Protease inhibitors regulate various biological processes and prevent host tissue/organ damage. Specific inhibition/regulation of proteases is clinically valuable for treating several diseases. Psoriasis affects the skin in the limbs and scalp of the body, and the contribution of cysteine and serine proteases to the development of skin inflammation is well documented. Cysteine protease inhibitors from ticks have high specificity, selectivity, and affinity to their target proteases and are efficient immunomodulators. However, their potential therapeutic effect on psoriasis pathogenesis remains to be determined. Therefore, we tested four tick cystatins (Sialostatin L, Sialostatin L2, Iristatin, and Mialostatin) in the recently developed, innate immunity-dependent mannan-induced psoriasis model. We explored the effects of protease inhibitors on clinical symptoms and histological features. In addition, the number and percentage of immune cells (dendritic cells, neutrophils, macrophages, and γδT cells) by flow cytometry, immunofluorescence/immunohistochemistry and, the expression of pro-inflammatory cytokines (TNF-a, IL-6, IL-22, IL-23, and IL-17 family) by qPCR were analyzed using skin, spleen, and lymph node samples. Tick protease inhibitors have significantly decreased psoriasis symptoms and disease manifestations but had differential effects on inflammatory responses and immune cell populations, suggesting different modes of action of these inhibitors on psoriasis-like inflammation. Thus, our study demonstrates, for the first time, the usefulness of tick-derived protease inhibitors for treating skin inflammation in patients.

• Keywords
• autoimmune disease, immune responses, protease inhibitors, psoriasis, tick,
• MeSH
• Dermatitis * MeSH
• Endopeptidases MeSH
• Cysteine Proteinase Inhibitors MeSH
• Protease Inhibitors MeSH
• Humans MeSH
• Mannans MeSH
• Immunity, Innate MeSH
• Peptide Hydrolases MeSH
• Psoriasis * chemically induced   drug therapy   MeSH
• Inflammation drug therapy   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Endopeptidases MeSH
• Cysteine Proteinase Inhibitors MeSH
• Protease Inhibitors MeSH
• Mannans MeSH
• Peptide Hydrolases MeSH

Serpins are widely distributed and functionally diverse inhibitors of serine proteases. Ticks secrete serpins with anti-coagulation, anti-inflammatory, and immunomodulatory activities via their saliva into the feeding cavity to modulate host's hemostatic and immune reaction initiated by the insertion of tick's mouthparts into skin. The suppression of the host's immune response not only allows ticks to feed on a host for several days but also creates favorable conditions for the transmission of tick-borne pathogens. Herein we present the functional and structural characterization of Iripin-1 (Ixodes ricinus serpin-1), whose expression was detected in the salivary glands of the tick Ixodes ricinus, a European vector of tick-borne encephalitis and Lyme disease. Of 16 selected serine proteases, Iripin-1 inhibited primarily trypsin and further exhibited weaker inhibitory activity against kallikrein, matriptase, and plasmin. In the mouse model of acute peritonitis, Iripin-1 enhanced the production of the anti-inflammatory cytokine IL-10 and chemokines involved in neutrophil and monocyte recruitment, including MCP-1/CCL2, a potent histamine-releasing factor. Despite increased chemokine levels, the migration of neutrophils and monocytes to inflamed peritoneal cavities was significantly attenuated following Iripin-1 administration. Based on the results of in vitro experiments, immune cell recruitment might be inhibited due to Iripin-1-mediated reduction of the expression of chemokine receptors in neutrophils and adhesion molecules in endothelial cells. Decreased activity of serine proteases in the presence of Iripin-1 could further impede cell migration to the site of inflammation. Finally, we determined the tertiary structure of native Iripin-1 at 2.10 Å resolution by employing the X-ray crystallography technique. In conclusion, our data indicate that Iripin-1 facilitates I. ricinus feeding by attenuating the host's inflammatory response at the tick attachment site.

• Keywords
• anti-inflammatory protein, cell migration, iripin, ixodes ricinus, serpin, tick saliva, tick-host interaction, ticks,
• MeSH
• Anti-Inflammatory Agents pharmacology   MeSH
• Chemokines MeSH
• Endothelial Cells metabolism   MeSH
• Ixodes * metabolism   MeSH
• Monocytes metabolism   MeSH
• Mice MeSH
• Serpins * metabolism   MeSH
• Trypsin MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Intramural MeSH
• Names of Substances
• Anti-Inflammatory Agents MeSH
• Chemokines MeSH
• Serpins * MeSH
• Trypsin MeSH

Tick saliva has been extensively studied in the context of tick-host interactions because it is involved in host homeostasis modulation and microbial pathogen transmission to the host. Accumulated knowledge about the tick saliva composition at the molecular level has revealed that serine protease inhibitors play a key role in the tick-host interaction. Serpins are one highly expressed group of protease inhibitors in tick salivary glands, their expression can be induced during tick blood-feeding, and they have many biological functions at the tick-host interface. Indeed, tick serpins have an important role in inhibiting host hemostatic processes and in the modulation of the innate and adaptive immune responses of their vertebrate hosts. Tick serpins have also been studied as potential candidates for therapeutic use and vaccine development. In this review, we critically summarize the current state of knowledge about the biological role of tick serpins in shaping tick-host interactions with emphasis on the mechanisms by which they modulate host immunity. Their potential use in drug and vaccine development is also discussed.

• Keywords
• anti-tick vaccine, immunomodulation, serpins, therapeutic effects, tick host interaction, tick saliva,
• MeSH
• Serine Proteinase Inhibitors physiology   MeSH
• Ticks * metabolism   MeSH
• Serpins * metabolism   MeSH
• Salivary Glands metabolism   MeSH
• Saliva metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Serine Proteinase Inhibitors MeSH
• Serpins * MeSH

It has been demonstrated that impairing protein synthesis using drugs targeted against tRNA amino acid synthetases presents a promising strategy for the treatment of a wide variety of parasitic diseases, including malaria and toxoplasmosis. This is the first study evaluating tRNA synthetases as potential drug targets in ticks. RNAi knock-down of all tested tRNA synthetases had a strong deleterious phenotype on Ixodes ricinus feeding. Our data indicate that tRNA synthetases represent attractive, anti-tick targets warranting the design of selective inhibitors. Further, we tested whether these severely impaired ticks were capable of transmitting Borrelia afzelii spirochaetes. Interestingly, biologically handicapped I. ricinus nymphs transmitted B. afzelii in a manner quantitatively sufficient to develop a systemic infection in mice. These data suggest that initial blood-feeding, despite the incapability of ticks to fully feed and salivate, is sufficient for activating B. afzelii from a dormant to an infectious mode, enabling transmission and dissemination in host tissues.

• Keywords
• Borrelia, Lyme disease, borreliosis, tRNA synthetase, tick, transmission,
• MeSH
• Acaricides pharmacology   MeSH
• Amino Acyl-tRNA Synthetases antagonists & inhibitors   genetics   MeSH
• Borrelia burgdorferi Group MeSH
• Ticks drug effects   microbiology   MeSH
• Humans MeSH
• Lyme Disease drug therapy   microbiology   transmission   MeSH
• Protein Biosynthesis drug effects   MeSH
• Drug Development MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Acaricides MeSH
• Amino Acyl-tRNA Synthetases MeSH

Hemostatic disorders are caused either by platelet-related dysfunctions, defective blood coagulation, or by a combination of both, leading to an increased susceptibility to cardiovascular diseases (CVD) and other related illnesses. The unique specificity of anticoagulants from hematophagous arthropods, such as ticks, suggests that tick saliva holds great promise for discovering new treatments for these life-threatening diseases. In this study, we combined in silico and in vitro analyses to characterize the first recombinant serpin, herein called Dromaserpin, from the sialotranscriptome of the Hyalomma dromedarii tick. Our in silico data described Dromaserpin as a secreted protein of ~43 kDa with high similarities to previously characterized inhibitory serpins. The recombinant protein (rDromaserpin) was obtained as a well-structured monomer, which was tested using global blood coagulation and platelet aggregation assays. With this approach, we confirmed rDromaserpin anticoagulant activity as it significantly delayed plasma clotting in activated partial thromboplastin time and thrombin time assays. The profiling of proteolytic activity shows its capacity to inhibit thrombin in the micromolar range (0.2 to 1 μM) and in the presence of heparin this inhibition was clearly increased. It was also able to inhibit Kallikrein, FXIa and slightly FXIIa, with no significant effect on other factors. In addition, the rDromaserpin inhibited thrombin-induced platelet aggregation. Taken together, our data suggest that rDromaserpin deserves to be further investigated as a potential candidate for developing therapeutic compounds targeting disorders related to blood clotting and/or platelet aggregation.

• Keywords
• Hyalomma dromedarii, anticoagulants, salivary glands, serpin, thrombin inhibitor,
• MeSH
• Anticoagulants chemistry   metabolism   MeSH
• Phylogeny MeSH
• Blood Coagulation drug effects   MeSH
• Ixodidae metabolism   MeSH
• Protein Conformation MeSH
• Models, Molecular MeSH
• Computer Simulation MeSH
• Amino Acid Sequence MeSH
• Serpins chemistry   metabolism   pharmacology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Anticoagulants MeSH
• Serpins MeSH

In Europe, Ixodes ricinus is the most important vector of human infectious diseases, most notably Lyme borreliosis and tick-borne encephalitis virus. Multiple non-natural hosts of I. ricinus have shown to develop immunity after repeated tick bites. Tick immunity has also been shown to impair B. burgdorferi transmission. Most interestingly, multiple tick bites reduced the likelihood of contracting Lyme borreliosis in humans. A vaccine that mimics tick immunity could therefore potentially prevent Lyme borreliosis in humans. A yeast surface display library (YSD) of nymphal I. ricinus salivary gland genes expressed at 24, 48 and 72 h into tick feeding was constructed and probed with antibodies from humans repeatedly bitten by ticks, identifying twelve immunoreactive tick salivary gland proteins (TSGPs). From these, three proteins were selected for vaccination studies. An exploratory vaccination study in cattle showed an anti-tick effect when all three antigens were combined. However, immunization of rabbits did not provide equivalent levels of protection. Our results show that YSD is a powerful tool to identify immunodominant antigens in humans exposed to tick bites, yet vaccination with the three selected TSGPs did not provide protection in the present form. Future efforts will focus on exploring the biological functions of these proteins, consider alternative systems for recombinant protein generation and vaccination platforms and assess the potential of the other identified immunogenic TSGPs.

• MeSH
• Antigens blood   immunology   isolation & purification   MeSH
• Borrelia burgdorferi isolation & purification   MeSH
• Immunization MeSH
• Tick Infestations immunology   parasitology   MeSH
• Ixodes immunology   MeSH
• Tick Bites immunology   MeSH
• Rabbits MeSH
• Humans MeSH
• Lyme Disease blood   parasitology   transmission   MeSH
• Cell Surface Display Techniques methods   MeSH
• Peptide Library MeSH
• Peptide Fragments immunology   MeSH
• Saccharomyces cerevisiae MeSH
• Cattle MeSH
• Salivary Proteins and Peptides immunology   MeSH
• Salivary Glands immunology   MeSH
• Animals MeSH
• Check Tag
• Rabbits MeSH
• Humans MeSH
• Male MeSH
• Cattle MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antigens MeSH
• Peptide Library MeSH
• Peptide Fragments MeSH
• Salivary Proteins and Peptides MeSH

Tick saliva is a rich source of pharmacologically and immunologically active molecules. These salivary components are indispensable for successful blood feeding on vertebrate hosts and are believed to facilitate the transmission of tick-borne pathogens. Here we present the functional and structural characterization of Iripin-3, a protein expressed in the salivary glands of the tick Ixodes ricinus, a European vector of tick-borne encephalitis and Lyme disease. Belonging to the serpin superfamily of protease inhibitors, Iripin-3 strongly inhibited the proteolytic activity of serine proteases kallikrein and matriptase. In an in vitro setup, Iripin-3 was capable of modulating the adaptive immune response as evidenced by reduced survival of mouse splenocytes, impaired proliferation of CD4+ T lymphocytes, suppression of the T helper type 1 immune response, and induction of regulatory T cell differentiation. Apart from altering acquired immunity, Iripin-3 also inhibited the extrinsic blood coagulation pathway and reduced the production of pro-inflammatory cytokine interleukin-6 by lipopolysaccharide-stimulated bone marrow-derived macrophages. In addition to its functional characterization, we present the crystal structure of cleaved Iripin-3 at 1.95 Å resolution. Iripin-3 proved to be a pluripotent salivary serpin with immunomodulatory and anti-hemostatic properties that could facilitate tick feeding via the suppression of host anti-tick defenses. Physiological relevance of Iripin-3 activities observed in vitro needs to be supported by appropriate in vivo experiments.

• Keywords
• Ixodes ricinus, X-ray crystallography, adaptive immunity, blood coagulation, inflammation, saliva, serpin, tick,
• MeSH
• Adaptive Immunity drug effects   MeSH
• Lymphocyte Activation drug effects   MeSH
• Anticoagulants isolation & purification   pharmacology   MeSH
• Cytokines metabolism   MeSH
• Blood Coagulation drug effects   MeSH
• Insect Proteins isolation & purification   pharmacology   MeSH
• Immunologic Factors isolation & purification   pharmacology   MeSH
• Protease Inhibitors isolation & purification   pharmacology   MeSH
• Ixodes metabolism   MeSH
• Rabbits MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Lymphocytes drug effects   immunology   metabolism   MeSH
• Guinea Pigs MeSH
• Mice, Inbred C3H MeSH
• Mice, Inbred C57BL MeSH
• Mice, Transgenic MeSH
• Mice MeSH
• Cell Proliferation drug effects   MeSH
• Spleen drug effects   immunology   metabolism   MeSH
• Salivary Proteins and Peptides isolation & purification   pharmacology   MeSH
• Saliva metabolism   MeSH
• Animals MeSH
• Check Tag
• Rabbits MeSH
• Humans MeSH
• Guinea Pigs MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Anticoagulants MeSH
• Cytokines MeSH
• Insect Proteins MeSH
• Immunologic Factors MeSH
• Protease Inhibitors MeSH
• Salivary Proteins and Peptides MeSH

Ixodes ricinus is the vector for Borrelia afzelii, the predominant cause of Lyme borreliosis in Europe, whereas Ixodes scapularis is the vector for Borrelia burgdorferi in the USA. Transcription of several I. scapularis genes changes in the presence of B. burgdorferi and contributes to successful infection. To what extend B. afzelii influences gene expression in I. ricinus salivary glands is largely unknown. Therefore, we measured expression of uninfected vs. infected tick salivary gland genes during tick feeding using Massive Analysis of cDNA Ends (MACE) and RNAseq, quantifying 26.179 unique transcripts. While tick feeding was the main differentiator, B. afzelii infection significantly affected expression of hundreds of transcripts, including 465 transcripts after 24 h of tick feeding. Validation of the top-20 B. afzelii-upregulated transcripts at 24 h of tick feeding in ten biological genetic distinct replicates showed that expression varied extensively. Three transcripts could be validated, a basic tail protein, a lipocalin and an ixodegrin, and might be involved in B. afzelii transmission. However, vaccination with recombinant forms of these proteins only marginally altered B. afzelii infection in I. ricinus-challenged mice for one of the proteins. Collectively, our data show that identification of tick salivary genes upregulated in the presence of pathogens could serve to identify potential pathogen-blocking vaccine candidates.

• MeSH
• Arachnid Vectors microbiology   MeSH
• Bacterial Vaccines administration & dosage   MeSH
• Borrelia burgdorferi Group drug effects   MeSH
• Tick Infestations genetics   microbiology   prevention & control   transmission   MeSH
• Ixodes drug effects   MeSH
• Lyme Disease genetics   microbiology   prevention & control   transmission   MeSH
• Mice MeSH
• Arthropod Proteins genetics   MeSH
• Salivary Glands microbiology   MeSH
• Transcriptome * MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bacterial Vaccines MeSH
• Arthropod Proteins MeSH

Immunodeficiency disorders and autoimmune diseases are common, but a lack of effective targeted drugs and the side-effects of existing drugs have stimulated interest in finding therapeutic alternatives. Naturally derived substances are a recognized source of novel drugs, and tick saliva is increasingly recognized as a rich source of bioactive molecules with specific functions. Ticks use their saliva to overcome the innate and adaptive host immune systems. Their saliva is a rich cocktail of molecules including proteins, peptides, lipid derivatives, and recently discovered non-coding RNAs that inhibit or modulate vertebrate immune reactions. A number of tick saliva and/or salivary gland molecules have been characterized and shown to be promising candidates for drug development for vertebrate immune diseases. However, further validation of these molecules at the molecular, cellular, and organism levels is now required to progress lead candidates to clinical testing. In this paper, we review the data on the immuno-pharmacological aspects of tick salivary compounds characterized in vitro and/or in vivo and present recent findings on non-coding RNAs that might be exploitable as immunomodulatory therapies.

• Keywords
• drug discovery, host immunity, immunomodulation, salivary glands, tick saliva,
• MeSH
• Autoimmune Diseases immunology   therapy   MeSH
• Immunomodulation immunology   MeSH
• Ticks immunology   MeSH
• Humans MeSH
• Immune System Diseases immunology   therapy   MeSH
• Arthropod Proteins immunology   MeSH
• Saliva immunology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Arthropod Proteins MeSH