Anti-IL-4, Anti-IL-17, and Anti-IFN-Gamma Activity in the Saliva of Amblyomma sculptum Ticks

Abstract

The saliva of hematophagous arthropods, such as ticks and triatomines, contains bioactive ligands capable of modulating immune molecules, including cytokines. Cytokines play a critical role in immune regulation and have therapeutic relevance in inflammatory and immune-mediated diseases. Despite recent advances, identifying cytokine-binding molecules remains a significant challenge. Interferon-gamma (IFN-γ), interleukin-4 (IL-4), and interleukin-17 (IL-17) are key cytokines involved in inflammation, adaptive immunity, and host defense. This study evaluated the ability of salivary components from Amblyomma sculptum and compared the results to the triatomine Rhodnius neglectus (used as control) to bind to IL-2, IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF-α using ELISA assays with human cytokines. Saliva samples were tested at dilutions of 1:25, 1:50, and 1:100. Saliva from A. sculptum, which demonstrated significant anti-cytokine activity, was fractionated via HPLC to identify the active components. The results confirmed the inhibitory capacity of A. sculptum saliva on IFN-γ, IL-4, and IL-17, with inhibition rates ranging from 30% to 70%, depending on the cytokine and dilution. No inhibitory activity was observed against IL-2, IL-6, IL-10, or TNF-α. These findings underscore the immunomodulatory role of A. sculptum saliva during tick feeding and suggest its potential for the development of novel immunobiologics to treat inflammatory and immune-mediated diseases.

Keywords: Amblyomma sculptum; cytokine-binding proteins; immunomodulation; saliva; ticks.

Figure 1

Inhibition of (A) IL-4, (B) IL-17, and (C) IFN-γ binding by Amblyomma sculptum saliva. The cytokine binding levels were compared to control levels (white bar) using triplicate values and are represented in pg/mL at saliva dilutions of 1/25 (light gray bar), 1/50 (gray bar), and 1/100 (dark gray bar). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars labeled with different letters (e.g., “a” and “b”) indicate statistically significant differences between groups with p < 0.05.

Figure 2

Inhibition of (A) IL-2, (B) IL-6, (C) IL-10, and (D) TNF-α binding activity of Amblyomma sculptum saliva. The cytokine binding levels were compared to control levels (white bar) using triplicate values and are represented in pg/mL at saliva concentrations of 1/25 (light gray bar), 1/50 (gray bar), and 1/100 (dark gray bar). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars labeled with different letters (e.g., “a” and “b”) indicate statistically significant differences between groups with p < 0.05.

Figure 3

Anti-binding IL-4 (A), anti-binding IL17 (B), and anti-binding IFN-γ (C) activity of Amblyomma sculptum saliva fractions. Cytokine binding levels of each fraction are compared. Red arrows highlight fractions with higher anti-cytokine activity. The dashed line serves to better visualize the reduction in cytokine level and is aligned with the control result.

Figure 4

Anti-cytokine binding activity of Rhodnius neglectus saliva against: (A) IL-2, (B) IL-4, (C) IL-6, (D) IL-10, (E) IL-17, (F) IFN-γ, and (G) TNF-α. The cytokine binding levels were compared to control levels (white bar) using triplicate values and are represented in pg/mL at saliva concentrations of 1/25 (light gray bar), 1/50 (gray bar), and 1/100 (dark gray bar). Statistical analysis was performed using one-way ANOVA followed by Tukey’s post hoc test. Bars labeled with different letters (e.g., “a” and “b”) indicate statistically significant differences between groups with p < 0.05.