Department of Veterinary Pathobiology, School of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX, United States.
Department of Diagnostic Medicine/Pathobiology, College of Veterinary Medicine, Kansas State University, Manhattan, KS, United States.
Front Cell Infect Microbiol. 2023 Oct 26;13:1253670. doi: 10.3389/fcimb.2023.1253670. eCollection 2023.
Tick serine protease inhibitors (serpins) play crucial roles in tick feeding and pathogen transmission. We demonstrate that Ixodes scapularis (Ixs) nymph tick saliva serpin (S) 41 (IxsS41), secreted by Borrelia burgdorferi (Bb)-infected ticks at high abundance, is involved in regulating tick evasion of host innate immunity and promoting host colonization by Bb. Recombinant (r) proteins were expressed in Pichia pastoris, and substrate hydrolysis assays were used to determine. Ex vivo (complement and hemostasis function related) and in vivo (paw edema and effect on Bb colonization of C3H/HeN mice organs) assays were conducted to validate function. We demonstrate that rIxsS41 inhibits chymase and cathepsin G, pro-inflammatory proteases that are released by mast cells and neutrophils, the first immune cells at the tick feeding site. Importantly, stoichiometry of inhibition analysis revealed that 2.2 and 2.8 molecules of rIxsS41 are needed to 100% inhibit 1 molecule of chymase and cathepsin G, respectively, suggesting that findings here are likely events at the tick feeding site. Furthermore, chymase-mediated paw edema, induced by the mast cell degranulator, compound 48/80 (C48/80), was blocked by rIxsS41. Likewise, rIxsS41 reduced membrane attack complex (MAC) deposition via the alternative and lectin complement activation pathways and dose-dependently protected Bb from complement killing. Additionally, co-inoculating C3H/HeN mice with Bb together with rIxsS41 or with a mixture (rIxsS41 and C48/80). Findings in this study suggest that IxsS41 markedly contributes to tick feeding and host colonization by Bb. Therefore, we conclude that IxsS41 is a potential candidate for an anti-tick vaccine to prevent transmission of the Lyme disease agent.
蜱虫丝氨酸蛋白酶抑制剂（serpins）在蜱虫的进食和病原体传播中发挥着关键作用。我们证明，伯氏疏螺旋体（Bb）感染的蜱虫大量分泌的Ixodes scapularis（Ixs）若虫唾液丝氨酸蛋白酶抑制剂（S）41（IxsS41），参与调节蜱虫逃避宿主固有免疫，并促进伯氏疏螺旋体对宿主的定殖。重组（r）蛋白在毕赤酵母中表达，并进行底物水解实验来测定。进行了体外（补体和止血功能相关）和体内（爪垫水肿和对 C3H/HeN 小鼠器官中伯氏疏螺旋体定殖的影响）实验来验证功能。我们证明 rIxsS41 抑制糜酶和组织蛋白酶 G，这两种是由肥大细胞和中性粒细胞释放的促炎蛋白酶，是在蜱虫进食部位的第一类免疫细胞。重要的是，抑制分析的化学计量学表明，rIxsS41 分子需要 2.2 和 2.8 个才能分别 100%抑制 1 个糜酶和组织蛋白酶 G，这表明这里的发现可能是在蜱虫进食部位发生的。此外，由肥大细胞脱颗粒剂化合物 48/80（C48/80）诱导的糜酶介导的爪垫水肿被 rIxsS41 阻断。同样，rIxsS41 通过替代和凝集素补体激活途径减少膜攻击复合物（MAC）沉积，并剂量依赖性地保护伯氏疏螺旋体免受补体杀伤。此外，将 rIxsS41 或混合物（rIxsS41 和 C48/80）与伯氏疏螺旋体一起共同接种 C3H/HeN 小鼠。本研究的结果表明，IxsS41 显著促进蜱虫的进食和伯氏疏螺旋体对宿主的定殖。因此，我们得出结论，IxsS41 是预防莱姆病病原体传播的抗蜱疫苗的潜在候选物。
