Microvascular repair following a modified crush-avulsion injury in a rat model: effect of recombinant human tissue-type plasminogen activator on the patency rate

The failure rate of replantations following a crush-avulsion type injury is high. This study has been designed to reproduce an effective standardized crush-avulsion injury model to the femoral artery of the rat and evaluate the antithrombotic efficacy of systemic intravenous administration of recombinant human tissue-type plasminogen activator (rt-PA). The crush-avulsion injury was reproduced by using a bulldog clamp and two hemostats and followed by microvascular repair. The animals were divided into three groups of 20 rats each and received either normal saline, heparin 100 U/kg body weight, or rt-PA 3.5 mg/kg body weight intravenously. Patency tests were performed 20 min and 48 h after blood flow reestablishment. Results showed that this experimental crush-avulsion injury model ensures low patency in the control group, whereas systemic rt-PA administration improves the patency rate statistically significantly compared to control and heparin groups at both 20 min and 48 h postrevascularization.     

Microsurgical repair after crush-avulsion injury of the femoral vein in rats: prevention of microvascular thrombosis with recombinant human tissue-type plasminogen activator (rt-PA).

Vein thrombosis is often encountered in microsurgery, especially in the case of crush-avulsion injuries. The aim of this study was to investigate the effect of systemic administration of recombinant tissue-type plasminogen activator (rt-PA) on the patency of the femoral vein of the rat, which had previously sustained a crush-avulsion injury. The study consisted of 3 groups of male Wistar rats, 20 animals each. A standardized crush-avulsion injury model was used. After microvascular repair of the femoral vein, the animals received either normal saline (group A), heparin 100 U/kg body weight (group B), or rt-PA 3.5 mg/kg body weight (group C) systemically. Patency tests were performed at 20 minutes, 48 hours, and 1 week after blood flow reestablishment. According to our results, the patency rate of the rt-PA group was significantly higher than in both the control and heparin groups.     

Microvascular repair following crush-avulsion type injury with vein grafts: effect of direct inhibitors of thrombin on patency rate

The aim of this study was to develop a standardized effective thrombogenic arterial anastomosis model, as usually encountered in clinical practice, and to offer a detailed evaluation of the antithrombotic effect of thrombin's direct inhibitors, antithrombin III and hirudin, as locally applied. Wistar rats were divided into four groups of 12 animals each. The carotid artery sustained a standardized crush-avulsion-type injury (groups B-D). A segment of the afflicted area was removed and replaced by a microvenous graft. Group A had no crush-avulsion injury inflicted; a microvenous graft replaced a simple resection from the center of the carotid artery. During microvascular anastomoses, normal saline (groups A and B), recombinant hirudin (group C), or antithrombin III (group D) were locally applied. Bleeding times were recorded, and patency tests were performed 20 min, 48 h, and 1 week after blood flow reestablishment. All grafts were harvested and examined histologically. Patency tests, 1 week postrevascularization, demonstrated that this experimental crush-avulsion injury model ensured low patency in group B (25%), whereas group A, which had no injury inflicted, achieved a 100% patency rate. The local application of hirudin and antithrombin III significantly increased bleeding times as well as the patency rate (92% and 75%, respectively) compared to group B. These findings indicate the efficiency of the experimental model and the potential use of thrombin's direct inhibitors in microvascular surgery.     

Assessment of the microsurgical skills: 30 minutes versus 2 weeks patency

The aim of this study is to evaluate the amount of training needed by a trainee, with no background in microsurgery, in order to achieve proper skills for microvascular anastomosis. A protocol based on the rat femoral artery was established to provide a quantitative representation. Five inexperienced subjects started performing microvascular anastomosis. Patency was assessed at 30 min. The final assessment was performed at 2 weeks when rats were reoperated and the patency below the anastomosis was checked. The experiment was discontinued for one subject when he/she succeeded to have two series of four anastomosis with 100% patency at 2 weeks. The results were: 47.5% patency rate at 30 min and 7.5% at 2 weeks (series 1-2); 67.5 and 32.5% (3-4); 82.5 and 35% (5-6); 100 and 70% (7-8); 100 and 87.5% (9-10). Two trainees obtained 100% patency at 2 weeks after series 9-10. Other three needed two more series. There is a significant statistic difference (P < 0.01) between the results at 30 min and 2 weeks for the series (1-2, 3-4, 5-6, 7-8). The patency rate at 2 weeks reflects in a better way the microsurgical skills of a trainee. For long term functioning anastomosis, the training period needs an extension beyond that necessary for 100% patency at 30 min.     

Microvascular grafts: effect of diameter discrepancy on patency rates

This study examined the effects of diameter discrepancy between interpositional vein grafts within a range of 0.25 to 2 mm. One hundred rats underwent isolation of a 10 mm segment of the femoral artery, from which a 4 mm segment was removed. Venous grafts measuring 8 +/- 2 mm were interposed. The rats were categorized by their graft to artery diameter ratios. Patency was assessed upon completion and 4-6 days postoperatively. All surgical specimens were submitted to the pathology department for microscopic evaluation. Results showed Group I: graft to artery ratio 1:1, patency rate 90% (18/20); Group II: ratio 0.75:1, patency rate 80% (16/20); Group III: ratio 0.5:1, patency rate 60% (12/20), Group IV: ratio 0.25:1, patency rate 20% (4/20); Group V: ratio 2:1, patency rate 60% (12/20). It is concluded that 1:1 venous graft to artery ratios are optimal. When this is impossible, diameter discrepancies should be at least 0.75:1 for acceptable patency rates to be attained in microvascular grafting.     

Comparison of single-dose antithrombotic agents in the prevention of microvascular thrombosis.

A model of thrombosis was used to evaluate the efficacy of single-dose heparin, hemodilution, and 40,000 milliwatts dextran in the prevention of microvascular thrombosis. Forty Lewis rats (275 gm body weight) were divided into four groups (n = 10): hemodilution (6 ml NS), single-dose heparin (1 mu/gm), 40,000 mw dextran (0.3 gm/100 gm), and control (0.275 ml NS). After exposure of the superficial femoral arteries, a model of arterial crush with arteriotomy followed by an intimal abrasion was used. The animals randomly received one of the four solutions above. Experimental results included patency rates of 90% at 20 minutes and 10% at 24 hours in the hemodilution group, 100% at 20 minutes, and 70% at 24 hours in the single-dose heparin group, and 100% at both 20 minutes and 24 hours in the dextran group. A 100% thrombosis rate was observed in the control group at 20 minutes and 24 hours. Single-dose heparin and dextran significantly improved patency in comparison to both the control and hemodilution groups at 24 hours (p less than 0.01). Scanning electron microscopy of the vessels revealed thrombus deposition consistent with these findings. These results indicate that single-dose heparin and single-dose dextran reduce thrombosis in this model of microvascular injury.     

Effects of early pre- and postoperative irradiation on the healing of microvascular anastomoses

A comparative study was undertaken to evaluate the possible effects of time selection of irradiation on the healing of microvascular anastomoses. In this experimental study, 40 adult male Sprague-Dawley rats were divided into four groups, 10 animals in each. In Group 1, the rats were exposed to radiotherapy with a dose of 20 Gy in a single fraction at a 170,916 cby/min dose rate, at an 80-cm source to a skin distance from 3 to 4 cm in the right groin fields involving the femoral artery, using a Co60 megavoltage radiotherapy machine. The animals were operated on under general anesthesia 2 weeks later. During the operation, femoral microarterial anastomoses were performed. In Group 2, the rats were irradiated and operated on as in Group 1, on the same day. In Group 3, the rats were operated on and, 2 weeks later, irradiated, as in Group 1. In Group 4 (control group), the rats were operated on as in Group 1. No radiotherapy was performed. All rats were explored under general anesthesia after 1 month. Patency was evaluated, and the anastomotic sites were taken for histopathologic and electron microscopy studies. The authors found that there were no major differences in the healing of microvascular anastomoses in the early preoperative, immediate preoperative, and early postoperative irradiated groups. The patency rates of all radiotherapy groups were not statistically different from control arteries (p > 0.05).     

Effect of the two-wall-stitch mistake upon patency of rat femoral vein anastomosis: Preliminary observations

Anastomotic patency is believed to be the most important factor in microvascular surgery. The two-wall stitch is a technical error commonly considered to cause thrombosis of the anastomosis, especially on the venous side. In order to demonstrate the real effect on vein patency of the two-wall stitch, the authors performed a standardized mistake after correct microanastomosis on the femoral vein of 15 rats, with one stitch passing through the whole thickness of the two walls of the vein. Traditional correct anastomoses on the contralateral side were used as controls. Patency was assessed at 5, 20, and 60 min and at 24 h by the milking test, and by direct section of the vessel at 24 h. The results showed no statistically significant difference between the two techniques. Histological examination confirmed the clinical judgment about the vessel's patency, and ultrastructural microscopy evidenced only mild signs of endothelial activation. In conclusion, this study indicates that the occasional two-wall stitch does not necessarily increase the risk of venous occlusion in anastomoses of the rat femoral vein.     

Antithrombotic efficacy of direct oral anticoagulants on patency rate following microsurgical anastomosis in crushed rat arteries

BackgroundDuring the last decade direct oral anticoagulants (DOAC) have been established in various fields of medicine.Their use in microsurgery has not been evaluated yet though. This study aims to evaluate their efficacy in microsurgery and additionally compare them with a well established antithrombotic agent.Materials and methodsThe right femoral artery of 101 rats divided into 4 groups, was crushed and anastomosed. Group A (20 rats) received placebo therapy (1 ml NaCl 0.9%, orally), while Group B (27 rats), Group C (27 rats) and Group D (27 rats) received rivaroxaban (3 mg/kg, orally), dabigatran (30 mg/kg, orally) and enoxaparin (30 mg/kg, subcutaneously) respectively. All drugs were administered 3 h preoperatively and once daily for the following postoperative days until the sacrifice of the animals. Patency was evaluated at 1st, 7th and 20th postoperative day. Following patency evaluation the rats were sacrificed and the vessels were harvested for histological examination.ResultsNone of the rats died postoperatively. Patency rates of rivaroxaban group (78%), dabigatran group (70%) and enoxaparin group (63%) were statistically similar, but significantly higher than the placebo-treated control group (p < 0.05). Cells with morphologic features of endothelial cells were evident 7 days after the injury.ConclusionThe results of this study demonstrate the following: (1) rivaroxaban and dabigatran through inhibition of thrombus formation significantly enhanced the patency rate compared to placebo treatment (2) the antithrombotic efficacy of rivaroxaban and dabigatran in compromised microvessels was similar to that of enoxaparin, the most widely used antithrombotic agent.     

Microvascular repair with 1-mm polytetrafluoroethylene (PTFE) grafts: effect of recombinant tissue-type plasminogen activator (rt-PA) on the patency rate and healing process

The present study assesses the effect of recombinant tissue-type plasminogen activator (rt-PA) on the patency rate and healing process of microvascular polytetrafluoroethylene (PTFE) grafts. Wistar rats were used, divided into four groups of 25 animals each. After dissection of the carotid artery a segment of the vessel, 1 cm long, was resected and replaced by equal length graft. Two different type fibril length (30- or 60-microm) grafts of the same wall thickness (0.18 mm) were used. Normal saline or 3 mg/kg of body weight of rt-PA was applied locally in each group of different fibril length grafts. Patency tests were performed at 15 min and 4 weeks after blood flow was reestablished. All grafts were harvested and examined histologically. The results showed that local application of rt-PA improves patency statistically significantly in both types of fibril length grafts. Patency in 60-microm fibril length grafts was statistically significantly higher than that of 30-microm fibril length grafts, whether rt-PA was used or not. The use of rt-PA had no influence on the healing process of either type of graft.     

Local heparin is superior to systemic heparin in preventing arterial thrombosis

Thrombosis poses a significant problem in microvascular surgery, despite antithrombotic therapy. The purpose of the present study was to investigate whether a topical application of unfractionated heparin is equally efficient as a systemic bolus in avoiding thrombosis. A rat femoral artery model was used. Three different doses of systemic heparin (50, 100, and 200 U/kg) and one dose of locally administered heparin (100 U/ml) were evaluated and compared to a control group receiving isotonic saline. A thrombogenic injury, simulating poor microsurgical technique, was applied to the artery. The thrombus area was visualized by transillumination, and recorded for 60 min on video, for subsequent measurement. In addition, the level of activated partial thromboplastin time (APTT) and of anti-activated clotting factor X (aXa) was determined. Local heparin significantly reduced thrombus size as compared to isotonic saline and systemic heparin (50 and 100 U/kg). High-dose systemic heparin (200 U/kg) was equally potent, but local heparin had significantly less influence on the hemostatic parameters.     

乌司他丁对肝移植术患者凝血功能的影响

目的 评价乌司他丁对肝移植术患者凝血功能的影响.方法 拟行原位肝移植术的终末期肝病患者20例,年龄36～59岁,ASA Ⅲ或Ⅳ级,随机分为2组:对照组(C组)和乌司他丁组(U组).U组切皮后静脉输注乌司他丁2万U/min,持续1 h,乌司他丁输注量120万U(加入400 ml生理盐水中),每隔4 h重复输注;C组给予等容量生理盐水.分别于麻醉后切皮前、无肝前期30 min、无肝期30 min、新肝期30 min和术毕时采集中心静脉血样,测定Sonoclot凝血功能指标、常规凝血功能指标及血浆Ca2+浓度,记录术中出血量、输血量及术后24 h引流量.结果 与麻醉后切皮前比较,两组凝血酶时间、凝血酶原时间、活化部分凝血活酶时间、激活凝固时间延长,纤维蛋白原浓度、Ca2+浓度、纤维蛋白凝集率和血小板功能降低,C组D-二聚体浓度升高(P<0.05);与C组比较,U组D-二聚体浓度降低,血小板功能增强,术后24 h引流量降低(P<0.05),其余指标差异无统计学意义(P>0.05).结论 术中静脉输注乌司他丁可改善肝移植术患者凝血功能,减少术后出血.     

Prevention of hypocalcemia by administration of homologous plasma during experimental hemorrhagic pancreatitis in the pig

Pancreatitis was induced in fifteen immature pigs while five additional pigs underwent sham laparotomy. Animals with pancreatitis were separated into three groups of five each with respect to fluids administered after pancreatitis was induced. Each pig in group A and the control group received normal saline, 300 ml/hour for 8 hours. Pigs in group B received pig plasma at 150 ml/hour plus normal saline at 150 ml/hour for 8 hours. Pigs in group C received pig plasma at 50 ml/hour plus normal saline at 50 ml/hour for 24 hours. Values for serum hematocrit, total protein, and total calcium were measured preoperatively and at 4,8, and 24 hours postoperatively. Control animals and group B animals experienced no change in any parameter. Group A animals sustained transient severe hemoconcentration, permanent severe hypoproteinemia, and hypocalcemia. Group C animals displayed a transient moderate hemoconcentration and a moderate but sustained decrease in calcium concentration.     

去细胞异体神经和碱性成纤维细胞生长因子修复神经缺损

目的　研究碱性成纤维细胞生长因子 (bFGF)对去细胞异体神经修复神经缺损的促进作用。　方法　根据bFGF因子用药浓度分为 10 0 0、5 0 0、2 5 0和 10 0U/ml与盐水对照组每组 5只日本大耳白兔。采用抗神经微丝免疫组化染色观察各组神经纤维再生的距离 ,抗S 10 0免疫组化染色观察许旺细胞的分布。研究不同浓度和剂量的外源性bFGF对去细胞支架移植后早期神经再生的影响。　结果　高浓度bFGF组 (10 0 0U/ml和 5 0 0U/ml)的神经再生距离在术后 10d明显大于盐水对照组 ,(P <0 0 1)。而低浓度bFGF组 (2 5 0U/ml和 10 0U/ml)与盐水对照组无明显差别 (P >0 0 5 )。　结论　去细胞异体神经移植后早期应用一定浓度和剂量的bFGF能明显提高再生轴突在支架内的生长速度。     

PEG-hirudin/iloprost coating of small diameter ePTFE grafts effectively prevents pseudointima and intimal hyperplasia development.

OBJECTIVES: Small diameter PTFE grafts are prone to thrombosis and intimal hyperplasia development. Heparin graft coating has beneficial effects but also potential drawbacks. The purpose of this study was to evaluate the experimental efficacy of PEG-hirudin/iloprost coated small caliber PTFE grafts. METHODS: Thirty-six femoro-popliteal ePTFE grafts (expanded polytetrafluoroethylene, diameter 4 mm) were inserted into 18 pigs. Grafts were randomised individually for each leg and grouped for 3 groups. Group I consisted of native ePTFE grafts, group II were grafts coated with a polylactide polymer (PLA) without drugs and group III grafts were coated with PLA containing a polyethylene glycol (PEG)-hirudin/iloprost combination. The follow-up period was 6 weeks. Patency rates were calculated and development of pseudointima inside the grafts was noted. Thickness of intimal hyperplasia at the distal anastomoses was measured using light microscopy. RESULTS: Patency rates for group I were 6/9 (67%), for group II 9/10 (90%) and 12/12 (100%) for group III. In groups I and II there was a significant reduction of blood flow proximal to the graft at graft harvest, to 29+/-12 and 28+/-20 ml/min respectively (both p<0.01 versus preoperative value), whilst in group III blood flow, 99+/-21 ml/min, remained at the preoperative level. Subtotal stenosis due to development of pseudointima was noted in each of the native and PLA coated grafts but not in group III grafts. Intimal hyperplasia at the distal anastomosis was lowest in group III. CONCLUSIONS: The PEG-hirudin/iloprost coating of ePTFE prostheses effectively reduced pseudointima and intimal hyperplasia development and led to superior graft patency.     

Improved microvessel repair: laser welding with an anti-thrombotic solder

BACKGROUND AND OBJECTIVES: Concentrated protein solutions can be used as thermally polymerized solders in laser welding. Solders supplemented with biologically active chemicals may provide in situ drug delivery for localized therapeutics. These studies characterize a serum albumin (SA) solder containing heparin, designed to reduce microvascular thrombosis rates. STUDY DESIGN/MATERIALS AND METHODS: Samples of heparin added to 30% SA to obtain heparin-to-albumin molar ratios (HAMR) of 4:1 and 2:1 were thermally polymerized, and heparin release into saline was measured. Using a rat thrombosis model, patency was determined for suture, and 0 U/ml (control), 2.5 U/ml, 50 U/ml heparin solder repairs. RESULTS: Heparin release was five times higher for 4:1 than 2:1 HAMR solder acutely, but was equivalent after 2 days. Animal patency rates were: 50% suture, 0% control, 50% low heparin, 66% high heparin (P < 0.05 vs. control). CONCLUSIONS: Solders incorporating heparin should provide in situ anti-thrombotic therapy reducing the risk of microvascular thromboses.     

The effects of intraoperative infusion of dexmedetomidine on early renal function after percutaneous nephrolithotomy

Background: Percutaneous nephrolithotomy (PCNL) may interfere with renal function because of continuous fluid irrigation and compression. The aim of this study was to evaluate the effects of an intraoperative infusion of dexmedetomidine on renal function in patients undergoing PCNL. Methods: This study included 40 patients between the ages of 18 and 65 years who underwent PCNL. After induction of anesthesia, we administered 100 ml of normal saline to the patients in the control group (n=20) and 1 mcg/kg dexmedetomidine in 100 ml of normal saline to the patients in the dexmedetomidine group (n=20) over 10 min. Throughout the surgery, 1 mcg/kg/h dexmedetomidine and 1 ml/kg/h normal saline infusions were given to the dexmedetomidine and control groups, respectively. Renal function, electrolytes, serum levels of neutrophil gelatinase‐associated lipocalin (NGAL) and cystatin C were analyzed after induction and post‐operatively at 2, 8 and 24 h. Renin levels and blood gas analyses were performed after induction and before extubation. Results: No statistically significant differences were found between the two groups with regard to renal function, creatinine clearance (CrCl), NGAL, cystatin C and serum electrolyte levels at 0, 2, 8 and 24 h post‐operatively. End‐surgery renin levels in the dexmedetomidine group were significantly lower than the baseline levels in the control group. Conclusion: In PCNL, an intraoperative infusion of dexmedetomidine was not found to have beneficial effects on CrCl, NGAL or cystatin C levels early after the procedure; however, it reduced renin levels.     

肺灌洗及外源性肺表面活性物质治疗烟雾吸入性损伤的实验研究

为探讨肺灌洗与外源性肺表面活性物质（ＰＳ）治疗对严重烟雾吸入伤后内源性ＰＳ功能障碍和急性呼吸衰竭的防治效果,作者将重度烟雾吸入伤后Ｗｉｓｔａｒ大鼠经气管插管注入含ＰＳ的等渗盐水或等量盐水行肺灌洗,机械通气４小时,观察２４小时,检测动脉血气、肺水量、静态肺顺应性（Ｃｓｔ）、支气管肺泡灌洗液（ＢＡＬＦ）蛋白含量、ＢＡＬＦ表面张力特性和２４小时病死率等。结果显示：动物伤后立即出现严重缺氧和一氧化碳中毒。烟雾吸入组发生急性呼吸衰竭、高通透性肺水肿和ＰＳ功能障碍。烟雾吸入加灌洗加ＰＳ加机械通气组Ｃｓｔ和ＢＡＬＦ表面张力特性明显改善,氧合能力显著增强,肺水量和ＢＡＬＦ蛋白含量降低,２４小时病死率明显下降。烟雾吸入加灌洗加机械通气组也有一定疗效。作者认为,早期肺灌洗和外源性ＰＳ治疗能有效恢复烟雾吸入所致内源性ＰＳ功能抑制,改善肺功能,防止高通透性肺水肿和呼吸衰竭,降低早期病死率。     

Adverse effects of topical prostacyclin application in microvascular surgery: an experimental study

The efficacy of topical prostacyclin as an antithrombotic agent was tested in a model of microvascular trauma. Preliminary measurements were made to determine a suitable dosage. Twenty-seven central arteries of rabbit ears were then prepared and 32P-labelled platelets infused intraaortically. Arteriotomy (7 mm) was followed by intimectomy (5 mm). Fifteen vessels in a control group were irrigated with Ringer's lactate and 12 vessels in an experimental group were treated with prostacyclin (10 ng/ml) in normal saline. Bleeding times at the sites of arteriotomy intimectomy, in vivo accumulations of isotope-labelled platelets, amounts of red thrombotic material, and patency were recorded. Patency was lower following prostacyclin treatment (1/12 as against 5/15) but not significantly so, and there were no statistically significant differences in other parameters. Prostacyclin treatment decreased vessel wall tone, interfering with blood-flow and promoting thrombus formation.     

The management of bleeding following transurethral prostatic resection by local irrigation with a thrombin solution

The clinical value of local irrigation of thrombin was studied in 30 patients with urethral bleeding following transurethral prostatic resection. A thrombin solution consisting of 25,000 units of thrombin in 500 ml of saline was irrigated in 20 patients through the especially designed three-way balloon catheter at a speed of 8.3 ml/min for five hours after the operation while sterile saline was irrigated in the same manner in 10 patients as a control. The hemostatic effect of thrombin solution was evaluated as excellent in 2 patients (10%), good in 5 (25%), moderate in 6 (30%), and poor in 2 (10%) with 5 drop outs (25%) in the thrombin irrigating group. In the control group, the effect of irrigation was evaluated as good in 1 patient (10%), moderate in 4 (40%), poor in 2 (20%), and unable to be estimated in 2 (20%) with 1 drop out (10%). Therefore the rate of overall effectiveness was 65% in the thrombin irrigating group and 50% in the control group and volume of postoperative blood loss was 0.58 ml/g X hour and 0.62 ml/g X hour, respectively. Though almost all of the patients in both groups showed a moderate elevation of fibrinogen in serum, there was no adverse effect considered to be caused by thrombin irrigation. The local irrigation of the thrombin solution is a useful method and safe in the management of bleeding following transurethral prostatic resection.