Svetlana Kaplan

Effect of differential shear stress on platelet aggregation, surface thrombosis, and endothelialization of bilateral carotid-femoral grafts in the dog

PURPOSE The purpose of this study was to determine the effects of increased shear stress on the aggregability of platelets as they traverse a long, small-caliber (6 mm) Dacron graft in the dog and on the surface thrombosis and endothelialization of such a graft. METHODS Each of nine dogs received bilateral carotid-femoral artery grafts, approximately 75 cm long, for 3 months; one graft of each pair had a distal femoral arteriovenous fistula to produce a higher shear rate than the contralateral graft. Platelet aggregation scores were determined on blood withdrawn from the external jugular vein and from the proximal and distal ends of the grafts in each animal. Graft flow rates, which were used in the computation of shear stress, and luminal pressure gradients through grafts were measured during surgery and specimen retrieval. Specimens were studied with light microscopy after hematoxylin and eosin and immunocytochemical staining and by scanning electron and transmission electron microscopy to evaluate the nature, composition, and thickness of the flow surface lining, as well as the transmural healing. RESULTS Two high-shear stress and two low-shear stress grafts occluded unilaterally; five dogs had bilaterally patent grafts, allowing comparative analyses. All subjects had low platelet aggregability with aspirin. Platelet aggregation scores taken from proximal and distal ends of the grafts were not significantly different. The high-shear stress grafts had significantly more endothelial-like cell coverage (p < 0.0371) than the low-shear stress grafts, less flow-surface thrombus (p < 0.0056), and a thinner surface lining (p < 0.0029), on both the neointima and pseudointima. CONCLUSIONS In subjects with low platelet aggregation scores, long Dacron grafts do not elevate platelet aggregability of blood flowing through them. High-shear stress grafts have less flow surface thrombus, more endothelialization, and a thinner surface lining than do low-shear stress grafts.

The direct effect of graft compliance mismatch per se on development of host arterial intimal hyperplasia at the anastomotic interface

To study thedirect andsole effect of compliance mismatch on anastomotic intimal hyperplasia of the host arterial wall and to minimize possible confounding factors, dogs with a low thrombotic potential were selected as experimental subjects. Externally supported 6 cm × 5 mm Dacron grafts with a compliance value of approximately 1/300 of the host artery were implanted into the carotid arteries with end-to-end anastomoses on one side and end-to-side anastomoses on the other. The control graft was an autogenous carotid artery segment 4 cm in length transplanted into the femoral artery. Eight cases (24 grafts) were studied for 1 year and three (nine grafts) for 6 months. All were patent throughout the study period except for two noncompliant grafts with end-to-end anastomoses; thrombosis was the documented cause of occlusion. For the patent grafts, follow-up arteriograms showed no progressive narrowing of noncompliant anastomoses. Whether compliant or noncompliant, light microscopy studies showed slight intimal thickening within 1 to 2 mm of the anastomotic line, possibly the result of the normal healing response to stitch and surgical trauma. Quantitatively, 22 measurements representing longitudinal and circumferential thickness of the neointima were taken at each of the 40 patent noncompliant and 22 patent compliant control anastomoses. There was no statistically significant difference in anastomotic neointimal thickness in compliant and noncompliant grafts or for the different implantation periods. These data suggest that graft/host artery compliance mismatch does not cause arterial intimal hyperplasia at the anastomotic interface.

Platelet aggregometry can accurately predict failure of externally supported knitted Dacron femoropopliteal bypass grafts

PURPOSE Our purpose was to evaluate whether a method for quantification of platelet aggregability will predict failure of knitted Dacron femoropopliteal bypass grafts. METHODS A numerically derived platelet aggregation (PA) score, based on the aggregation pattern and platelet count, was determined in the 40 patients available for platelet analysis who underwent 53 femoropopliteal bypass grafts with preclotted, 6 mm, externally supported knitted Dacron grafts from 1981 to 1991 (mean follow-up 50 months). The preoperative score was found to remain stable after surgery, enabling the use of postoperative values when preoperative values were not available. The PA score was available in 19 patients (23 grafts) before surgery and 23 patients (30 grafts) after surgery. The following factors were analyzed for predicting graft failure by the Cox proportional hazards regression model: PA score, age, gender, history of smoking, coronary artery disease, hypertension, hyperlipidemia, cerebrovascular disease, diabetes, claudication versus limb salvage, site of the distal anastomosis, previous ipsilateral bypass, and state of the runoff as determined by preoperative angiography. RESULTS Of the studied risk factors, the value of the PA score was the most significant predictor of graft closure (p < 0.0001). An increase of 10 units was associated with an increased relative risk of 2.02. Throughout the follow-up period, 15 of 16 grafts remained patent in patients with a PA score of 15 or less, but only 2 grafts out of 17 remained patent in patients with a PA score of 30 or greater. CONCLUSIONS These data suggest that the PA score is a potential risk factor for failure of femoropopliteal bypass with externally supported knitted Dacron grafts.

Aspirin therapy in small-caliber arterial prostheses: Long-term experimental observations*

To study the therapeutic effects of 3 mg/kg aspirin given at the time of surgery and postoperatively, Dacron carotid grafts with an internal diameter of 4 mm and a length of 6 cm were implanted bilaterally in mongrel dogs. Sixteen control grafts in eight subjects and 20 grafts in 10 subjects treated with aspirin were followed by serial angiograms until consecutive studies showed stable patency rates in both groups. Platelet aggregations, malondialdehyde production, serum salicylate levels, and thromboxane A2 and prostacyclin secretion (measured as thromboxane B2 and 6-keto-prostaglandin F1 alpha) were monitored prior to and throughout the experiment. Surface mapping, indium-111 uptake, factor VIII-related antigen staining, and scanning and transmission electron microscopy were performed on the grafts at sacrifice. This study demonstrates a protective effect on the early patency of small-caliber prostheses in the canine model with daily oral aspirin administration. The degree and duration of this effect depends on the preoperative baseline ratio of thromboxane to prostacyclin in each subject.

Implant site: a determinant of completeness of arterial prosthesis healing in the dog and possibly in humans.

This paper compares the healing of supported knitted Dacron prostheses implanted in the descending thoracic aorta and in the subcutaneous carotid-femoral positions in each of 10 dogs. The descending thoracic aorta prostheses were 6 cm long and the carotid-femoral prostheses averaged 76 cm in length. The healing of four porous clinical axillofemoral grafts is compared to that of the experimental carotid-femoral grafts. There was little similarity between the healing observed in the descending thoracic aortic and subcutaneous positions; the descending thoracic aorta grafts healed rapidly and completely by eight weeks, with thrombus-free flow surfaces covered with confluent endothelial cells. However, the inner wall of the carotidfemoral grafts remained largely unhealed at one year, with endothelialization largely confined to a limited pannus ingrowth across the anastomoses. This indicates that the implant site in the dog is a primary determinant of the rapidity and completeness of healing. The clinical axillofemoral grafts explanted after 20 months to six and one-half years exhibited healing characteristics that were similar to those seen in the carotid-femoral position in the dog. These findings suggest that valid comparisons of interspecies healing require data obtained from similar implant locations. They present a challenge to the previously held concepts of major interspecies differences in speed and extent of tissue ingrowth based on data obtained from noncomparable implant sites.

Glutaraldehyde preparation of coronary artery bypass bioprostheses

A method for glutaraldehyde (GA) fixation of canine carotid arteries has been developed for the preparation of small caliber biologic prostheses for coronary artery bypass. The biologic grafts were preserved by a static inflation technique that proved to be more advantageous than the standard stenting method. The most suitable static inflation pressure was found to be 120 mm Hg. By means of colorimetric measurements the minimal tanning time and the amount of GA required for complete fixation for canine vascular tissue were established. Stabilization of the vessel collagen and confirmation of GA-collagen cross-linking were verified by evaluation of the elastic properties and shrinkage temperature of the grafts. Stress-strain measurements were evaluated to determine the number of cross-links introduced in the vascular tissue by GA. This number was shown to be proportional to the inflation pressure. Ethyl alcohol was chosen as the storage solution because it maintained the best physical, chemical, and histologic characteristics of the grafts. Biological evaluations were performed with carotid implants that were examined following acute low flow studies and implantations up to 112 days. All implantations have yielded 100% patencies.

Implant site influence on arterial prosthesis healing: A comparative study with a triple implantation model in the same dog

PURPOSE The purpose of this study was to develop a cost-effective canine graft healing model that gives information on various implant sites and controls for variable factors between graft locations and between animals and to compare the influence of implant site (retropleural, retroperitoneal, and subcutaneous areas) on arterial graft healing in the same subject under such controlled study conditions. METHODS Five mongrel dogs were studied for 8 weeks, and one was studied for 3 years. Each received three porous Dacron grafts during the same surgery: a carotid-femoral bypass (C-FB) and interposition grafts in the descending thoracic aorta and abdominal aorta. To produce comparable shear stress calibers of the C-FB and abdominal aorta grafts were 2 mm less than those of the descending thoracic aorta, and a distal arterio-venous fistula was created to further increase the C-FB flow. For comparable blood aggregation status platelet aggregation was preevaluated and adjusted with antiplatelet agents. Graft flow surfaces were assessed for thrombus-free surface and endothelial-like cell coverage scores. Tissue samples were studied with hematoxylin-eosin, factor VIII/ von Willebrand factor, smooth muscle alpha-actin staining, and scanning electron microscopy and transmission electron microscopy. RESULTS All grafts were patent. Shear stress for the three grafts and platelet aggregation among the study subjects were comparable. Healing of descending thoracic aorta and abdominal aorta grafts was similar, but C-FB healing was slow, incomplete, and uneven, with a high incidence of seroma. Eight-week and 3-year results were comparable. CONCLUSIONS This model gives broad healing information about the areas where grafts are often implanted in humans. Eight weeks appears to be a sufficient period to reflect basic and general healing characteristics. Grafts heal better in the retropleural and retroperitoneal areas than in the subcutaneous tissues.

Reendothelialization of isolated segments of the canine carotid artery with reference to the possible role of the adventitial vasa vasorum

This study was designed to determine if the adventitial vasa vasorum contribute to re-endothelialization of the canine carotid artery after removal of the endothelial flow surface. Casting studies demonstrated that vasa vasorum are present only in the adventitia of the canine carotid artery. Ninety autograft segments of the carotid artery from which the endothelium had been removed were implanted in both carotid arteries of 45 dogs. Glutaraldehyde-processed canine carotid allografts were positioned at each end to prevent pannus ingrowth and a Gore-Tex wrap to prevent periarterial tissue growth was placed into the outer wall. Three methods were used, with observations at 4 and 8 weeks. In method 1 the test segment was treated with superficial endarterectomy. In method 2 a balloon catheter was used to remove the endothelium. In method 3 balloon catheter denudation of the flow surface was also employed and, in addition, the adventitia was removed surgically as completely as possible, although a few vasa vasorum were shown to remain in some grafts. In method 1 all the patent endarterectomized arteries were partially re-endothelialized at both the 4- and 8-week intervals. In method 2, 72% of the balloon-denuded patent arteries with intact adventitial vasa vasorum were partially re-endothelialized at 4 weeks and 84% at 8 weeks. However, in method 3 the flow surfaces had no endothelium at 4 weeks and 83% still had none at 8 weeks. These findings suggest that, in the absence of pannus ingrowth, re-endothelialization of the canine carotid artery depends on not only the presence but also the number of adventitial vasa vasorum.

The Evolution of a Clinically Relevant Experimental Model for the Realistic Evaluation of Long-Length (80-cm) Arterial Prostheses

A new carotid-femoral experimental model has been developed which has significant clinical relevance to human limb salvage bypass grafting in the areas of graft length, hemodynamics and tissue environment. More significantly, the graft healing pattern observed with this model is similar to what is seen in humans. Therefore, this experimental model provides a challenging and critical site for dynamic healing studies and comparisons of different types of vascular prostheses.

Citric Acid Enhances the Antithrombotic Effect of Aspirin in Many Aspirin-Resistant Subjects

This study had three objectives: (1) to determine the frequency of high platelet aggregators in a consecutive series of 268 apparently healthy volunteers who presented to our Center; (2) to assess the inhibitory effect of aspirin (ASA) on these high aggregators; (3) to determine, in a double-blind trial, whether or not the addition of citric acid (CTA) to ASA would increase its inhibitory effect in subjects who had a suboptimal response to aspirin alone. A platelet aggregation-scoring methodology developed for turbidimetric platelet aggregometry was used to quantify baseline aggregation and medicinal effects. We define a high aggregator as one whose unmedicated PA score is ≥30. We define the response of a high aggregator to ASA as poor if the medicated PA score stays at ≥30. We found that 58 of 268 apparently healthy unmedicated volunteers (22%) had PA scores ≥30. and that 27 of these (47%) had a poor response to 325 mg ASA, with an average drop in their PA scores from 49.5 ± 13.1 to 41.1 ± 8.6 (16%). Twenty-five of these 27 people were enrolled in the double-Mind study comparing the effect of ASA and ASA + CTA on platelet aggregability. Of these high aggregators who had a poor response to ASA, 12 of 25 (50%) had a good response to 162.5 mg of ASA plus 162.5 mg of CTA, with an average drop of their PA scores from 46.7 ± 13.2 to 22.0 ± 5.2 (53%). CTA alone had no effect on the PA score, which was similar to the control placebo. Our data suggest that a 1:1 combination of ASA and CTA may offer significantly greater protection agairtst arterial thrombotic events than ASA alone in subjects who respond poorly to ASA. Key Words: Platelet aggregation—Antithrombotic medication—Thrombosis.