Endovascular Procedures
The effect of Saratin on the endovascular healing of surgically-injured vessels was studied in rats. In this model, saratin was applied locally at the site of an open surgical endarterectomy in the carotid artery and its effect on platelet deposition acutely (3 and 24 hours) and on intimal hyperplasia at two weeks was examined. Rats were anesthetized and the carotid arteries were exposed. The rats were randomized to receive either Saratin or vehicle control. Blood flow was stopped by cross-claming the carotid artery and a surgical flap was created. The endothelium was removed by mechanical rubbing, and Saratin or vehicle was placed on the wound with a pipette (5 mL total volume, various concentrations (see below)) and allowed to set for five (5) minutes. At the end of this exposure period, the surgical flap was closed and blood flow was reestablished by removing the clamps. The area was examined for control of bleeding and the skin was closed. A representation of the surgical procedure and clinical endpoints is depicted in the figure.
Platelet adhesion to the wound site was examined at 3 and 24 hours post surgery. In addition, at the end of a two-week post-surgical period, the rats were euthanized and the carotid arteries were removed for determination of the effects of Saratin on the development of intimal hyperplasia.
Rat carotid endarterectomy injury model and end points.
Saratin caused a marked decrease in the adhesion of platelets and other cellular elements to the damaged vessel wall which was evident at 3 hours and persisted until at least 24 hours. This effect was dose-dependant, with the maximum effect occurring at saratin doses of 5 µg and above. Persistence of the effect to 24 hours and beyond may reflect the ability of saratin to inhibit the initiation of the platelet aggregation cascade by interfering with the early adhesion mediated by vWF-collagen interactions. The reduction of intimal hyperplasia and hence luminal stenosis at 2 weeks (11% stenosis on saratin vs 30% on control, P = 0.0042) suggests that saratin inhibits the longer term sequelae of this interaction and may have clinical utility in preventing vascular stenosis. Saratin also decreased the rate of thrombosis in this model (0/15 saratin-treated rats vs 5/15 controls, P = 0.0156).
In humans, cardiovascular risk is increased in patients with raised homocysteine levels. In rats fed high levels of homocysteine, intimal hyperplasia in response to carotid endarterectomy is much more pronounced, correlating strongly with plasma homocysteine levels. The mechanism for this effect is not completely understood but is believed to involve platelet adhesion. In hyperhomocysteinemic rats, saratin was found to attenuate the effect of the raised homocysteine on intimal hyperplasia and luminal stenosis back to control levels. Saratin had no effect on homocysteine metabolism.
Electron micrograph (2000X) of endarterectomized rat carotid artery 3 hours after carotid endarterectomy.
A) Control surface. B) Surface receiving topical saratin (5 µL, 1 mg/mL).
Electron micrograph (2000X) of endarterectomized rat carotid artery 24 hours after carotid endarterectomy.
A) Control surface. B) Surface receiving topical saratin (5 µL, 1 mg/mL).
This model was used to determine the optimal dose of Saratin for prevention of platelet adhesion to damaged vascular endothelial surfaces. A concentration-response curve is presented in which the optimal concentration was determined to be 1 mg/mL in this model.
Concentration-response curve of Saratin prevention of platelet adhesion in the rat carotid endarterectomy injury model.
At two weeks post surgical-induced injury, intimal hyperplasia was significantly reduced in Saratin-treated tissues compared to vehicle control-treated tissues. This highly statistically significant result is presented in the figure.
Reduction of intimal hyperplasia with the use of Saratin in the rat carotid endarterectomy injury model.
