphate-buffered formaldehyde (PBF) and stained with Picrosirius Red solution. Photomicrographs showed the degree of red color intensity connoting collagen density as observed in Control (a), Ag S (b), 1% ivermectin (c), 0.30% ivermectin (d), 0.10% ivermectin (e), 0.03% ivermectin (f). Image Scale bar is 40 μm. Micrographs were captured at 400x magnification with a resolution of 12.0 Megapixels

Fig. 3

Effects of ivermectin cream on cutaneous wound tissue levels of MPO and hydroxyproline. Sprague-Dawley rats were anaesthetised and excision wounds created as described in the methods. Animals were sacrificed on either day 7 or day 21 post-wounding. The levels of myeloperoxidase on day 7 as well as levels of hydroxyproline on day 7 and day 21 were measured and presented as mean ± SEM. The statistical analysis for MPO levels was by One-Way ANOVA. * means p < 0.05, ** means p < 0.01,*** means p < 0.001 for treated groups when compared to the vehicle (naïve) control group. The statistical analysis for hydroxyproline was by Two-Way ANOVA followed by Bonferroni's post hoc test. * means p < 0.05,** means p < 0.01 when compared to the corresponding day control levels whilst # means p < 0.05, ## means p < 0.01, #### means p < 0.0001 when comparing levels on day 7 to day 21 within a group

Fig. 4

Effects of ivermectin cream on cutaneous wound levels of TGF-B1 and VEGF. Sprague Dawley rats were anaesthetised and excision wounds created as described in the methods. Animals was sacrificed either on day 7 or day 21 post-wounding. The levels of TGF-ß1 on day 7 and day 21 as well as the levels of VEGF on day 7 and day 21 were measured and presented as mean ± SEM. The statistical analysis was by Two-Way ANOVA. Results shown as *p < 0.05,**p < 0.01,***p < 0.001, ****p < 0.0001, for all treated groups compared to vehicle (naïve) control group and # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001 represent comparison between levels on Day 7 and 21 within a group

Fig. 5

Effects of ivermectin on IL-1α, IL-4, IL-10, and TNF-α levels in cutaneous wounds. Sprague Dawley rats were anaesthetised and excision wounds created as described in the methods. Animals were sacrificed either on day 7 or day 21 post-wounding. The levels of the cytokines on day 7 and day 21 are presented graphically as mean ± SEM. The statistical analysis is by two-Way ANOVA followed by Bonferrroni's post hoc test. Results presented as *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 when comapred to the vehicle (naïve) control group and # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001 represent comparison between levels on Day 7 and 21 within a group

Fig. 6

Effects of ivermectin on LTB₄, PGE₂ and PGD₂ levels in cutaneous wound. Sprague Dawley rats were anaesthetised and excision wounds created as described in the methods. Animals were sacrificed either on day 7 or day 21 post-wounding. The levels of eicosanoids on day 7 and day 21 are presented graphically as as mean ± SEM. The Two-Way ANOVA statistical analysis results are shown as; # p< 0.05, #### p< 0.0001 when levels on day 7 are compared to levels on day 21 of the same group