**3.1. Components affecting coagulation**

Ticks saliva has other strategies besides inhibiting blood coagulation factors, in order to facilitate the feeding. After injury, subendothelial tissue get exposed, activated platelets bind to exposed von Willebrand factor and collagen through its surface receptors and platelets release soluble vasoconstrictor mediators (ADP, serotonin, and thromboxane A2 ). Physiologically, there are three major mechanisms that regulate anticoagulation: TFPI, antithrombin III (ATIII), and protein C/thrombomodulin/activated protein C. Until now, there are no description of tick saliva components interfering with or imitating antithrombin, protein S, protein C, heparin, or thrombomodulin [24]. However, many ticks can inhibit thrombin-induced platelet aggregation. On the other hand, anticoagulant molecules from tick saliva also regulate hemostasis by inhibiting blood coagulation factors (FXa or thrombin) or tenase complexes (FVIIa/ TF and FIXa) and/or platelet aggregation [132].

fibrinogen or other ligands to platelet receptors such as savignygrin from *O. savignyi* [141]. Variabilin is another anti-platelet RGD-containing peptide from *Dermacentor variabilis* [142]. Some inhibitors identified in *I. pacificus* and *I. scapularis*, known as ixodegrins, display some differences with variabilin by having cysteines flanking the RGD motif, and with savignygrin,

Anticoagulants from Hematophagous http://dx.doi.org/10.5772/intechopen.78025 51

Other anti-platelet molecules from ticks were reported: monogrin from *Argas monolakensis* [144], moubatin, a lipocalin derived from *O. moubata* which inhibits collagen-induced platelet aggregation by scavenging thromboxane A2 [139, 140, 144], longicornin, isolated from the salivary gland of *Haemaphysalis longicornis*, which also inhibits collagen-mediated platelet

To target blood coagulation, components from tick saliva have inhibitory activities on the extrinsic tenase complex in blood coagulation [132]. From the studies in *I. scapularis* tick (Acari: Ixodidae) [146], two classes of extrinsic tenase complex inhibitors were identified acting similarly, but not identically, to the physiological inhibitor, tissue factor pathway inhibitor (TFPI) [136]. The first group is represented by ixolaris [147], a 15.7 kDa molecule obtained from the cDNA library of the salivary glands of *I. scapularis* consisting of 140 amino acid residues containing 10 cysteine and two-Kunitz tandem domain which does not bind to FXa active site, in contrast TFPI. It was hypothesized that the second Kunitz domain of ixolaris binds first to FX/FXa (on a heparin binding proexosite/exosite) before binding to the FVIIa-TF complex via the first Kunitz domain. The native inhibitor has a molecular mass of 24 kDa, and both forms are equally effective as anticoagulants. Functionally, the Ixolaris is structurally distinct from human tissue factor pathway inhibitor (TFPI) [146]. The second group is represented by penthalaris [148], a five-Kunitz tandem domain which uses FX or FXa as scaffold to inhibit the FVIIa-TF complex.

One of the main classes of FXa inhibitors characterized from soft tick saliva is the atypical, non-canonical Kunitz-type inhibitors including the tick anticoagulant peptide (TAP), obtained from the *Ornithodoros moubata* tick [9] and FXa-inhibitor (FXaI) from *O. savignyi* tick [149] (Acari: Argasidae). Both inhibitors possess a single Kunitz domain, in contrast to the tandem Kunitz type thrombin inhibitors. Kinetically, both are slow, tight-binding, competitive inhibitors of FXa. The recombinant (rTAP) TAP has a single-chain acidic polypeptide composed of 60 amino acids including 6 cysteine residues, and is a competitive FXa inhibitor highly selective and reversible. Its molecular weight is 6.8 kDa, pI 4.5 and Ki of 0.588 for the native form, and Ki of 0.18 nM for the recombinant form, expressed in *Saccharomyces cerevisiae* [150–152].

Amblyomin-X is a FXa inhibitor identified molecule in the transcriptomics profile of the salivary glands by Expressed Sequence Tags (ESTs) from the hard tick *Amblyomma cajennense* (currently *Amblyomma sculptum*) [153], containing an unique structure with a N-terminal Kunitz-type domain of 60 amino acids and a C-terminal with 49 amino acids. Amblyomin-X is able to inhibit factor Xa, prothrombinase and tenase activities. As FXa inhibitor, Amblyomin-X

which have a non-canonical RGD peptide inserted into a Kunitz fold [127, 143].

aggregation [145].

*3.1.2. Tenase complex inhibitors*

*3.1.3. Factor Xa inhibitors*

Anticoagulants from tick saliva can be classified according with their biochemical characteristics and structure, some of them belonging to the Kunitz-type domain inhibitors and Serpin domain inhibitors [127]. Members of those families can modulate coagulation, inflammation, or vasoconstriction. For example, the Serpin IRS-2 (*I ricinus* Serpin-2) from *I. ricinus* inhibits cathepsin G and chymase, both known as mediators of platelet aggregation and inflammation [133], as well as to mediate vascular permeability [25]. Besides, Kunitz domain inhibitors are widely expressed and characterized as anticoagulants, some of them having just one Kunitz domain being able to inhibit factor Xa [134] or thrombin, such as savigin from *Ornithodoros savignyi* [135].

Depending on the mechanism of action, they can include platelet inhibitors, factor Xa inhibitors and thrombin inhibitors, since they are able to prevent blood clotting and maintain blood incoagulable. Those blood coagulation inhibitors from tick are the major focus of this section.
