一种新的腹腔粘连动物模型及其关联指标的定量分析
作者：郑青山 桂常青 孙瑞元 王民
单位：郑青山(中国科学技术大学生命科学学院，合肥 230027)；桂常青 孙瑞元(皖南医学院药物研究所，芜湖 241001)；王民(山西皮尔复临床医药研究所，太原 230006)
关键词：动物模型；粘连；联合药物治疗；ROC曲线；尿囊素；甲硝唑；地塞米松
中国临床药理学与治疗学000202 目的 制作一种新的大鼠腹腔粘连模型，并确定腹腔渗液中组织纤溶酶原激活物活性(PAA)是否具有判断粘连形成的能力。方法 大鼠随机分为2组，麻醉后不作任何消毒，打开腹腔，找到回盲部。距盲端约1～2 cm处结扎，剪去盲端的多余部分，挤去敞口盲端内的肠内容物，不作消毒，放回腹腔。在腹腔内放置直径约4 mm的引流软管。作两层缝合。皮肤层缝合前，用药组注入抗粘连组方(AMD)(尿囊素+甲硝唑+地塞米松)，剂量为1.5 ml.100 g-1体重。对照组注射等容量5 % GS。6 h后抽取腹腔液1 ml，拨除引流管。1 w后脱臼处死大鼠，打开腹腔检查粘连的形成及其程度。用相关操作特征图 (ROC) 分析PAA是否具有判断粘连形成的能力。结果 对照组的20只大鼠均形成腹腔粘连，腹腔渗液量为(1.25±0.09) ml,WBC(×103) =(23.1±6.6) mm3，PAA=(0.9±0.4) IU.ml-1。用药组没有形成腹腔粘连(0/20)，腹腔渗液量为(0.52±0.04) ml (P<0.01),WBC (×103) 为(10.6±4.2) mm3 (P<0.01), PAA 为(23.1±6.6) IU.ml-1(P<0.01)。由于此3药联用的抗粘连作用与腹腔炎性渗液量减少，抗菌抗炎作用，以及提高渗液中PAA有关，从而反映了本模型的形成机理。ROC分析认为， PAA可作为判断粘连是否形成的重要指标，当PPA>1.24 IU.ml-1时，粘连不易形成，反之则较易形成。结论 本动物模型是研究大鼠腹腔粘连的有效工具，腹腔渗液中PAA活性的变化对大鼠腹腔粘连的形成具有判断能力。
, 百拇医药
中图分类号 R965.1
A novel animal model of intra-abdominal adhesion and
quantitative evaluation with related indices
ZHENG Qing-Shan
(School of Life Science，University of Science and Technology of China, Hefei 230027)
GUI Chang-Qing
(Institute of Materia Medica, Wannan Medical College, Wuhu 241001)
, http://www.100md.com
SUN Rui-Yuan
(Institute of Materia Medica, Wannan Medical College, Wuhu 241001)
WANG Ming
(Pierfu Institute of Clinical Medical Research, Taiyuan 030006)
Aim To set up a novel animal model of intra-abdominal adhesion and to determine whether the tissue plasminogen activator activity (PAA) in exudate can be taken as an indicator to judge the formation of the adhesion. Methods Rats were randomly divided into 2 groups. Each animal in both groups was opened the abdominal cavity via midline laparotomy without any disinfectant measures. 2-cm section from the cecal end was clamped and ligated, 1-cm cecum of the section was cut, and another 1-cm end from the ligated site was kept. After the content in the end was extruded, the cecum was put back without using any antibacterial agent. Before the skin closure, an effective combination AMD (allantoin, metronidazole and dexamethasone in combination), was given (ip) according to 1.5 ml per 100 g body weight (60.6 mg.kg-1). The control group was injected (ip) the same volume of normal saline. After 6 h, the exudate was extracted from drainage-tube, with the rats varying posture, and after 1 kw, the rats were killed for examining the intra-abdominal adhesion. The values of PAA of exudate in both groups were analyzed by the relative operating characteristic curve (ROC). Results In the control group, all 20 rats formed the adhesions, the amount of exudate=(1.25±0.09) ml, the number of WBC(×103)=(23.1±6.6) mm3 and PAA=(0.9±0.4) IU.ml-1 in the exudate of abdominal cavity. In AMD group, however, there was not the adhesion formations (0/20), the amount of exuade was (0.52±0.04) ml (P<0.01), the number of WBC (×103) (10.6±4.2) mm3 (P<0.01), and PAA (23.1±6.6) IU.ml-1(P<0.01) in exuade.ROC analysis indicated that if PAA >1.24 IU.ml-1 in the exuade, the adhesion was difficult to form, and vice versa. Conclusion This animal model can be taken as an effective tool to evaluate the human adhesion related to multi-links on the pathogenesis, and the PAA in exudate an indicator to judge intra-abdominal adhesion formation.
, http://www.100md.com
Key words animal model; adhesion; combination drug therapy; ROC curve; allantoin; metronidazole; dexamethasone
The etiology of intra-abdominal adhesion formation involves many factors, such as tissue ischemia, infection, inflammation and exudation, and the mechanisms have been attributed to peritoneal plasminogen activator activity (PAA) decreased in peritoneum over the past 3 decades[1,2]. On therapeutics and prevention, however, there has been just a little advance because numerous agents and surgical techniques often obtain conflicting results. The cause may be mainly, at least partly, multi-factor in the adhesion etiology and multi-pathway in the adhesion mechanism, which makes the adhesions become difficult to prevent by using a single drug or certain a measure. In this study, we firstly made the animal model of intra-abdominal adhesion caused by multi-factor,then an effective combination (AMD)[3], allantoin (Alt), metronidazole (Met) and dexamethasone sodium phosphate (Dex), of preventing the adhesion formation was used to evaluated the model and to explore its pathway. Lastly, the relationship between the PAA in exudate of abdominal cavity and the adhesion would be assessed to determine whether the PAA could be taken as an indicator to judge the formation of intra-abdominal adhesion.
, 百拇医药
1 Experiment
1.1 Materials Alt powder was obtained from Jiangsu Huanghai Pharmaceutical Factory and the purity=99.6 %,Met powder from Tianjing Hebei Pharmaceutical Factory and the purity=99.85 %, and Dex powder from Roussel Uclaf Co and the purity=99.6 %.The compatibility AMD(the patent register code: 97110167.1), provided by Prof. Wang M, was composed of Alt,Met and Solution was prepared with 5 % GS. The assay kit of tissue plasminogen activator (PPA) was purchased from FJ Sun Biotech Co, №990406.
, http://www.100md.com
Wistar rats(weighing 190～210 g) of both sex were purchased from the Animal Center of Shanghai Medical University (Grade Ⅱ, Certificate №02-22-12).
1.2 Animal models of intra-abdominal adhesions Rats were randomly divided into 2 groups of 10 rats. Anesthetized with 3 % sodium pentobarbital (1 ml.kg-1, ip), each animal in both groups was opened the abdominal cavity via midline laparotomy without any disinfectant measures. 2-cm section from the cecal end was clamped and ligated, 1-cm cecum of the section was cut, and another 1-cm end from the ligated site was kept. After the content in the end was extruded, the cecum was put back without using any antibacterial agent. Then the end of operation knife was used to grind the parietal peritoneum on the anterior abdominal wall for 5 times. A soft drainage-tube, 4 mm in diameter, was put. The wound closure was accomplished using 3-0 silk for the abdominal medline and 4-0 silk for the skin. Before the skin closure, AMD was given (ip) according to 1.5 ml per 100 g body weight (60.6 mg.kg-1). The control group was injected (ip) the same volume of normal saline. Then the drainage-tube was clamped, fixed and covered. After 6 h, the exudate was extracted from drainage-tube, with the rats varying posture. If the volume did not reach to 1 ml was obtained, a small amount of normal saline was injected via the tube to wash the abdominal cavity, and the washing liquid kept for 3～5 min was collected until 1 ml. The sample contained a lot of blood would be discarded. The collected exudate was put into anticoagulated silicic tube and centrifuged at 1500×g for 10 min. 200 μl supernatant mixed with the same volume of a kind of acidic liquid was kept at 2～8 ℃ for use. Then the drainage- tube was taken out. After 1 wk, the rats were killed for examining the intra- abdominal adhesion. The adhesions were scored by a modified scale devised by Swolin [4]: 0 (no adhesions),1 (filmy connections separated spontaneously),2 (filmy adhesions separated by gravity),3 (filmy adhesions by traction), and 4 (Dense adhesions requiring sharp dissection).
, 百拇医药
1.3 Determination of indices 1) PAA: it was determined by the chromogenic substance method. The procedure of the experiment was done according to the PPA assay kit introduction. Exudate absorbance (A) in 96-well plate was determined at 405 nm with a microplate photometer (Yutai Yanghang Co, Hong Kong). If the exudate was diluted,A must multiply the multiple of the dilution; 2) the exudate volume in abdominal cavity; 3) the number of WBC in the exudate.
1.4 Data analysis A cutoff of PAA between the AMD group and the control group was the maximum sum of the sensitivity and the specificity of PAA.It denoted that the rats with PAA >the cutoff were not easily to form intra-abdominal adhesions, and vice versa. The experimental performance was determined by the relative operating characteristic curve (ROC)[5,6]plot,on which x-coordinate was 1 minus specificity and y-coor- dinate was sensitivity. The area under ROC and its standard error were expressed as W±SEw, and analyzed by z-test.
, 百拇医药
2 Results
2.1 Preventive effect of AMD on intra-ab- dominal adhesions in rats The 20 control rats were formed strip or round adhesions of intestinal canal (Fig 1), each score > 1 and the mean score was 2.4±0.6. But there were not found obvious adhesions in AMD group, each score<2 and the mean score was 0.3±0.4 (P<0.01vs control).
Fig 1 Formation of the rat model of intra-abdominal adhesion
, 百拇医药
Left:round intestinal canal; Right:strip intestinal canal
2.2 Influence of AMD to some indices related intra-abdominal adhesions Compared with the control group,the PAA increased (P<0.01), the exudation volume and the number of WBC decreased (P<0.01) (Tab 1). It indicated that AMD could decrease fibrin into abdominal cavity and increase fibrinogenolysis.
Tab 1 Preventive effect of AMD(Alt+Met+Dex) on intra-abdominal adhesion in rats (
±s,n =19) Groups
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Exudate
PAA
WBC(×103)
AMD
0.52±0.11
1.6±0.8
10.6±4.2
Saline
1.25±0.09*
0.9±0.4*
23.1±6.6*
, http://www.100md.com
*P<0.01 by t test.PAA=the tissue plasminogen activitor activity
2.3 Cutoff of PAA between adhesions and no adhesions, and analysis of experimental performance According to the order from small to large, each value of PAA was selected as a cutoff, respectively, to form a series of four-cells like Tab 2, and all the sum of the sensitivity and the specificity in each point were calculated. The maximum sum was the real cutoff (C point on Fig 2), at which sensitivity + specificity = 0.84+0.74 =1.58, and the cutoff = 1.24 IU.ml-1. χ2=10.19, which was also the largest value in all four-cell tables. It indicated that the adhesion was not easily formed when PAA was over 1.241 IU.ml-1.
, 百拇医药
Fig 2 Each value of the tissue plasminogen activator activity (PAA) corresponded to sums of sensitivity and specificity.
Point C corresponding to the maximum sum (1.58) in y-axis and PAA ( 1.241 IU.ml-1) in x-axis.
Tab 2 Four-cell table Cutoff/IU.ml-1
Adhesion
No adhesion
, 百拇医药
<1.214
16(a)
5(b)
≥1.241
3(c)
14(d)
χ2 =10.19,P<0.01；Sensitivity=a/(a+c)=16/(16+3)=0.84；
Specificity=d/(b+d)=14/(14+5)=0.74
The results of the experimental performance (Fig 3):W=0.764, SEw=0.0837, z=3.16, P<0.01. It meant that there was a significant difference between the PAA in adhesion animals and that in no adhesion animals, indicating the adhesions were distinguished by the PAA (< cutoff= 1.241 IU.ml-1) of exudate in abdominal cavity.
, 百拇医药
Fig 3 Analysis of ROC curve plot to the ability of the tissue plasminogen activator activity (PAA) in judgement of the intra-abdominal adhesion formation.
3 Discussion
Intra-abdominal adhesion is a common irreversible syndrome, so the goal of new drug development is how to prevent it. The PAA decreased in peritoneum has been taken as main mechanism of adhesion formation by many investigators[1,7,8]. The availability of recombination tissue plasminogen activator (rtPA) has also been conformed[9].If an investigator take a piece of peritoneum from no general peritonitis to determine the PAA, the location of sample possibly affects the result. In addition, it is a kind of traumatic technique and difficult to apply in clinical study. So the determination of PAA in exudate has some advantages for use. However,the relevance between PAA in peritoneum and in exudate is unclear, so it is necessary for quantitatively evaluate the relationship between PAA and the adhesion with ROC analysis. The analysis has been taken as the most important method for judging the experimental performance in recent decades, especially for determining a cutoff. The PAA in abdominal cavity can be taken as a valid indicator in judgement of the adhesion formation in this study.
, 百拇医药
The process of making animal model in this study reflected the complexity of etiology, which involved ischemia, mechanical stimulation,trauma,exudation,infection,and inflammation. It may reflect the nature of
intra- abdominal adhesion much effectively.
The compound drug, AMD, can prohibit the fibrin rich exudate into the abdominal cavity and at the same time increase fibrinogenolysis by some positive approaches, such as anti-inflammation, anti-bacteria,anti-exudation, and increasing PAA[3].These factors may show some pathways of the animal model formation.
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Project supported by the National Natural Science Foundation of China, № 39670845 and the Natural Science Foundation of Anhui Province, № 98454735.
Correspondence to Dr ZHENG Qing-Shan.
Phn: 86-551-360-3754. Fax:86-551-360-3754.
E-mail: zhengqs@mail.ahwhptt.edu.cn
References
1，Buckman RF, Woods M, Sargent L, et al.A unifying pathogentic mechanism in the etiology of intraperitoneal adhesion. J Sur Res，1976; 20:1
, http://www.100md.com
2，Menzies D, Ellis H. Intro-abdominal adhesions and
their prevention by topical tissue plasminogen activator. J Roy Soc Med， 1989; 82:534
3，Wang PJ,Zheng QS,Wang M, et al. Quantitative analysis of effects of allantoin, dexamethasone and metronidazole in compatibility to indices related introabdominal adhesion. Chin Pharmacol Bul， 2000; 16
(accepted)
4，Swolin K. Experimental study of prophylaxis of intraabdominal adhesions.Acta Obstet Gynecol Scand, 1966;45:473
, http://www.100md.com
5，Beck JR, Shulz EK. The relative operating charac-
teristic (ROC) curves in test performance evaluation. Arch Pathol Lab Med， 1986; 110:13
6，Zheng QS, Cheng NN, Sun RY. Evaluation on experimental performance with the relative operating characteristic curve plot. Chin J Lab Diagn， 1998;2:
171
7，Raftery AT. Effect of peritoneal trauma on peritoneal fibrinolytic activity and intraperioneal adhesion formation. Eur Surg Res， 1981;13:397
, 百拇医药
8，Vipond MN, Whawell SA, Thompson JN, et al.
Peritoneal fibrinolytic activity and intra-abdominal adhesions. Lancet,1990;335:1120
9，Evans DM, McAree K, Guyton DP,et al. Dose dependency and wound healing aspects of the use of tissue plasminogen activator in the prevention of intra-abdominal adhesions. Am J Surg,1993;165:229
2000-01-24 received, http://www.100md.com
单位：郑青山(中国科学技术大学生命科学学院，合肥 230027)；桂常青 孙瑞元(皖南医学院药物研究所，芜湖 241001)；王民(山西皮尔复临床医药研究所，太原 230006)
关键词：动物模型；粘连；联合药物治疗；ROC曲线；尿囊素；甲硝唑；地塞米松
中国临床药理学与治疗学000202 目的 制作一种新的大鼠腹腔粘连模型，并确定腹腔渗液中组织纤溶酶原激活物活性(PAA)是否具有判断粘连形成的能力。方法 大鼠随机分为2组，麻醉后不作任何消毒，打开腹腔，找到回盲部。距盲端约1～2 cm处结扎，剪去盲端的多余部分，挤去敞口盲端内的肠内容物，不作消毒，放回腹腔。在腹腔内放置直径约4 mm的引流软管。作两层缝合。皮肤层缝合前，用药组注入抗粘连组方(AMD)(尿囊素+甲硝唑+地塞米松)，剂量为1.5 ml.100 g-1体重。对照组注射等容量5 % GS。6 h后抽取腹腔液1 ml，拨除引流管。1 w后脱臼处死大鼠，打开腹腔检查粘连的形成及其程度。用相关操作特征图 (ROC) 分析PAA是否具有判断粘连形成的能力。结果 对照组的20只大鼠均形成腹腔粘连，腹腔渗液量为(1.25±0.09) ml,WBC(×103) =(23.1±6.6) mm3，PAA=(0.9±0.4) IU.ml-1。用药组没有形成腹腔粘连(0/20)，腹腔渗液量为(0.52±0.04) ml (P<0.01),WBC (×103) 为(10.6±4.2) mm3 (P<0.01), PAA 为(23.1±6.6) IU.ml-1(P<0.01)。由于此3药联用的抗粘连作用与腹腔炎性渗液量减少，抗菌抗炎作用，以及提高渗液中PAA有关，从而反映了本模型的形成机理。ROC分析认为， PAA可作为判断粘连是否形成的重要指标，当PPA>1.24 IU.ml-1时，粘连不易形成，反之则较易形成。结论 本动物模型是研究大鼠腹腔粘连的有效工具，腹腔渗液中PAA活性的变化对大鼠腹腔粘连的形成具有判断能力。
, 百拇医药
中图分类号 R965.1
A novel animal model of intra-abdominal adhesion and
quantitative evaluation with related indices
ZHENG Qing-Shan
(School of Life Science，University of Science and Technology of China, Hefei 230027)
GUI Chang-Qing
(Institute of Materia Medica, Wannan Medical College, Wuhu 241001)
, http://www.100md.com
SUN Rui-Yuan
(Institute of Materia Medica, Wannan Medical College, Wuhu 241001)
WANG Ming
(Pierfu Institute of Clinical Medical Research, Taiyuan 030006)
Aim To set up a novel animal model of intra-abdominal adhesion and to determine whether the tissue plasminogen activator activity (PAA) in exudate can be taken as an indicator to judge the formation of the adhesion. Methods Rats were randomly divided into 2 groups. Each animal in both groups was opened the abdominal cavity via midline laparotomy without any disinfectant measures. 2-cm section from the cecal end was clamped and ligated, 1-cm cecum of the section was cut, and another 1-cm end from the ligated site was kept. After the content in the end was extruded, the cecum was put back without using any antibacterial agent. Before the skin closure, an effective combination AMD (allantoin, metronidazole and dexamethasone in combination), was given (ip) according to 1.5 ml per 100 g body weight (60.6 mg.kg-1). The control group was injected (ip) the same volume of normal saline. After 6 h, the exudate was extracted from drainage-tube, with the rats varying posture, and after 1 kw, the rats were killed for examining the intra-abdominal adhesion. The values of PAA of exudate in both groups were analyzed by the relative operating characteristic curve (ROC). Results In the control group, all 20 rats formed the adhesions, the amount of exudate=(1.25±0.09) ml, the number of WBC(×103)=(23.1±6.6) mm3 and PAA=(0.9±0.4) IU.ml-1 in the exudate of abdominal cavity. In AMD group, however, there was not the adhesion formations (0/20), the amount of exuade was (0.52±0.04) ml (P<0.01), the number of WBC (×103) (10.6±4.2) mm3 (P<0.01), and PAA (23.1±6.6) IU.ml-1(P<0.01) in exuade.ROC analysis indicated that if PAA >1.24 IU.ml-1 in the exuade, the adhesion was difficult to form, and vice versa. Conclusion This animal model can be taken as an effective tool to evaluate the human adhesion related to multi-links on the pathogenesis, and the PAA in exudate an indicator to judge intra-abdominal adhesion formation.
, http://www.100md.com
Key words animal model; adhesion; combination drug therapy; ROC curve; allantoin; metronidazole; dexamethasone
The etiology of intra-abdominal adhesion formation involves many factors, such as tissue ischemia, infection, inflammation and exudation, and the mechanisms have been attributed to peritoneal plasminogen activator activity (PAA) decreased in peritoneum over the past 3 decades[1,2]. On therapeutics and prevention, however, there has been just a little advance because numerous agents and surgical techniques often obtain conflicting results. The cause may be mainly, at least partly, multi-factor in the adhesion etiology and multi-pathway in the adhesion mechanism, which makes the adhesions become difficult to prevent by using a single drug or certain a measure. In this study, we firstly made the animal model of intra-abdominal adhesion caused by multi-factor,then an effective combination (AMD)[3], allantoin (Alt), metronidazole (Met) and dexamethasone sodium phosphate (Dex), of preventing the adhesion formation was used to evaluated the model and to explore its pathway. Lastly, the relationship between the PAA in exudate of abdominal cavity and the adhesion would be assessed to determine whether the PAA could be taken as an indicator to judge the formation of intra-abdominal adhesion.
, 百拇医药
1 Experiment
1.1 Materials Alt powder was obtained from Jiangsu Huanghai Pharmaceutical Factory and the purity=99.6 %,Met powder from Tianjing Hebei Pharmaceutical Factory and the purity=99.85 %, and Dex powder from Roussel Uclaf Co and the purity=99.6 %.The compatibility AMD(the patent register code: 97110167.1), provided by Prof. Wang M, was composed of Alt,Met and Solution was prepared with 5 % GS. The assay kit of tissue plasminogen activator (PPA) was purchased from FJ Sun Biotech Co, №990406.
, http://www.100md.com
Wistar rats(weighing 190～210 g) of both sex were purchased from the Animal Center of Shanghai Medical University (Grade Ⅱ, Certificate №02-22-12).
1.2 Animal models of intra-abdominal adhesions Rats were randomly divided into 2 groups of 10 rats. Anesthetized with 3 % sodium pentobarbital (1 ml.kg-1, ip), each animal in both groups was opened the abdominal cavity via midline laparotomy without any disinfectant measures. 2-cm section from the cecal end was clamped and ligated, 1-cm cecum of the section was cut, and another 1-cm end from the ligated site was kept. After the content in the end was extruded, the cecum was put back without using any antibacterial agent. Then the end of operation knife was used to grind the parietal peritoneum on the anterior abdominal wall for 5 times. A soft drainage-tube, 4 mm in diameter, was put. The wound closure was accomplished using 3-0 silk for the abdominal medline and 4-0 silk for the skin. Before the skin closure, AMD was given (ip) according to 1.5 ml per 100 g body weight (60.6 mg.kg-1). The control group was injected (ip) the same volume of normal saline. Then the drainage-tube was clamped, fixed and covered. After 6 h, the exudate was extracted from drainage-tube, with the rats varying posture. If the volume did not reach to 1 ml was obtained, a small amount of normal saline was injected via the tube to wash the abdominal cavity, and the washing liquid kept for 3～5 min was collected until 1 ml. The sample contained a lot of blood would be discarded. The collected exudate was put into anticoagulated silicic tube and centrifuged at 1500×g for 10 min. 200 μl supernatant mixed with the same volume of a kind of acidic liquid was kept at 2～8 ℃ for use. Then the drainage- tube was taken out. After 1 wk, the rats were killed for examining the intra- abdominal adhesion. The adhesions were scored by a modified scale devised by Swolin [4]: 0 (no adhesions),1 (filmy connections separated spontaneously),2 (filmy adhesions separated by gravity),3 (filmy adhesions by traction), and 4 (Dense adhesions requiring sharp dissection).
, 百拇医药
1.3 Determination of indices 1) PAA: it was determined by the chromogenic substance method. The procedure of the experiment was done according to the PPA assay kit introduction. Exudate absorbance (A) in 96-well plate was determined at 405 nm with a microplate photometer (Yutai Yanghang Co, Hong Kong). If the exudate was diluted,A must multiply the multiple of the dilution; 2) the exudate volume in abdominal cavity; 3) the number of WBC in the exudate.
1.4 Data analysis A cutoff of PAA between the AMD group and the control group was the maximum sum of the sensitivity and the specificity of PAA.It denoted that the rats with PAA >the cutoff were not easily to form intra-abdominal adhesions, and vice versa. The experimental performance was determined by the relative operating characteristic curve (ROC)[5,6]plot,on which x-coordinate was 1 minus specificity and y-coor- dinate was sensitivity. The area under ROC and its standard error were expressed as W±SEw, and analyzed by z-test.
, 百拇医药
2 Results
2.1 Preventive effect of AMD on intra-ab- dominal adhesions in rats The 20 control rats were formed strip or round adhesions of intestinal canal (Fig 1), each score > 1 and the mean score was 2.4±0.6. But there were not found obvious adhesions in AMD group, each score<2 and the mean score was 0.3±0.4 (P<0.01vs control).
Fig 1 Formation of the rat model of intra-abdominal adhesion
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Left:round intestinal canal; Right:strip intestinal canal
2.2 Influence of AMD to some indices related intra-abdominal adhesions Compared with the control group,the PAA increased (P<0.01), the exudation volume and the number of WBC decreased (P<0.01) (Tab 1). It indicated that AMD could decrease fibrin into abdominal cavity and increase fibrinogenolysis.
Tab 1 Preventive effect of AMD(Alt+Met+Dex) on intra-abdominal adhesion in rats (
±s,n =19) Groups, http://www.100md.com
Exudate
PAA
WBC(×103)
AMD
0.52±0.11
1.6±0.8
10.6±4.2
Saline
1.25±0.09*
0.9±0.4*
23.1±6.6*
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*P<0.01 by t test.PAA=the tissue plasminogen activitor activity
2.3 Cutoff of PAA between adhesions and no adhesions, and analysis of experimental performance According to the order from small to large, each value of PAA was selected as a cutoff, respectively, to form a series of four-cells like Tab 2, and all the sum of the sensitivity and the specificity in each point were calculated. The maximum sum was the real cutoff (C point on Fig 2), at which sensitivity + specificity = 0.84+0.74 =1.58, and the cutoff = 1.24 IU.ml-1. χ2=10.19, which was also the largest value in all four-cell tables. It indicated that the adhesion was not easily formed when PAA was over 1.241 IU.ml-1.
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Fig 2 Each value of the tissue plasminogen activator activity (PAA) corresponded to sums of sensitivity and specificity.
Point C corresponding to the maximum sum (1.58) in y-axis and PAA ( 1.241 IU.ml-1) in x-axis.
Tab 2 Four-cell table Cutoff/IU.ml-1
Adhesion
No adhesion
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<1.214
16(a)
5(b)
≥1.241
3(c)
14(d)
χ2 =10.19,P<0.01；Sensitivity=a/(a+c)=16/(16+3)=0.84；
Specificity=d/(b+d)=14/(14+5)=0.74
The results of the experimental performance (Fig 3):W=0.764, SEw=0.0837, z=3.16, P<0.01. It meant that there was a significant difference between the PAA in adhesion animals and that in no adhesion animals, indicating the adhesions were distinguished by the PAA (< cutoff= 1.241 IU.ml-1) of exudate in abdominal cavity.
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Fig 3 Analysis of ROC curve plot to the ability of the tissue plasminogen activator activity (PAA) in judgement of the intra-abdominal adhesion formation.
3 Discussion
Intra-abdominal adhesion is a common irreversible syndrome, so the goal of new drug development is how to prevent it. The PAA decreased in peritoneum has been taken as main mechanism of adhesion formation by many investigators[1,7,8]. The availability of recombination tissue plasminogen activator (rtPA) has also been conformed[9].If an investigator take a piece of peritoneum from no general peritonitis to determine the PAA, the location of sample possibly affects the result. In addition, it is a kind of traumatic technique and difficult to apply in clinical study. So the determination of PAA in exudate has some advantages for use. However,the relevance between PAA in peritoneum and in exudate is unclear, so it is necessary for quantitatively evaluate the relationship between PAA and the adhesion with ROC analysis. The analysis has been taken as the most important method for judging the experimental performance in recent decades, especially for determining a cutoff. The PAA in abdominal cavity can be taken as a valid indicator in judgement of the adhesion formation in this study.
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The process of making animal model in this study reflected the complexity of etiology, which involved ischemia, mechanical stimulation,trauma,exudation,infection,and inflammation. It may reflect the nature of
intra- abdominal adhesion much effectively.
The compound drug, AMD, can prohibit the fibrin rich exudate into the abdominal cavity and at the same time increase fibrinogenolysis by some positive approaches, such as anti-inflammation, anti-bacteria,anti-exudation, and increasing PAA[3].These factors may show some pathways of the animal model formation.
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Project supported by the National Natural Science Foundation of China, № 39670845 and the Natural Science Foundation of Anhui Province, № 98454735.
Correspondence to Dr ZHENG Qing-Shan.
Phn: 86-551-360-3754. Fax:86-551-360-3754.
E-mail: zhengqs@mail.ahwhptt.edu.cn
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2000-01-24 received, http://www.100md.com