Jan B. Wieslander, Lars Salemark, and Peter Dougan
HYDROXYETHYL STARCH INCREASES PATENCY AND REDUCES THROMBUS INTIMECTOMY IN SMALL ARTERIES: A N EXPERIMENTAL STUDY IN THE RABBIT ABSTRACT Twenty-four arteries of rabbit ears, divided into two groups of 12 vessels each, were prepared and 32Plabelled platelets were infused. Arteriotomy/intimectomy was performed after 1 hr and in vivo platelet accumulation recorded for 2 hr. Group A comprised untreated control animals and group B was treated with 1 g hydroxyethyl starch (HES), MW 450,000 in 17 ml saline/kg b.w. (Plasma-steril®). Vessel bleeding-times were normal, patency was improved, and intraluminal thrombotic material was reduced after HES treatment. Initial in vivo platelet accumulation was rapid and reached similar levels in both groups. However, the platelet accumulation curves decreased more frequently following HES than in the control group. HES does not prevent platelet accumulation at trauma sites, but reduces the sizes of the thrombi formed and may enhance disaggregation/fibrinolysis.
A number of colloid solutions are used for blood and plasma replacement and for improving tissue perfusion. Of these, dextran, originally introduced for volume substitution, was later found to be an effective agent against postoperative pulmonary embolism.1 Dextran is therefore widely used today as an antithrombotic agent, and earlier studies by our group have confirmed that it is an effective antithrombotic agent following trauma of different degrees of severity in small rabbit arteries.2 3 Hydroxyethyl starch (HES) is another colloid whose effects on volume substitution and tissue perfusion are well-documented,45 and there is some evidence that it causes less bleeding6 7 and fewer allergic reactions than dextran.4 However, the influence of HES on thrombus formation in microvascular surgery has
not previously been investigated. The aim of this study was to examine the effects of HES in vivo on platelet function, thrombus formation, and patency rates, following severe trauma in small rabbit arteries. To facilitate comparison with earlier results, the same techniques were used as in studying dextran 70 and saline solutions.2
MATERIALS AND METHODS Rabbits, males and females weighing 2.5 to 5.5 kg, were kept on a standard pellet diet and given water ad libitum. They were anesthetized intravenously with sodium pentobarbital (Mebumal® 60 mg/ml, ACO Lake- 357
Department of Experimental Research and Department of Plastic and Reconstructive Surgery, Malmo General Hospital, Malmo, Sweden Reprint requests: Dr. Wieslander, Dept. of Plastic and Reconstructive Surgery, Malmo General Hospital, S-214, 01 Malmo, Sweden Accepted for publication lune 3, 1990 Copyright © 1990 by Thieme Medical Publishers, Inc., 381 Park Avenue South, New York, NY 10016. All rights reserved.
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FORMATION FOLLOWING ARTERIOTOMY/
JOURNAL OF RECONSTRUCTIVE MICROSURGERY/VOLUME 6, NUMBER 4
Arleriotomy 7 mm
^ *~;
1 mm
Figure 2. The exposed artery is positioned in a double vascular clamp and a 7-mm long arteriotomy made using a microscope and microscissors. Intimectomy is next performed at high magnification on the central 5-mm segment using a surgical blade, and the arteriotomy is closed with a running microsuture. Blood flow is then reestablished by removing the clamp. The same clamp is used in all cases.
transferred to another Fenwal bag. Platelet-poor plasma (PPP) was prepared by centrifuging for later use in washing labelled platelets. Twenty-five MBq (1 millicurie = 37 MBq) of the beta-emitter 32P (phosphorus) as orthophosphate in dilute hydrochloric acid were dissolved in modified Ringer's solution (ACO Lakemedel, Solna, Sweden), added to the PRP in the Fenwal bag, and the resulting mixture incubated for 2 hr at 37°C in a rocker. The solution was removed after centrifuging and 15 ml of PPP added. The contents of the plastic bag were homogenized and PPP removed after centrifuging. Washing in this manner was carried out three times. Finally, a small amount of PPP (3 to 4 ml) was added, and the solution of 32P-labelled platelets infused into the rabbit being studied. The stability in blood of 32P-labelled platelets labelled in this way was studied earlier and COMPUTER FEN RECORDER found to be satisfactory.8 RADIOACTIVITY. The prepared arteries were placed in grooves in chromium-plated brass holders at body activity signal Ringor s solution temperature, and thin-window beta-counting GeigerMuller (GM) tubes (Philips ZP 1410) were positioned just above them (see Fig. 1) to detect emitted radiation. Care was taken to remove any blood present. Throughout the measurements, repeated checks were made to insure that radioactive blood had not leaked from the prepared vessel segments or surrounding skin/wound edges into the field-of-view of the detectors. The sigSKIN /CARTILAGE FLAP nals from the GM tubes were monitored continuously on a pen recorder and stored for later analysis using Figure 1. The exposed central artery of the ear is sealers (Canberra Model 2072) on-line to a Luxor moistened, wrapped in plastic foil, and placed in a groove in ABC806 computer. Independent counting channels a chromium-plated brass holder maintained at 39.8°C. The permitted measurements to be made on both ears radioactivity originating from 32P-labelled platelets in the simultaneously. vessel segment studied is registered on a Geiger-Muller Two hours after reperfusion, the segments of vesdetector. The output signals are monitored on a pen re358 corder and stored for later analysis using a computer. sel containing the arteriotomy/intimectomy regions
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medel, Solna, Sweden), and anesthesia maintained by intermittent injections of the same compound. During the measurements, the animals were maintained at 39.8 ± 0.2°C by thermostatically-regulated heating pads. Other than the local application of lidocaine (Xylocaine®, 10 mg/ml, AB Astra, Sodertalje, Sweden) to release vascular spasm, neither anticoagulants nor vasodilating agents were used Ringer's solution at body temperature was used as a local irrigant. The central arteries (diameters 0.8 to 1.2 mm) of the ears of 14 rabbits were prepared (Fig. 1). Platelets labelled with 32P, as described below, were infused into the aorta via a catheter inserted into the arteria femoralis. HES solution was next infused over an interval of 20 to 30 min. About 1 hr later, longitudinal arteriotomy (7 mm) followed by intimectomy (5 mm) were performed under magnification using a microscope (Fig. 2). Conventional microsurgical instruments were used, arteriotomy being performed with microscissors and intimectomy with a surgical blade using a standardized technique. The arteriotomy was closed with a running 10-0 monofilament nylon suture (Dermalon 10-0, TE 70, Davis & Geek, American Cyanamid Co, Pearl River, NY, USA) and blood flow reestablished by removing the vascular clamp. PLATELET LABELLING. A plastic bag (Transfer Pack Unit, Fenwal Laboratories, Deerfield, IL, USA) was prepared with 20 ml sodium citrate solution. A plateletdonor rabbit was anesthetized by intravenous administration of sodium pentobarbital and 100 to 150 ml whole blood drawn via an intraaortically-positioned cannula. A plastic tube connected the cannula to the bag, which was gently shaken during sampling to facilitate the mixing of sodium citrate and blood. Platelets were isolated as platelet-rich plasma (PRP), using standard centrifugation techniques, and
OCTOBER 1990
HyDROXYETHYL STARCH/WIESLANDER, SALEMARK, DOUGAN
RECORDING OF ARTERIOTOMY/INTIMECTOMY PLATELET ACCUMULATION. Platelet labelling, detecting of radio-
activity, and the experimental set-up have been described above. The trauma model used has been shown to be both reproducible and capable of yielding qualitative and quantitative information about platelet accumulation in the immediate post-reperfusion time period. RED THROMBOTIC MATERIAL. Intramural red thrombotic material was qualified as - / + (clean or small amounts), + + (moderate amounts) or + + + (large amounts). The amounts of material were estimated using a microscope at high magnification. HEMATOCRIT VALUES. In Group B, hematocrit values were measured before infusion of Plasmasteril®, at 10 min postinfusion, and then at hourly intervals for a further two hours (Table 1).
RESULTS BLEEDING TIMES. The bleeding time in the control group was 5±] min (median and quartiles) and in the HES group 4±j min (median and quartiles). Statistical comparison using the Mann-Whitney U-test showed that the bleeding times in the untreated control and HES groups were completely indistinguishable. PATENCY (Fig. 3). In the control group, good patency was recorded in five and poor or no patency in seven vessels. All 12 vessels in the HES group were patent, but two (both in one rabbit) had reduced patency. Comparison of these values using Fisher's Exact Test shows that the groups are significantly different (p < 0.05). PLATELET ACCUMULATION. Qualitative Features of the Platelet Accumulation Curves. In the untreated control group, activities in all cases increased rapidly just after restoration of blood flow and later decreased in five, remained constant in five, and increased in two vessels. In general, the accumulations decreased in vessels with good patency and were constant in those with reduced patency. In the HES group, activities also increased rapidly after releasing the vascular clamps and later decreased in 11 of the 12 vessels studied. Activity in the twelfth case, one of the two vessels with reduced patency, remained constant.
Quantitative Platelet Accumulation (Fig. 4). After re-
leasing the vascular clamps, the median values of radioactivity in both groups increased rapidly to 500 to 600 percent of their preoperative values. The median value in the control group remained high while that in the HES group decreased during the 2 hr radiation
No of vessels
• Control £23 HES
10 8 6
Tablet. Rabbit
O-Sample
210 211 212 213 214
34 30 19 30 28
Mean
(100%) (100%) (100%) (100%) (100%)
Hematocrit Values 10 min 23 22 14 18 18
(68%) (74%) (74%) (60%) (64%)
68%
70 min 29 22 14 16 21
(85%) (74%) (74%) (53%) (75%) 75%
130 min 31 22 15 13 23
(91%) (74%) (79%) (43%) (82%)
73%
The results are listed above as absolute values and as percentages of initial values. The degree of hemodilution obtained using HES is thus 20 to 30 percent. HES infusions were started immediately afater taking the 0-samples.
4 2
Good
Poor/No
Figure 3. Patency. Treatment with HES increases patency rates, compared to the control group. "Good" denotes rapid refilling; "Poor," slow refilling; and "No," no refilling when patency is checked using the "empty/refill" test.
359
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were excised and the levels of residual background activity recorded. Relative values of platelet accumulation in the two groups, expressed as the ratios of the counting rates in the postoperative periods to the "steadystate" preoperative values in percent, at intervals of 15 min post-reperfusion, were derived and compared statistically using the Mann-Whitney U-test. GROUPS. Group A was an untreated control group of 12 vessels in eight rabbits. Group B comprised 12 vessels in six rabbits treated with 1 g hydroxyethyl starch MW 450,000 in 17 ml saline/kg b. w. (Plasmasteril®, 6 percent in NaCl, Fresenius AG, Bad Homburg, FRG) administered through an intraaorta catheter about 2 hr prior to vessel reperfusion. BLEEDING TIMES. The time intervals elapsing between restoring blood flow (release of vascular clamps) and cessation of bleeding from the arteriotomies were noted in minutes. PATENCY. Vessel patency was determined at the end of all experiments and recorded as either good (rapid refilling) or poor/not patent (slow or no refilling) using the empty/refill test, performed as follows. Using a pair of microforceps, the vessel was occluded just distal to the arteriotomy region and emptied of blood over a short distance using a second pair of microforceps. The first pair of microforceps was then opened and patency assessed using the above criteria for refilling.
JOURNAL OF RECONSTRUCTIVE MICROSURGERY/VOLUME 6, NUMBER 4 OCTOBER 1990 °
No of vessels
CONTROL
Control 10 o
HES
8
o
6 4 Median values Quartile ranges
—
2
|Arteriotomy
minutes
Figure 5. Thrombotic material. Decreased amounts of red thrombotic material were observed following HES infuFigure 4. \n vivo accumulation of 32P platelets at arteri- sion. The classification used is: - clean, + small amounts, + + otomy/intimectomy trauma sites. Median values and inter- moderate amounts, and + + + large amounts of material. quartile ranges of the post-reperfusion increases in platelet activity. The values are plotted as percentages of the steadystate values. Platelet accumulation at trauma sites was rapid and reached about the same level for both groups initially. The larger proportion of decreasing curves in the HES group is reflected by the gradual decrease in its median value. The groups are, however, statistically indistinguishable. intimectomy
360
detection period. There were no statistically significant ence of dextran 70 has been reported to be deficient, differences in platelet accumulation between the groups. with reduced tensile strength compared to untreated RED THROMBOTIC MATERIAL (Fig. 5). Red thromcontrol animals 1213 and it has been suggested that botic material was observed in all vessels. In Group A, HES has a similar fibroplastic effect.14 The potential the material visible at inspection was quantified as 3 usefulness of HES as an antithrombotic agent has, - / + , 5 + + and 4+ + + and in Group B as 9 - / + , 2 + + however, not been documented. and 1 + + + . Comparison of these values using the Our results show that, compared to the control Mann-Whitney U-test with an ordinal scale yields p < group, HES treatment does not increase vessel bleed0.05, so that the deposits are significantly smaller ing time, while in dextran-treated animals, the correwhen HES is present. sponding bleeding time is considerably prolonged.2 Dextran 70 and HES cause similar degrees of hemodilution, a colloid effect known to increase peripheral flow rates, which is possibly an important DISCUSSION factor in reducing thrombus deposition and improving patency. In our model, treatment with HES or dextran The particular trauma model used (arteriotomy/ 70 leads to considerable improvements in patency, intimectomy) was chosen because it simulates the consistent with the reduced amounts of thrombotic clinical microvascular trauma occurring, for example, material observed. That similar numbers of platelets in avulsion injuries, and to facilitate comparison with accumulate at reperfusion as rapidly as in untreated saline and dextran solutions previously tested using controls shows that platelets apparently function normally (adhere and aggregate) even in the presence of the same model. HES is fairly new and interesting colloid reported these substances. This suggests that the positive efto be a clinically safe plasma volume expander.7 Unlike fects the agents produce must occur at later stages of dextran, it does not introduce clinical bleeding diathe- thrombus deposition, e.g., enhanced fibrinolysis. The ses6-7 at doses of less than 1.5 g/kg b. w./day unless a effect must be specific to these colloids, since saline hemostatic defect, e.g., von Willebrand's disease, is solution (used as carrier of the colloids) does not 2 already present.9 Like dextran,10 it has been reported produce antithrombotic effects. to enhance fibrinolysis.1' The fibrin formed in the presTo summarize, our study shows for the first time
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Steady s
HYDROXYETHYL STARCH/WIESLANDER, SALEMARK, DOUGAN
that HES, without endangering hemostasis, produces antithrombotic effects and leads to improvements in patency and related parameters in severely traumatized small arteries.
6. 7. 8.
REFERENCES Gruber UF: Prevention of thromboembolic complications The problem and alternatives. In Lewis DH (ed): Dextran—30 Years. Stockholm: Almqvist & Wiksell, 1977, p 55 Wieslander IB, Dougan P, Stjernquist U, Mecklenburg C: Effect of dextran 70 and saline on thrombus formation following arteriotomy and intimectomy in small arteries. Microsurg 7:168, 1986 Wieslander IB, Dougan P, Stjernquist U, et al: The influence of dextran and saline solution on platelet behaviour following microarterial anastomosis Surg Gynecol Obstet 163256, 1986
9. 10. 11. 12. 13
Mishler JM: Synthetic plasma volume expanders—Their pharmacology, safety and clinical efficacy. Clin Haematol 13:75, 1984 Munsch CM, Macintyre E, Machin SI, et al.: Hydroxyethyl starch: An alternative to plasma for post-operative volume expansion after cardiac surgery. Br ) Surg 75:675, 1988 Wieslander IB, Aberg M, Dougan P: Accumulation of isotopelabelled platelets in small arteries after end-to-end and end-in-end anastomoses in the rabbit Br I Plast Surg 35: 158, 1982 Strauss RG, Stump DC, Henriksen RA: Hydroxyethyl starch accentuates von Willebrand's disease. Transfusion 25:235, 1985 Carlin G, Saldeen T: On the interaction between dextran and the primary fibrinolysis inhibitor, a 2 -antiplasmin. Thromb Res 19:103, 1980 Strauss RG, Stump DC, Henriksen RA, Saunders R. Effect of hydroxyethyl starch on fibrinogen, fibrin clot formation and h'brinolysis. Transfusion 25:230, 1985 Aberg M, Bergentz SE, Hedner U: Effect of dextran on the iysability of ex vivo thrombi. Ann Surg 181:342, 1975 Carlin G, Viik K-O, Arfors K-E, et al.: Influences on the formation and structure of fibrin. Thromb Res 9:623, 1980
senius Stiftung, 1981 Kohler H, Zschiedrich H, Clasen R, et al: Blutvolumen, Kolloidosmotischer Druck- und Nierenfunktion von Probanden nach Infusion mittelmolekularer 10% Hydroxy-ethyl Starke 200/0.5 und 10% Dextran 40. Anaesthesist 31:61, 1982
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GahvVR: Aktuelle Volumentherapie mit Kolloidalen Losungen. Wissenschaftlige Informationen, Anestesie 2/81, Bad Homburg: Fre-
This research was supported by grants from the Swedish Medical Research Council (B87-17X-00759-22B), the Medical Faculty, University of Lund, Lund, Sweden, and Syskonen Svensson's Foundation, Malmo, Sweden
361
HYDROXYETHYL STARCH INCREASES PATENCY AND REDUCES THROMBUS INTIMECTOMY IN SMALL ARTERIES: A N EXPERIMENTAL STUDY IN THE RABBIT ABSTRACT Twenty-four arteries of rabbit ears, divided into two groups of 12 vessels each, were prepared and 32Plabelled platelets were infused. Arteriotomy/intimectomy was performed after 1 hr and in vivo platelet accumulation recorded for 2 hr. Group A comprised untreated control animals and group B was treated with 1 g hydroxyethyl starch (HES), MW 450,000 in 17 ml saline/kg b.w. (Plasma-steril®). Vessel bleeding-times were normal, patency was improved, and intraluminal thrombotic material was reduced after HES treatment. Initial in vivo platelet accumulation was rapid and reached similar levels in both groups. However, the platelet accumulation curves decreased more frequently following HES than in the control group. HES does not prevent platelet accumulation at trauma sites, but reduces the sizes of the thrombi formed and may enhance disaggregation/fibrinolysis.
A number of colloid solutions are used for blood and plasma replacement and for improving tissue perfusion. Of these, dextran, originally introduced for volume substitution, was later found to be an effective agent against postoperative pulmonary embolism.1 Dextran is therefore widely used today as an antithrombotic agent, and earlier studies by our group have confirmed that it is an effective antithrombotic agent following trauma of different degrees of severity in small rabbit arteries.2 3 Hydroxyethyl starch (HES) is another colloid whose effects on volume substitution and tissue perfusion are well-documented,45 and there is some evidence that it causes less bleeding6 7 and fewer allergic reactions than dextran.4 However, the influence of HES on thrombus formation in microvascular surgery has
not previously been investigated. The aim of this study was to examine the effects of HES in vivo on platelet function, thrombus formation, and patency rates, following severe trauma in small rabbit arteries. To facilitate comparison with earlier results, the same techniques were used as in studying dextran 70 and saline solutions.2
MATERIALS AND METHODS Rabbits, males and females weighing 2.5 to 5.5 kg, were kept on a standard pellet diet and given water ad libitum. They were anesthetized intravenously with sodium pentobarbital (Mebumal® 60 mg/ml, ACO Lake- 357
Department of Experimental Research and Department of Plastic and Reconstructive Surgery, Malmo General Hospital, Malmo, Sweden Reprint requests: Dr. Wieslander, Dept. of Plastic and Reconstructive Surgery, Malmo General Hospital, S-214, 01 Malmo, Sweden Accepted for publication lune 3, 1990 Copyright © 1990 by Thieme Medical Publishers, Inc., 381 Park Avenue South, New York, NY 10016. All rights reserved.
Downloaded by: University of Iowa Libraries. Copyrighted material.
FORMATION FOLLOWING ARTERIOTOMY/
JOURNAL OF RECONSTRUCTIVE MICROSURGERY/VOLUME 6, NUMBER 4
Arleriotomy 7 mm
^ *~;
1 mm
Figure 2. The exposed artery is positioned in a double vascular clamp and a 7-mm long arteriotomy made using a microscope and microscissors. Intimectomy is next performed at high magnification on the central 5-mm segment using a surgical blade, and the arteriotomy is closed with a running microsuture. Blood flow is then reestablished by removing the clamp. The same clamp is used in all cases.
transferred to another Fenwal bag. Platelet-poor plasma (PPP) was prepared by centrifuging for later use in washing labelled platelets. Twenty-five MBq (1 millicurie = 37 MBq) of the beta-emitter 32P (phosphorus) as orthophosphate in dilute hydrochloric acid were dissolved in modified Ringer's solution (ACO Lakemedel, Solna, Sweden), added to the PRP in the Fenwal bag, and the resulting mixture incubated for 2 hr at 37°C in a rocker. The solution was removed after centrifuging and 15 ml of PPP added. The contents of the plastic bag were homogenized and PPP removed after centrifuging. Washing in this manner was carried out three times. Finally, a small amount of PPP (3 to 4 ml) was added, and the solution of 32P-labelled platelets infused into the rabbit being studied. The stability in blood of 32P-labelled platelets labelled in this way was studied earlier and COMPUTER FEN RECORDER found to be satisfactory.8 RADIOACTIVITY. The prepared arteries were placed in grooves in chromium-plated brass holders at body activity signal Ringor s solution temperature, and thin-window beta-counting GeigerMuller (GM) tubes (Philips ZP 1410) were positioned just above them (see Fig. 1) to detect emitted radiation. Care was taken to remove any blood present. Throughout the measurements, repeated checks were made to insure that radioactive blood had not leaked from the prepared vessel segments or surrounding skin/wound edges into the field-of-view of the detectors. The sigSKIN /CARTILAGE FLAP nals from the GM tubes were monitored continuously on a pen recorder and stored for later analysis using Figure 1. The exposed central artery of the ear is sealers (Canberra Model 2072) on-line to a Luxor moistened, wrapped in plastic foil, and placed in a groove in ABC806 computer. Independent counting channels a chromium-plated brass holder maintained at 39.8°C. The permitted measurements to be made on both ears radioactivity originating from 32P-labelled platelets in the simultaneously. vessel segment studied is registered on a Geiger-Muller Two hours after reperfusion, the segments of vesdetector. The output signals are monitored on a pen re358 corder and stored for later analysis using a computer. sel containing the arteriotomy/intimectomy regions
Downloaded by: University of Iowa Libraries. Copyrighted material.
medel, Solna, Sweden), and anesthesia maintained by intermittent injections of the same compound. During the measurements, the animals were maintained at 39.8 ± 0.2°C by thermostatically-regulated heating pads. Other than the local application of lidocaine (Xylocaine®, 10 mg/ml, AB Astra, Sodertalje, Sweden) to release vascular spasm, neither anticoagulants nor vasodilating agents were used Ringer's solution at body temperature was used as a local irrigant. The central arteries (diameters 0.8 to 1.2 mm) of the ears of 14 rabbits were prepared (Fig. 1). Platelets labelled with 32P, as described below, were infused into the aorta via a catheter inserted into the arteria femoralis. HES solution was next infused over an interval of 20 to 30 min. About 1 hr later, longitudinal arteriotomy (7 mm) followed by intimectomy (5 mm) were performed under magnification using a microscope (Fig. 2). Conventional microsurgical instruments were used, arteriotomy being performed with microscissors and intimectomy with a surgical blade using a standardized technique. The arteriotomy was closed with a running 10-0 monofilament nylon suture (Dermalon 10-0, TE 70, Davis & Geek, American Cyanamid Co, Pearl River, NY, USA) and blood flow reestablished by removing the vascular clamp. PLATELET LABELLING. A plastic bag (Transfer Pack Unit, Fenwal Laboratories, Deerfield, IL, USA) was prepared with 20 ml sodium citrate solution. A plateletdonor rabbit was anesthetized by intravenous administration of sodium pentobarbital and 100 to 150 ml whole blood drawn via an intraaortically-positioned cannula. A plastic tube connected the cannula to the bag, which was gently shaken during sampling to facilitate the mixing of sodium citrate and blood. Platelets were isolated as platelet-rich plasma (PRP), using standard centrifugation techniques, and
OCTOBER 1990
HyDROXYETHYL STARCH/WIESLANDER, SALEMARK, DOUGAN
RECORDING OF ARTERIOTOMY/INTIMECTOMY PLATELET ACCUMULATION. Platelet labelling, detecting of radio-
activity, and the experimental set-up have been described above. The trauma model used has been shown to be both reproducible and capable of yielding qualitative and quantitative information about platelet accumulation in the immediate post-reperfusion time period. RED THROMBOTIC MATERIAL. Intramural red thrombotic material was qualified as - / + (clean or small amounts), + + (moderate amounts) or + + + (large amounts). The amounts of material were estimated using a microscope at high magnification. HEMATOCRIT VALUES. In Group B, hematocrit values were measured before infusion of Plasmasteril®, at 10 min postinfusion, and then at hourly intervals for a further two hours (Table 1).
RESULTS BLEEDING TIMES. The bleeding time in the control group was 5±] min (median and quartiles) and in the HES group 4±j min (median and quartiles). Statistical comparison using the Mann-Whitney U-test showed that the bleeding times in the untreated control and HES groups were completely indistinguishable. PATENCY (Fig. 3). In the control group, good patency was recorded in five and poor or no patency in seven vessels. All 12 vessels in the HES group were patent, but two (both in one rabbit) had reduced patency. Comparison of these values using Fisher's Exact Test shows that the groups are significantly different (p < 0.05). PLATELET ACCUMULATION. Qualitative Features of the Platelet Accumulation Curves. In the untreated control group, activities in all cases increased rapidly just after restoration of blood flow and later decreased in five, remained constant in five, and increased in two vessels. In general, the accumulations decreased in vessels with good patency and were constant in those with reduced patency. In the HES group, activities also increased rapidly after releasing the vascular clamps and later decreased in 11 of the 12 vessels studied. Activity in the twelfth case, one of the two vessels with reduced patency, remained constant.
Quantitative Platelet Accumulation (Fig. 4). After re-
leasing the vascular clamps, the median values of radioactivity in both groups increased rapidly to 500 to 600 percent of their preoperative values. The median value in the control group remained high while that in the HES group decreased during the 2 hr radiation
No of vessels
• Control £23 HES
10 8 6
Tablet. Rabbit
O-Sample
210 211 212 213 214
34 30 19 30 28
Mean
(100%) (100%) (100%) (100%) (100%)
Hematocrit Values 10 min 23 22 14 18 18
(68%) (74%) (74%) (60%) (64%)
68%
70 min 29 22 14 16 21
(85%) (74%) (74%) (53%) (75%) 75%
130 min 31 22 15 13 23
(91%) (74%) (79%) (43%) (82%)
73%
The results are listed above as absolute values and as percentages of initial values. The degree of hemodilution obtained using HES is thus 20 to 30 percent. HES infusions were started immediately afater taking the 0-samples.
4 2
Good
Poor/No
Figure 3. Patency. Treatment with HES increases patency rates, compared to the control group. "Good" denotes rapid refilling; "Poor," slow refilling; and "No," no refilling when patency is checked using the "empty/refill" test.
359
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were excised and the levels of residual background activity recorded. Relative values of platelet accumulation in the two groups, expressed as the ratios of the counting rates in the postoperative periods to the "steadystate" preoperative values in percent, at intervals of 15 min post-reperfusion, were derived and compared statistically using the Mann-Whitney U-test. GROUPS. Group A was an untreated control group of 12 vessels in eight rabbits. Group B comprised 12 vessels in six rabbits treated with 1 g hydroxyethyl starch MW 450,000 in 17 ml saline/kg b. w. (Plasmasteril®, 6 percent in NaCl, Fresenius AG, Bad Homburg, FRG) administered through an intraaorta catheter about 2 hr prior to vessel reperfusion. BLEEDING TIMES. The time intervals elapsing between restoring blood flow (release of vascular clamps) and cessation of bleeding from the arteriotomies were noted in minutes. PATENCY. Vessel patency was determined at the end of all experiments and recorded as either good (rapid refilling) or poor/not patent (slow or no refilling) using the empty/refill test, performed as follows. Using a pair of microforceps, the vessel was occluded just distal to the arteriotomy region and emptied of blood over a short distance using a second pair of microforceps. The first pair of microforceps was then opened and patency assessed using the above criteria for refilling.
JOURNAL OF RECONSTRUCTIVE MICROSURGERY/VOLUME 6, NUMBER 4 OCTOBER 1990 °
No of vessels
CONTROL
Control 10 o
HES
8
o
6 4 Median values Quartile ranges
—
2
|Arteriotomy
minutes
Figure 5. Thrombotic material. Decreased amounts of red thrombotic material were observed following HES infuFigure 4. \n vivo accumulation of 32P platelets at arteri- sion. The classification used is: - clean, + small amounts, + + otomy/intimectomy trauma sites. Median values and inter- moderate amounts, and + + + large amounts of material. quartile ranges of the post-reperfusion increases in platelet activity. The values are plotted as percentages of the steadystate values. Platelet accumulation at trauma sites was rapid and reached about the same level for both groups initially. The larger proportion of decreasing curves in the HES group is reflected by the gradual decrease in its median value. The groups are, however, statistically indistinguishable. intimectomy
360
detection period. There were no statistically significant ence of dextran 70 has been reported to be deficient, differences in platelet accumulation between the groups. with reduced tensile strength compared to untreated RED THROMBOTIC MATERIAL (Fig. 5). Red thromcontrol animals 1213 and it has been suggested that botic material was observed in all vessels. In Group A, HES has a similar fibroplastic effect.14 The potential the material visible at inspection was quantified as 3 usefulness of HES as an antithrombotic agent has, - / + , 5 + + and 4+ + + and in Group B as 9 - / + , 2 + + however, not been documented. and 1 + + + . Comparison of these values using the Our results show that, compared to the control Mann-Whitney U-test with an ordinal scale yields p < group, HES treatment does not increase vessel bleed0.05, so that the deposits are significantly smaller ing time, while in dextran-treated animals, the correwhen HES is present. sponding bleeding time is considerably prolonged.2 Dextran 70 and HES cause similar degrees of hemodilution, a colloid effect known to increase peripheral flow rates, which is possibly an important DISCUSSION factor in reducing thrombus deposition and improving patency. In our model, treatment with HES or dextran The particular trauma model used (arteriotomy/ 70 leads to considerable improvements in patency, intimectomy) was chosen because it simulates the consistent with the reduced amounts of thrombotic clinical microvascular trauma occurring, for example, material observed. That similar numbers of platelets in avulsion injuries, and to facilitate comparison with accumulate at reperfusion as rapidly as in untreated saline and dextran solutions previously tested using controls shows that platelets apparently function normally (adhere and aggregate) even in the presence of the same model. HES is fairly new and interesting colloid reported these substances. This suggests that the positive efto be a clinically safe plasma volume expander.7 Unlike fects the agents produce must occur at later stages of dextran, it does not introduce clinical bleeding diathe- thrombus deposition, e.g., enhanced fibrinolysis. The ses6-7 at doses of less than 1.5 g/kg b. w./day unless a effect must be specific to these colloids, since saline hemostatic defect, e.g., von Willebrand's disease, is solution (used as carrier of the colloids) does not 2 already present.9 Like dextran,10 it has been reported produce antithrombotic effects. to enhance fibrinolysis.1' The fibrin formed in the presTo summarize, our study shows for the first time
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Steady s
HYDROXYETHYL STARCH/WIESLANDER, SALEMARK, DOUGAN
that HES, without endangering hemostasis, produces antithrombotic effects and leads to improvements in patency and related parameters in severely traumatized small arteries.
6. 7. 8.
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This research was supported by grants from the Swedish Medical Research Council (B87-17X-00759-22B), the Medical Faculty, University of Lund, Lund, Sweden, and Syskonen Svensson's Foundation, Malmo, Sweden
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