Ixonnexin from Tick Saliva Promotes Fibrinolysis by Interacting with Plasminogen and Tissue-Type Plasminogen Activator, and Prevents Arterial Thrombosis

Tick saliva is a rich source of modulators of vascular biology. We have characterized Ixonnexin, a member of the “Basic-tail” family of salivary proteins from the tick Ixodes scapularis. Ixonnexin is a 104 residues (11.8 KDa), non-enzymatic basic protein which contains 3 disulfide bonds and a C-terminal rich in lysine. It is homologous to SALP14, a tick salivary FXa anticoagulant. Ixonnexin was produced by ligation of synthesized fragments (51–104) and (1–50) followed by folding. Ixonnexin, like SALP14, interacts with FXa. Notably, Ixonnexin also modulates fibrinolysis in vitro by a unique salivary mechanism. Accordingly, it accelerates plasminogen activation by tissue-type plasminogen activator (t-PA) with Km 100 nM; however, it does not affect urokinase-mediated fibrinolysis. Additionally, lysine analogue ε-aminocaproic acid inhibits Ixonnexin-mediated plasmin generation implying that lysine-binding sites of Kringle domain(s) of plasminogen or t-PA are involved in this process. Moreover, surface plasmon resonance experiments shows that Ixonnexin binds t-PA, and plasminogen (KD 10 nM), but not urokinase. These results imply that Ixonnexin promotes fibrinolysis by supporting the interaction of plasminogen with t-PA through formation of an enzymatically productive ternary complex. Finally, in vivo experiments demonstrates that Ixonnexin inhibits FeCl3-induced thrombosis in mice. Ixonnexin emerges as novel modulator of fibrinolysis which may also affect parasite-vector-host interactions

Amylin induces hypoglycemia in mice

Amylin is a 37-aminoacid pancreatic protein that exerts control over several metabolic events such as glycemia and lacticemia. Amylin has long been shown to induce increases in arterial plasma glucose. We decided to investigate whether amylin plays additional roles in the glucose metabolism. We evaluated glucose homeostasis using whole blood from the tail tip of fasting, conscious, unrestrained normal and streptozotocyn-induced diabetic mice following subcutaneous administration of mouse amylin. Subcutaneous injection of 1 μg mouse amylin caused a transient decrease in whole blood glucose in both normal and diabetic mice in the absence of insulin. The blood glucose levels were lowest approximately 2 hours after amylin administration, after that they gradually recovered to the levels of the control group. The hypoglycemic effect followed a dose-dependent response ranging from 0.1 to 50 µg / mouse. These results reveal the ability for amylin in the direct control of glycemia at low doses in the absence of insulin

Dipetalodipin and triplatin are antithrombotic <i>in vivo</i>.

<p>Thrombosis was induced in the carotid artery of mice via local application with 7.5% FeCl₃. Blood flow was monitored with a perivascular flow probe for 60 min or until stable occlusion occurred. (A) Dipetalodipin or (B) triplatin was injected into the caudal vein 15 min before injury. Each symbol represents one individual. *<i>P</i> < 0.05 vs control, **<i>P</i> < 0.01 vs control; ANOVA with the Dunnett posttest.</p

Dipetalodipin and triplatin abolish the collagen-mediated acceleration of PRP clotting.

<p>Human citrated-anticoagulated (A) PRP or (B) PPP supplemented with PC/PS was pretreated with dipetalodipin (1 μM) or triplatin (2 μM). Preparations were then incubated with collagen (50 μg/ml, final concentration) or vehicle solvent (control) and activated with 16.6 mM CaCl₂. Mean ± SEM (n = 5); *<i>P</i> < .05; **<i>P</i> < .01; ***<i>P</i> < .001; NS, non-significant; analysis of variance (ANOVA) with Tukey's posttest.</p

Effect of dipetalodipin and triplatin on the pulmonary embolism model.

<p>(A-B) Kaplan-Meier survival curves. Mortality associated with i.v. injection of collagen (0.8 mg/kg) and epinephrine (60 μg/kg) after administration of PBS, (A) dipetalodipin or (B) triplatin. Animals still alive 30 min after the challenge were considered survivors. **<i>P</i> < 0.01 vs control (log-rank test).</p