结肠液囊管一期修复结直肠穿孔及吻合口漏的实验研究
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摘要
研究背景与目的
结直肠穿孔发病率约为结直肠癌总数的3-8%,包括肿瘤在内的各种疾病自发性肠穿孔约为1.2-4.6%。自发性结直肠穿孔发生于乙状结肠及乙状结肠直肠交界处的系膜对侧缘>50%,原因与乙状结肠特殊生理解剖因素有关。自发性结肠穿孔多发于反应相对迟钝的60岁以上老年人,病史难于叙述清楚,加上结肠穿孔缺乏特异临床表现,早期正确诊断不足10%。由于结肠内容物含有大量细菌,穿孔确诊后已产生严重弥漫性腹膜炎,加上细菌毒素的大量吸收,形成感染抗体并导致多器官功能衰竭,故病死率可高达到35-47%。
各种方式的结直肠吻合术后的吻合口漏(Anastigmatic Leakage,AL)发生率大约为3-9%,TME手术AL发生率2-19%,结肠AL病死率12.9%。
也就是说我国每年需进行结直肠穿孔或AL修补的病例相当庞大。而对于上述病人的传统治疗原则上均采取肠穿孔或AL全层间断缝合浆肌层加固,同时行近段结肠腹壁造瘘,2-3个月后二期再次行造瘘肠管回纳术(肿瘤除外)。这种处理方式病程长,痛苦大,特别是腹壁造瘘留下巨大的疤疤,两次手术易导致严重肠粘连并发症以及高额费用,给患者带来极大的心理压力和躯体痛苦。如何才能安全有效地简化此类疾病的治疗流程,缩短病程,降低费用,寻求到一种微创,美观,并发症低的手术治疗方式呢?目前许多学者特别是胃肠外科医务工作者正不竭努力地进行研究。特别是近年来随着以腹腔镜为主体的微创胃肠外科的发展及各种可吸收人工生物材料的临床应用增多,使此项研究工作的进展逐渐加快。正是在这种背景下,本实验设计出结肠液囊管(colon sac duct,CSD)来有效地解决需要解决的问题。并对此项技术进行了认真科学的研究,期望这一研究能在结直肠穿孔及AL的治疗领域有所突破。
研究目的：
1总结既往外科临床对于结直肠穿孔及结直肠AL成功进行各种手术修复的经验,比较传统手术成功的关键因素及存在的不足,结合人体结直肠具有肛门这一个通向体外的自然通道,通过实践研究设计并制作出一种能经肛门进出,可顺利到达人体结直肠各段,并能在腹腔镜协助下对结直肠穿孔及结直肠AL进行有效一期修复的医疗器械-CSD。并对CSD各项性能进行客观检测,以求证其用于实验及临床的安全性,可靠性,有效性。
2通过对实验用西藏小型猪进行动物实验研究CSD能否作为一种安全可靠的新技术对结直肠穿孔及结直肠AL进行有效的一期修复。特别是能否在结直肠穿孔或结直肠AL48-72小时以上,严重感染的腹腔环境下有效修复结直肠穿孔及结直肠AL。以传统外科手术作为对照,进一步求证CSD作为一种新技术的科学性,安全性,可靠性,有效性及微创优势。
研究内容与方法
1 CSD的样品制作
1.1制定预设方案：制定三种预设方案,查阅文献,预实验中对大约35kg的实验用西藏小型猪进行解剖,了解它的结直肠肠腔内径大小。作为CSD尺寸设计的依据,制定出CSD三种预设方案,并绘制出工程制作线图及立体彩图。
1.2材料选择：在高分子材料工程师的帮助下,对临床上已广泛使用且被证实具有良好生物组织相容性的几种医用级高分子材料的弹性模量、断裂强度、断裂伸长率进行充分比较后选择出硅橡胶、聚氨脂用于CSD的样品制作。
1.3样品制作：寻找选择样品制作单位,根据本实验提供三种设计方案的立体彩图,工程制作线图,进行模具制作,根据需要反复修改模具,制作出三种预设方案的CSD样品。
1.4确定最终样品：CSD的结构如下：主要由外套囊(1)与内管(2)组成,其中外套囊两端有环形凹槽(3),用于固定及隔绝,在外套囊与内管之间形成一密闭的环形空间(4),利用热融合技术使二者粘合成一体。外套囊一端的充气管(5)与所述环形空间相通用来注射空气充盈外套囊。外套囊质地柔韧,内管质地坚硬,外套囊两端向内收缩,延伸至内层管壁。这种设计模型可用于结直肠穿孔或结直肠AL的一期修复手术。它既可阻挡肠腔内压力向穿孔或漏口处传递,又可使病变肠管与肠内容物完全隔绝。形成一个非常有效的肠腔内导流管装置,故有利于病变肠管清洁低压的环境下一期愈合,达到甚至超过传统二期手术的效果。
1.52.3样品性能检测
利用三通管将CSD的充气管、精密压力表进行连接,向液囊管内匀速充气,观察CSD外套囊、内管是否变形。凹槽是否外凸变浅,液囊管是否漏气或破裂,同时记录压力表所显示的压力读数。并进行统计学分析。对样品的性能作出科学客观的分析比较。
2 CSD修复结直肠穿孔及结直肠AL动物实验
2.1实验设计
处理组采用CSD对结肠穿孔及结直肠AL进行一期修补,以传统一期穿孔或漏口缝合近段结肠造瘘,二期结肠回纳手术作为对照组,每组15例。比较两种手术方式的手术时间、术后腹腔粘连程度、安全性、可靠性、有效性。并在修补后7、14、21天检测穿孔及漏口的愈合情况：比较结直肠穿孔及漏口部位的愈合强度——爆破压,结肠愈合部位组织中微血管密度(microvessel density,MVD)及羟脯氨酸含量,评估CSD一期修补结直肠穿孔及结直肠AL的实用性、可靠性、安全性及微创效果。
2.2动物模型制造
电视腹腔镜监视下经肛门制作动物模型,采用实验用西藏小型猪,体重30-35kg。氯胺酮联合戊巴比妥麻醉。麻醉前半小时称体重,臀部肌肉内注射鲁米那钠100mg、阿托品0.5mg。麻醉诱导按氯胺酮6mg/kg耳根部注射给药进行,待实验猪安静少动时经行气管插管(7.0-7.5号)并固定,呼吸机辅助呼吸。手术过程中以氯胺酮,戊巴比妥或异丙酚静脉滴注维持。
连接各线路及管道,开启调制电视腹腔镜的冷光源,CO2气腹机、高频电刀,调节腹腔镜视管的黑白及清晰度。于肚脐右上缘切约11mm皮肤皮下切口,常规建立气腹,当腹内压达到10-12mmHg状态时。以10mmTrocar穿刺成功后,置入腹腔镜视管。观察腹腔内各脏器,排除腹腔内异常情况后,从肛门置入另一视管(代替肠镜)。在腹腔镜监视下进入到距肛门约25-30cm处于肠系膜对侧缘刺破肠壁,形成大约1.0×1.2cm2穿孔,制造结肠穿孔模型。或利用举肠器有效显露距肛门约25-30cm处结肠,以电凝勾切断肠管,以丝线缝合肠管两断端,留取大约三分之一的肠系膜对侧缘肠壁不缝合,制造结肠AL模型。
2.3应用CSD对结肠穿孔或AL进行一期修复
第一次手术后48小时,再次手术,在腹腔镜下应用CSD对存在穿孔或漏口的结肠进行一期修复。麻醉、体位固定及消毒,铺无菌巾单同前述动物模型制造。于原脐上切口处拆除缝线,以弯钳逐步分离腹壁各层,进入腹腔,注意防止此处有肠管粘连而被破损伤,直视下,将10mmTrocar置入腹腔内,并置入10mm腹腔镜视管,仔细观察腹腔内粘连情况,避免损伤粘连肠管。在腹腔镜监视下于左右下腹合适位置,各切一长约5mm切口,并置入5mm Trocar后,置入相应的腹腔镜器械。轻柔推拉分离腹腔内粘连,清除腹腔内存留的粪便、脓苔寻找穿孔或漏口部位。如果寻找困难则以温盐水轻柔冲洗腹腔,吸净后仔细寻找。找到穿孔或漏口后,以四号丝线园针,纵行方向上缝吊,切取少许异常肠壁组织送病理。另一手术组成员以手指轻柔扩肛,逐渐扩张至2指宽后置入肛管,以大量稀释碘伏冲洗肠腔,至流出的冲洗液基本不带粪碴后使用石蜡油润滑肠腔及CSD,然后在腹腔镜监视下应用推送器将液囊管轻柔推送至肠穿孔或漏口部位,并使肠穿孔或漏口处位于液囊管的中部,缓慢均匀地对液囊管进行充气,致液囊管充分膨胀具有一定的韧性,又不致过分粗硬扩张压迫肠管,造成肠壁张力过大,影响病变部位肠管的血运。液囊管在肠腔内能具有一定的活动度为佳。在推送器内置入金属杆,使推送器灵活转变为类似于妇产科举宫器相仿的举肠器,通过举肠器的辅助使穿孔部位结肠及其系膜能上下左右摆动,并形成一定的张力,清楚地显露出结肠系膜的无血管区,利于手术者操作。手术者使用分离钳于液囊管两环形凹槽部位紧贴肠外壁穿过结肠系膜无血管区,不损伤肠壁血运情况下,用可吸收羊肠线适度捆扎肠管于液囊管上,将穿孔或漏口部位肠管与肠腔完全隔绝,大量温盐水冲洗腹腔。仔细止血,清点纱布器械,放尽腹腔积气,撤除器械,关闭各切口,固定液囊管的冲气管后结束手术。术后1小时,麻醉作用消失,给予正常饮水,术后禁食24小时,第2天开始给予无渣流质饮食。
对照组的传统手术：麻醉、体位固定、消毒、铺巾、从左侧经腹直肌切口进腹、探查穿孔部位,全层缝合。手术切口前端左侧旁开大约5.0cm处,将皮肤剪一直径约3.0cm的圆形切口,电刀切开皮下组织、肌筋膜、钝性分离腹肌,切开腹膜,选择合适的结肠段,经造瘘口提出到腹外,以塑胶棒及橡胶管穿过肠系膜无血管区,将肠管固定于腹外,电刀纵行切开对系膜缘肠壁约3.0cm,将切开的肠管壁缝合固定于造瘘口上。腹腔冲洗,仔细止血,留腹腔引流管,清点纱布器械,关腹结束手术。
2.4第三次手术对修复效果进行检测
分别于手术后第7天、14天、21天对处理组及对照组的西藏小型猪结肠穿孔或AL修复的效果进行检测,腹腔镜下检测的项目为腹腔内粘连情况、肠AL部位有无狭窄、开腹检测肠穿孔或AL愈合部位的爆破压,并取样检测愈合肠壁组织的羟脯氨酸含量及MVD。
2.4.1活体状态下结肠AL或穿孔愈合部位爆破压的测定
确定CSD位置,于其近端约5.Ocm处用编织带结扎结肠,放尽CSD内气体,取出CSD,将精密压力表连接管经肛门插入直肠15.0-20.0cm,用7.0号丝线荷包缝合肛门,确保不会渗气,使穿孔或AL处结肠管与精密压力表处于同一水平,缓慢匀速地向肠腔内打气,助手密切观察并记录穿孔或AL结肠修复部位漏气时压力表的读数,即为结肠穿孔或AL修补部位的爆破压。
切取病变愈合部位肠壁组织,分为5份。1份浸泡在10%福尔马林中固定,进行HE染色,其余4份保存在-80℃冰箱中,分别用于检测病变愈合部位组织中羟脯氨酸含量及MVD。
2.4.2 AL或穿孔愈合处组织羧脯氨酸含量测定
使用碱水裂解法羟脯氨酸检测试盒：精确称取湿重组织,准确加碱水裂解液,调节PH值,在水解液中加入活性炭,取上清液,依次加入试剂,水浴,冷却,离心,取上清液,测吸光度,按公式计算样品中羟脯氨酸含量。
2.4.3 AL或穿孔愈合处组织MVD测定
组织OCT包埋切片,丙酮中固定,常规水化,PBS冲洗,消除内源性过氧化物酶的活性,PBS冲洗,山羊血清封闭室温孵育,倾去血清,滴加一抗工作液,PBS冲洗,滴加生物素标记山羊抗小鼠IgG, PBS冲洗,滴加辣根酶标记链霉卵白素,PBS冲洗,配置DAB工作液,滴加(DAB)显色剂显色,自来水充分轻柔冲洗,苏木素液一滴滴于组织上复染,PBS冲洗,脱水,玻片干燥后滴加中性树脂封片,微血管计数。
2.5统计学分析
应用SPSS13.0软件包统计分析,计量数据以均数±标准差(X±s夕表示,两组间比较采用两独立样本的t检验(双侧)分析,P<0.05为有显著性差异。
结果
1样品性能检测
1.1 CSD外套囊变形压力比较
医用硅橡胶与医用聚氨酯制作的CSD外套囊变形压力分别为：30.42+0.46Kpa与30.73±+0.29 Kpa。结果显示医用聚氨酯稍优于医用硅橡胶,但无统计学差别
1.2 CSD内管变形压力比较
医用硅橡胶制作的CSD内管变形压力值为5.96±0.22 Kpa, CSD的内管硬度不够,容易内突导致管腔狭窄闭合,而医用聚氨酯制作的CSD内管变形压力值大于100 Kpa时均无明显变形。故医用聚氨酯明显优于医用硅橡胶。
1.3 CSD漏气压力比较
用医用硅橡胶与医用聚氨酯制作的CSD漏气压力值为59.23±1.25Kpa与60.01±±1.45 Kpa,结果显示医用聚氨酯稍优于医用硅橡胶,但无统计学差别。
2结肠穿孔修复术后愈合部位检测
2.1结肠穿孔修复术后爆破压检测
术后第7、14、21天处理组与对照组爆破压分别为：26.20±+2.76、36.12±±1.67、38.58±1.46与2128±0.80、27.72±1.07、35.92±+2.01,处理组均优于对照组(t=3.83,P=0.005；t=9.46,P<0.001；t=2.39,P=0.044)。
2.2结肠穿孔愈合部位羟脯氨酸含量检测
术后第7、14、21天处理组与对照组羟脯氨酸含量分别为：21.42+0.76、25.96±±0.87、27.50±0.59与15.14+0.97、21.08±±1.08、26.90±0.56,术后第7、14天处理组羟脯氨酸含量较对照组高(t=11.39；t=7.82,P<0.001),术后第21天2组无显著差别(t=1.65,尸=0.139)。
2.3结肠穿孔愈合部位MVD检测
术后第7天处理组与对照组MVD分别为：12.40±0.89与10.40±±1.14,术后第7天MVD处理组高于对照组(t=3.09,P=0.015)。
3结肠AL修复术后愈合部位检测
3.1结肠AL修复术后爆破压检测
术后第7、14、21天处理组与对照组爆破压分别为：23.54±1.41、34.14+1.86、36.70±+1.52与19.10±0.57、25.96±1.17、34.00±2.15,术后第7、14天处理组均优于对照组(t=6.54,P<0.001；t=8.31,P<0.001),术后第21天2组无显著差别(t=2.29,P=0.051)。
3.2 AL愈合部位羟脯氨酸含量检测
术后第7、14、21天处理组与对照组羟脯氨酸含量分别为：20.46±±0.76、25.08±±1.07、26.10+0.61与14.20±+0.94、20.22±±1.22、26.70±0.67,术后第7、14天处理组羟脯氨酸含量较对照组高(t=0.41,P<0.001；t=0.82,P<0.001),术后第21天2组无显著差别(t=0.85,P=0.179)
3.3 AL愈合部位MVD检测
术后第7天处理组与对照组MVD分别为：11.20+0.84与8.80+0.84,第7天MVD处理组高于对照组(t=4.54,P=0.002)。
结论
1.采用聚氨酯制作的CSD结构简单、性能优良、操作方便、安全可靠,有临床应用价值。
2.应用CSD技术一期修复结肠AL或穿孔是安全、可靠且有效。
本研究创新之处
1自主研究设计的CSD能经肛门自由进出能到达人体结直肠各处,在腹腔镜下
对结直肠穿孔及AL进行一期修补。
2 CSD治疗效果安全、可靠,能有效取代传统一期修补造瘘,二期结肠回纳手术。
Background and objective
Incidence rate of colorectal perforation is about 3-8% of the total number of colorectal cancer, spontaneous colon perforation in all diseases is about 1.2-4.6% including various tumors. Spontaneous colon perforation occur at opposite mesentery edge of the sigmoid colon or the junction of the sigmoid colon and rectum,the rate is more than 50% due to special physiological and anatomical factors at sigmoid colon. Spontaneous perforation of the colon happens to mainly the 60 years old persons or elders,because the case history is difficult to describe clearly,and with the lack of specific clinical manifestations of colon perforation, early diagnosis is less than 10%. Since a large number of bacteria are in colon contents, serious diffuse peritonitis happens after diagnosis, a substantial absorption of bacterial toxins,the formation of infection antibodies and lead to multiple organ failure, so the mortality can reach 35-47%.
The rate of various forms colorectal anastomosis leakage(AL) is about 3-9%, TME is 2-19%, the mortality rate of AL is 12.9%, AL happens to mainly the elders and the mortality is about 35-47%.
The cases of colorectal perforation or AL repair are large each year in China. The traditional treatment for these patients is taking on the principle of full-thickness interruption suture and seromuscular layer strengthened,and the colostomy in proximal colon through abdominal wall,2-3 months later the colostomy is closed and returned back to the abdomen (except cancer).This approach is characteristic of longer duration, large pain, especially the huge scar left on the abdominal wall,the complications including colon adhesion,the high cost. tremendous psychological pressure and pain due to twice surgery. How to safely and effectively simplify the treatment of this disease, shortening the course, reducing costs and seeking to a minimally invasive, beautiful and low complications surgical treatment? In recent years, minimally invasive gastrocolon surgery with laparoscopy as the main body and clinical application of artificial absorption biological material are increasing. Therefore, colon sac duct (CSD)can effectively solve the problems we face.
Purpose:
1 Summarizing successful experience of the previous clinical surgery for repairing colorectal perforation and AL, analyzing traditional surgical critical successive factors and shortcomings,human anus which has a natural channel leading to out-of-body and practical researchs I designed and made medical equipment-CSD that can reach the any parts of the body and effectively repair colorectal perforation and AL through the anus,furthermore tested the performance of CSD objectively to verify safety, reliability and validity for experimental and clinical application.
2 Through experimental studies with Tibet mini-pig colon I verified that CSD can used as a safe and reliable new technology for repairing colorectal perforation and AL effectively,in particular when colorectal perforation or anastomosis leakage is more than 48 to 72 hours,there is severe abdominal infection. Traditional surgery as the control, and further verified if CSD has a new scientific technology, security, reliability, validity and minimally invasive advantage.
Contents and methods
1 CSD prototyping
1.1 Making the default program
Making three preset programs, reviewing literature in preliminary experiments. About 35kg Tibet mini pig was dissected to understand its colorectal lumen diameter size as basis of the colon sac design.making CSD three preset programs, and drawing projects map and three-dimensional color pictures.
1.2 Material selection
With the help of polymer material engineers, comparing elastic modulus, tensile strength, break elongation rate of several medical grade polymer material widely used in clinic and with good tissue compatibility, I selected the silastic, polyurethane for CSD samples at last.
1.3 Sample production
Looking for samples production units, according to our design that provide three-dimensional color pictures,engineering production line graph,We made the mold, repeatedly revised the mold as needed to produce the three preset CSD samples.
1.4 Determine the final sample
CSD structure is as follows up:mainly with the outer capsule (1) and inner duct (2),2 circular grooves at both ends of outer capsule (3) for fixing and isolateing, there is a closed annular space between the outer capsule and the inner duct (4), with hot fusion technology to bond the two into one capsule for filling capsule with saline/gas,The inflatable duct is at one side of the capsule (5) and communicates annular space. Outer capsule texture is flexibile, inner duct texture is hard, The outer capsule contracts at both ends, extending to the inner duct wall. This design model can be used for repairing colorectal perforation or AL in 1st stage, which can block the pressure of the perforation or leakage passing the wound, the wound can also completely isolated from the colon contents.These is good for healing.
1.5 sample performance test
The CSD,inflatable duct and precision pressure gauges were connected by three-way duct,then aerateed to observe if the CSD and inner duct were deformed,circular grooves were convex, sac leaked or broke,at the same time recording the pressure readings on pressure gauge.
2 CSD repairing colon perforation and AL in animal experiments
2.1 Experimental Design
Tibet mini-pigs were randomly divided into 2 groups, treatment group and control group,15 per group, repairing colon AL of treatment group with CSD, the control group with traditional suture and colostomy in a proximal colon in 1st stage,then put colostomy back in 2nd stage.Comparing operation time, postoperative abdominal adhesions,safety,reliability, validity.at the at 7th,14th,21st days later,detecting the anastomotic leak condition:Comparing of the healing of AL intensity:burst pressure, the tissue micro vessel density(MVD) and hydroxyproline contents in healing parts, assessing practicality, reliability, security, and minimally invasive effects of CSD repairing colorectal perforation and AL.
2.2 Making animal model
Animal models were made under the surveillance of laparoscopic transanal using experimental Tibet mini-pigs, weight 30-50kg. anesthesia wtih Ketamine combined with pentobarbital. Weighing before anesthesia, intramuscular injection luminal sodium 100mg, atropine 0.5mg at the hip. Anesthesia was induced by injection of ketamine 6mg/kg by ears, when the pig became quiet, endotracheal was dealed (No.7.0) and fixed. Maintaining pentobarbital or propofol by intravenous infusion during surgery.
Connecting lines and piping, opening the cold light, modulating laparoscope, CO2 pneumoperitoneum machines, high frequency electroscalpel, laparoscopic black and white and clarity adjustment. Conventional establishment of pneumoperitoneum at upper-right navel, when intra-abdominal pressure reached 10-12mmHg state, the 10mmTrocar puncture, laparoscopic vision duct were placed, observing the organs in the abdominal cavity to exclude intra-abdominal abnormalities. Inserting vision duct, from the anus,under the supervision of the laparoscope arriving about 25-30cm from the anus, at the opposite edge of mesenteric piercing the colon wall, formating about 1.0×1.2cm perforation, then colon perforation model was made. About 25-30cm from the anus at the colon, cutting off the colon with electric coagulation hook then suturing the two ends of colon with silk, but 1/3 circle colon was left for non-closure at contralateral margin of mesenterium, then colon AL model was made.
2.3 Application of CSD for the colon perforation or AL in repairing in 1st stage.
Re-operation at 48 hours after the first surgery, in the application of laparoscope repairing the leakage and perforation with CSD. Anesthesia, position and disinfection, sterile towel were the same foregoing. Through original suture incision separating the abdominal wall layers gradually with curved forceps, entering the abdominal cavity, there was attention to prevent colon injury due to adhesion, Inserting lmm Trocar into the abdominal cavity, and placing 10mm vison duct, carefully observing the intra-abdominal adhesions, avoid damaging colon. At proper position in the left and right lower abdomen under laparoscopic monitoring cutting a incision about 5mm, and implanting 5mm Trocar and the corresponding laparoscopic instruments. Separating intra-abdominal adhesions and clearing the pus and abdominal stool left gently, looking for perforation or leak. If you had difficulty in finding them, rinsing gently with warm water and sucking carefully. After finding perforation or leakage,longitudinally seaming with code 4 thread and circular needle, hanging, cutting a small abnormal tissue for pathology. Another members expanding the anal with fingers gently and gradually to 2 finger width, then inserting the anal speculum, washing the colon with a large number of diluted povidone-iodine,Lubricating CSD and colon lumen with paraffin oil, then under laparoscopic surveillance pushing the sac duct to the place of perforation or leakage with pushing device, making sure that the perforation or leakage was in center of the sac duct, inflating with aeration the sac duct to make it tough, but not too hard, avoiding expanding and oppressing bowel, causing too large bowel wall tension, effecting lesion revascularization. The sac duct in the colon lumen duct had a certain activity. Inserting metal rod into the pushing device, the device is similar to the lift-uterus device in obstetrics and gynecology, making perforation part of the colon and its mesentery swing up and down, and a certain tension with the auxiliary,clearly revealing the avascular zone, which will help surgery operation. With the separation clamp permeating the mesocolon avascular zone on the two circular groove location of the sac duct close the colon wall, not damage the colon blood flow, fixing colon in the sac duct moderately with absorbable catgut,making the parts of the colon perforation or leakage isolate from the colon lumen completely, rinsing the peritoneal cavity with a large number of warm salt water, careful hemostasis, inventorying gauzes equipment, exhausting pneumoperitoneum, removing equipment, closing the incision, fixing the aeration duct. After 1 hour, anesthesia disappeared, giving the pig drinking water and no residue liquid diet after fasting 24 hours.
Control group of traditional surgery:anesthesia, fixing position, disinfection, posing towels, opening the abdomen from the original incision,explorating perforation site, full-thickness sutures.making a round incision with diameter about 3.0cm at about 5.0cm front left side of the incision,opening the subcutaneous tissue, fascia with electroscalpel, bluntly separating the abdominal muscles, cutting peritoneum, taking out the suitable colon section through the stoma, making the plastic sticks and rubber ducts across the mesenteric vascular-free zone to fixed the colon on the abdominal wall, making a 3.0cm longitudinal incision on the edge of the colon wall with electroscalpel, suturing the cut colon wall onto the stoma. washing abdominal cavity, careful hemostasis, remaining peritoneal drainage duct, gauze inventorying equipment, closing the abdomen and ending surgery.
2.4 The third times surgery for detecting the effect of repairing
Detecting the effect of repairing colon perforation and AL in treatment and control groups at the 7th,14th,21st days after surgery,detecting intra-abdominal adhesions and stenosis at AL with laparoscope, detecting the bursting pressure at AL, and take healing tissue samples for detecting hydroxyproline contents and MVD.
2.4.1 Detecting bursting pressure at healed AL or perforation of the colon in vivo
Detecting the location of CSD, ligating the colon with a braided belt about 5.0cm at its proximal, exhausting gas in CSD, removing it, inserting connection duct of the precision pressure gauge through the anus into the rectum 15.0-20.0cm, suturing anus with 7.0 silk to ensure that no gas permeated and the part of perforation or anastomotic leak of the colon and the pressure gauge was the same level, slowly injecting gas into the colon, the assistant closely observed and recorded pressure gauge readings, When the colon perforation or anastomotic leak site began leak, the readings was the burst pressure.
Excising tissue at healed place of colon, divided them into 5 copies.1 copy was soaked in 10% formalin for HE staining, and the remaining 4 copies were stored in-80℃freezer, respectively for detecting hydroxyproline contents and MVD.
2.4.2 Detecting hydroxyproline content of healed tissue of AL or perforation
Alkaline lysis method of the hydroxyproline assay test box:Weighing wet weight tissue, accuratly adding alkaline lysis buffer, adjust PH value, adding activated carbon in the hydrolyzate,taking out supernatant,adding reagents,water bath,cooling, centrifuged,taking out supernatant,measureing absorbance,calculating hydroxyproline contents according to a formula.
2.4.3 Detecting the MVD at healed tissue of AL or perforation
Embedding tissue sections with OCT, fixing with acetone, regular hydration, PBS wash to remove endogenous peroxidase activity,blocking with goat serum,incubating at room temperature, tilting the serum, dropping working solution with the first antibody, washing with PBS,dropping biotinylated goat anti-mouse IgG, washing with PBS, dropping streptavidin labeled with horseradish, washing with PBS, configuration DAB working solution, dropping (DAB) color reagent, washing with tap water, restaining the organization with hematoxylin, washing with PBS, dehydration, dry glass slides were mounted with neutral resin, counting microvessel.
2.5 Statistical analysis
Statistical analysis with software package SPSS 13.0, measurement data displayed with mean±standard deviation (x±s), comparison between two groups with two independent sample t-test (bilateral) analysis, P<0.05 as significant difference.
Results
1 Testing sample performance
1.1 Comparing deformation pressure of outer capsule of CSD
The outer capsule deformation pressure values of CSD made of medical silastic and polyurethane were 30.42±0.46Kpa versus 30.73±0.29 Kpa,The results showed that medical polyurethane was slightly better than the medical silastic, but no statistical difference
1.2 Comparing deformation pressure of inner duct of CSD
The inner duct deformation pressure values of CSD made of medical silastic and polyurethane were 5.96±0.22 Kpa and>100 Kpa, the CSD made of medical silastic was not enough hard, easily lead to stenosis and closure.but the inner duct of CSD made of medical polyurethane do not deform When the pressure value is more than 100 Kpa. So the CSD made of medical polyurethane is better than that made of medical silastic.
1.3 Comparing leak pressure of CSD
The leakage gas pressure values of CSD made of medical silastic and polyurethane were 59.23±1.25 Kpa versus 60.01±1.45 Kpa, medical polyurethane was slightly better than the medical silastic, but no statistical difference.
2 Testing the healing parts of colon perforation
2.1 Measuring the bursting pressure after colon perforation repaired
The 7th,14th,21st days after surgery, bursting pressure in the treatment group and control group:26.20+2.76,36.12+1.67,38.58+1.46 versus 21.28+0.80,27.72±1.07,35.92+2.01. The bursting pressure in treatment groups was higher than that of the control group at 7th,14th,21st days after surgery (t=3.83,P=0.005;t=9.46,P <0.001;t=2.39,P=0.044)
2.2 Measuring the hydroxyproline contents in the healing site after colon perforation repaired
The 7th,14th,21st days after surgery, the hydroxyproline contents in the treatment group and control group:21.42±0.76,25.96±0.87,27.50±0.59 versus 15.14±0.97,21.08±1.08,26.90±0.56, It was higher in treatment group than that of the control group at 7th,14th days after surgery (t=11.39;t=7.82,P<0.001),no significant difference between 2 groups at 21st day (t=1.65,P=0.139)
2.3 Measuring the MVD in the healing site after colon perforation repaired
The 7th day after surgery,the MVD in the treatment group and control group:12.40+0.89 versus 10.40+1.14.It was higher in treatment group than that of the control group at the 7th after surgery (t=3.09,P=0.015)
3 Testing the healing parts of AL
3.1 Measuring the bursting pressure after colon AL repaired
The 7th,14th,21st days after surgery, the bursting pressure in the treatment group and control group:23.54+1.41,34.14±1.86,36.70±1.52 versus 19.10±0.57,25.96±1.17,34.00±2.15. The bursting pressure in treatment groups is higher than that of the control group at the 7th,14th days after surgery (t=6.54, P<0.001;t=8.31,P<0.001), No significant difference between 2 groups at 21st day(t=2.29,P=0.051).
3.2 Measuring the hydroxyproline contents in the healing site after colon AL repaired.
The 7th,14th,21st days after surgery,hydroxyproline contents in the treatment group and control group:20.46±0.76,25.08±1.07,26.10±0.61 versus 14.20±0.94, 20.22±1.22,26.70±0.67, It was higher in treatment group than that of the control group at 7th,14th days after surgery(t=0.41,P<0.001;t=0.82,P<0.001), no significant difference between 2 groups at 21st day(t=0.85, P=0.179).
3.3 Measuring the MVD in the healing site of AL repaired
The 7th day after surgery, MVD in the treatment group and control group:11.20±0.84 versus 8.80±0.84. It was higher in treatment group than that of the control group at 7th day after surgery (t=4.54,P=0.002).
Conclusion
1. The CSD made of polyurethane has simple structure, excellent performance, easy operation, safety, reliability,so has clinical value.
2. Repairing colon AL or perforation with the CSD in 1st stage is safe, reliable and effective.
The innovation
1 The CSD can reach every place of colorectal through the anus, repair colorectal perforation and AL with laparoscope in 1st stage.
2 Treatment effect of CSD is safe, reliable, and can effectively replace the traditional surgery-repair and stoma in 1st stage, put back the colon in 2nd stage.
结直肠穿孔发病率约为结直肠癌总数的3-8%,包括肿瘤在内的各种疾病自发性肠穿孔约为1.2-4.6%。自发性结直肠穿孔发生于乙状结肠及乙状结肠直肠交界处的系膜对侧缘>50%,原因与乙状结肠特殊生理解剖因素有关。自发性结肠穿孔多发于反应相对迟钝的60岁以上老年人,病史难于叙述清楚,加上结肠穿孔缺乏特异临床表现,早期正确诊断不足10%。由于结肠内容物含有大量细菌,穿孔确诊后已产生严重弥漫性腹膜炎,加上细菌毒素的大量吸收,形成感染抗体并导致多器官功能衰竭,故病死率可高达到35-47%。
各种方式的结直肠吻合术后的吻合口漏(Anastigmatic Leakage,AL)发生率大约为3-9%,TME手术AL发生率2-19%,结肠AL病死率12.9%。
也就是说我国每年需进行结直肠穿孔或AL修补的病例相当庞大。而对于上述病人的传统治疗原则上均采取肠穿孔或AL全层间断缝合浆肌层加固,同时行近段结肠腹壁造瘘,2-3个月后二期再次行造瘘肠管回纳术(肿瘤除外)。这种处理方式病程长,痛苦大,特别是腹壁造瘘留下巨大的疤疤,两次手术易导致严重肠粘连并发症以及高额费用,给患者带来极大的心理压力和躯体痛苦。如何才能安全有效地简化此类疾病的治疗流程,缩短病程,降低费用,寻求到一种微创,美观,并发症低的手术治疗方式呢?目前许多学者特别是胃肠外科医务工作者正不竭努力地进行研究。特别是近年来随着以腹腔镜为主体的微创胃肠外科的发展及各种可吸收人工生物材料的临床应用增多,使此项研究工作的进展逐渐加快。正是在这种背景下,本实验设计出结肠液囊管(colon sac duct,CSD)来有效地解决需要解决的问题。并对此项技术进行了认真科学的研究,期望这一研究能在结直肠穿孔及AL的治疗领域有所突破。
研究目的：
1总结既往外科临床对于结直肠穿孔及结直肠AL成功进行各种手术修复的经验,比较传统手术成功的关键因素及存在的不足,结合人体结直肠具有肛门这一个通向体外的自然通道,通过实践研究设计并制作出一种能经肛门进出,可顺利到达人体结直肠各段,并能在腹腔镜协助下对结直肠穿孔及结直肠AL进行有效一期修复的医疗器械-CSD。并对CSD各项性能进行客观检测,以求证其用于实验及临床的安全性,可靠性,有效性。
2通过对实验用西藏小型猪进行动物实验研究CSD能否作为一种安全可靠的新技术对结直肠穿孔及结直肠AL进行有效的一期修复。特别是能否在结直肠穿孔或结直肠AL48-72小时以上,严重感染的腹腔环境下有效修复结直肠穿孔及结直肠AL。以传统外科手术作为对照,进一步求证CSD作为一种新技术的科学性,安全性,可靠性,有效性及微创优势。
研究内容与方法
1 CSD的样品制作
1.1制定预设方案：制定三种预设方案,查阅文献,预实验中对大约35kg的实验用西藏小型猪进行解剖,了解它的结直肠肠腔内径大小。作为CSD尺寸设计的依据,制定出CSD三种预设方案,并绘制出工程制作线图及立体彩图。
1.2材料选择：在高分子材料工程师的帮助下,对临床上已广泛使用且被证实具有良好生物组织相容性的几种医用级高分子材料的弹性模量、断裂强度、断裂伸长率进行充分比较后选择出硅橡胶、聚氨脂用于CSD的样品制作。
1.3样品制作：寻找选择样品制作单位,根据本实验提供三种设计方案的立体彩图,工程制作线图,进行模具制作,根据需要反复修改模具,制作出三种预设方案的CSD样品。
1.4确定最终样品：CSD的结构如下：主要由外套囊(1)与内管(2)组成,其中外套囊两端有环形凹槽(3),用于固定及隔绝,在外套囊与内管之间形成一密闭的环形空间(4),利用热融合技术使二者粘合成一体。外套囊一端的充气管(5)与所述环形空间相通用来注射空气充盈外套囊。外套囊质地柔韧,内管质地坚硬,外套囊两端向内收缩,延伸至内层管壁。这种设计模型可用于结直肠穿孔或结直肠AL的一期修复手术。它既可阻挡肠腔内压力向穿孔或漏口处传递,又可使病变肠管与肠内容物完全隔绝。形成一个非常有效的肠腔内导流管装置,故有利于病变肠管清洁低压的环境下一期愈合,达到甚至超过传统二期手术的效果。
1.52.3样品性能检测
利用三通管将CSD的充气管、精密压力表进行连接,向液囊管内匀速充气,观察CSD外套囊、内管是否变形。凹槽是否外凸变浅,液囊管是否漏气或破裂,同时记录压力表所显示的压力读数。并进行统计学分析。对样品的性能作出科学客观的分析比较。
2 CSD修复结直肠穿孔及结直肠AL动物实验
2.1实验设计
处理组采用CSD对结肠穿孔及结直肠AL进行一期修补,以传统一期穿孔或漏口缝合近段结肠造瘘,二期结肠回纳手术作为对照组,每组15例。比较两种手术方式的手术时间、术后腹腔粘连程度、安全性、可靠性、有效性。并在修补后7、14、21天检测穿孔及漏口的愈合情况：比较结直肠穿孔及漏口部位的愈合强度——爆破压,结肠愈合部位组织中微血管密度(microvessel density,MVD)及羟脯氨酸含量,评估CSD一期修补结直肠穿孔及结直肠AL的实用性、可靠性、安全性及微创效果。
2.2动物模型制造
电视腹腔镜监视下经肛门制作动物模型,采用实验用西藏小型猪,体重30-35kg。氯胺酮联合戊巴比妥麻醉。麻醉前半小时称体重,臀部肌肉内注射鲁米那钠100mg、阿托品0.5mg。麻醉诱导按氯胺酮6mg/kg耳根部注射给药进行,待实验猪安静少动时经行气管插管(7.0-7.5号)并固定,呼吸机辅助呼吸。手术过程中以氯胺酮,戊巴比妥或异丙酚静脉滴注维持。
连接各线路及管道,开启调制电视腹腔镜的冷光源,CO2气腹机、高频电刀,调节腹腔镜视管的黑白及清晰度。于肚脐右上缘切约11mm皮肤皮下切口,常规建立气腹,当腹内压达到10-12mmHg状态时。以10mmTrocar穿刺成功后,置入腹腔镜视管。观察腹腔内各脏器,排除腹腔内异常情况后,从肛门置入另一视管(代替肠镜)。在腹腔镜监视下进入到距肛门约25-30cm处于肠系膜对侧缘刺破肠壁,形成大约1.0×1.2cm2穿孔,制造结肠穿孔模型。或利用举肠器有效显露距肛门约25-30cm处结肠,以电凝勾切断肠管,以丝线缝合肠管两断端,留取大约三分之一的肠系膜对侧缘肠壁不缝合,制造结肠AL模型。
2.3应用CSD对结肠穿孔或AL进行一期修复
第一次手术后48小时,再次手术,在腹腔镜下应用CSD对存在穿孔或漏口的结肠进行一期修复。麻醉、体位固定及消毒,铺无菌巾单同前述动物模型制造。于原脐上切口处拆除缝线,以弯钳逐步分离腹壁各层,进入腹腔,注意防止此处有肠管粘连而被破损伤,直视下,将10mmTrocar置入腹腔内,并置入10mm腹腔镜视管,仔细观察腹腔内粘连情况,避免损伤粘连肠管。在腹腔镜监视下于左右下腹合适位置,各切一长约5mm切口,并置入5mm Trocar后,置入相应的腹腔镜器械。轻柔推拉分离腹腔内粘连,清除腹腔内存留的粪便、脓苔寻找穿孔或漏口部位。如果寻找困难则以温盐水轻柔冲洗腹腔,吸净后仔细寻找。找到穿孔或漏口后,以四号丝线园针,纵行方向上缝吊,切取少许异常肠壁组织送病理。另一手术组成员以手指轻柔扩肛,逐渐扩张至2指宽后置入肛管,以大量稀释碘伏冲洗肠腔,至流出的冲洗液基本不带粪碴后使用石蜡油润滑肠腔及CSD,然后在腹腔镜监视下应用推送器将液囊管轻柔推送至肠穿孔或漏口部位,并使肠穿孔或漏口处位于液囊管的中部,缓慢均匀地对液囊管进行充气,致液囊管充分膨胀具有一定的韧性,又不致过分粗硬扩张压迫肠管,造成肠壁张力过大,影响病变部位肠管的血运。液囊管在肠腔内能具有一定的活动度为佳。在推送器内置入金属杆,使推送器灵活转变为类似于妇产科举宫器相仿的举肠器,通过举肠器的辅助使穿孔部位结肠及其系膜能上下左右摆动,并形成一定的张力,清楚地显露出结肠系膜的无血管区,利于手术者操作。手术者使用分离钳于液囊管两环形凹槽部位紧贴肠外壁穿过结肠系膜无血管区,不损伤肠壁血运情况下,用可吸收羊肠线适度捆扎肠管于液囊管上,将穿孔或漏口部位肠管与肠腔完全隔绝,大量温盐水冲洗腹腔。仔细止血,清点纱布器械,放尽腹腔积气,撤除器械,关闭各切口,固定液囊管的冲气管后结束手术。术后1小时,麻醉作用消失,给予正常饮水,术后禁食24小时,第2天开始给予无渣流质饮食。
对照组的传统手术：麻醉、体位固定、消毒、铺巾、从左侧经腹直肌切口进腹、探查穿孔部位,全层缝合。手术切口前端左侧旁开大约5.0cm处,将皮肤剪一直径约3.0cm的圆形切口,电刀切开皮下组织、肌筋膜、钝性分离腹肌,切开腹膜,选择合适的结肠段,经造瘘口提出到腹外,以塑胶棒及橡胶管穿过肠系膜无血管区,将肠管固定于腹外,电刀纵行切开对系膜缘肠壁约3.0cm,将切开的肠管壁缝合固定于造瘘口上。腹腔冲洗,仔细止血,留腹腔引流管,清点纱布器械,关腹结束手术。
2.4第三次手术对修复效果进行检测
分别于手术后第7天、14天、21天对处理组及对照组的西藏小型猪结肠穿孔或AL修复的效果进行检测,腹腔镜下检测的项目为腹腔内粘连情况、肠AL部位有无狭窄、开腹检测肠穿孔或AL愈合部位的爆破压,并取样检测愈合肠壁组织的羟脯氨酸含量及MVD。
2.4.1活体状态下结肠AL或穿孔愈合部位爆破压的测定
确定CSD位置,于其近端约5.Ocm处用编织带结扎结肠,放尽CSD内气体,取出CSD,将精密压力表连接管经肛门插入直肠15.0-20.0cm,用7.0号丝线荷包缝合肛门,确保不会渗气,使穿孔或AL处结肠管与精密压力表处于同一水平,缓慢匀速地向肠腔内打气,助手密切观察并记录穿孔或AL结肠修复部位漏气时压力表的读数,即为结肠穿孔或AL修补部位的爆破压。
切取病变愈合部位肠壁组织,分为5份。1份浸泡在10%福尔马林中固定,进行HE染色,其余4份保存在-80℃冰箱中,分别用于检测病变愈合部位组织中羟脯氨酸含量及MVD。
2.4.2 AL或穿孔愈合处组织羧脯氨酸含量测定
使用碱水裂解法羟脯氨酸检测试盒：精确称取湿重组织,准确加碱水裂解液,调节PH值,在水解液中加入活性炭,取上清液,依次加入试剂,水浴,冷却,离心,取上清液,测吸光度,按公式计算样品中羟脯氨酸含量。
2.4.3 AL或穿孔愈合处组织MVD测定
组织OCT包埋切片,丙酮中固定,常规水化,PBS冲洗,消除内源性过氧化物酶的活性,PBS冲洗,山羊血清封闭室温孵育,倾去血清,滴加一抗工作液,PBS冲洗,滴加生物素标记山羊抗小鼠IgG, PBS冲洗,滴加辣根酶标记链霉卵白素,PBS冲洗,配置DAB工作液,滴加(DAB)显色剂显色,自来水充分轻柔冲洗,苏木素液一滴滴于组织上复染,PBS冲洗,脱水,玻片干燥后滴加中性树脂封片,微血管计数。
2.5统计学分析
应用SPSS13.0软件包统计分析,计量数据以均数±标准差(X±s夕表示,两组间比较采用两独立样本的t检验(双侧)分析,P<0.05为有显著性差异。
结果
1样品性能检测
1.1 CSD外套囊变形压力比较
医用硅橡胶与医用聚氨酯制作的CSD外套囊变形压力分别为：30.42+0.46Kpa与30.73±+0.29 Kpa。结果显示医用聚氨酯稍优于医用硅橡胶,但无统计学差别
1.2 CSD内管变形压力比较
医用硅橡胶制作的CSD内管变形压力值为5.96±0.22 Kpa, CSD的内管硬度不够,容易内突导致管腔狭窄闭合,而医用聚氨酯制作的CSD内管变形压力值大于100 Kpa时均无明显变形。故医用聚氨酯明显优于医用硅橡胶。
1.3 CSD漏气压力比较
用医用硅橡胶与医用聚氨酯制作的CSD漏气压力值为59.23±1.25Kpa与60.01±±1.45 Kpa,结果显示医用聚氨酯稍优于医用硅橡胶,但无统计学差别。
2结肠穿孔修复术后愈合部位检测
2.1结肠穿孔修复术后爆破压检测
术后第7、14、21天处理组与对照组爆破压分别为：26.20±+2.76、36.12±±1.67、38.58±1.46与2128±0.80、27.72±1.07、35.92±+2.01,处理组均优于对照组(t=3.83,P=0.005；t=9.46,P<0.001；t=2.39,P=0.044)。
2.2结肠穿孔愈合部位羟脯氨酸含量检测
术后第7、14、21天处理组与对照组羟脯氨酸含量分别为：21.42+0.76、25.96±±0.87、27.50±0.59与15.14+0.97、21.08±±1.08、26.90±0.56,术后第7、14天处理组羟脯氨酸含量较对照组高(t=11.39；t=7.82,P<0.001),术后第21天2组无显著差别(t=1.65,尸=0.139)。
2.3结肠穿孔愈合部位MVD检测
术后第7天处理组与对照组MVD分别为：12.40±0.89与10.40±±1.14,术后第7天MVD处理组高于对照组(t=3.09,P=0.015)。
3结肠AL修复术后愈合部位检测
3.1结肠AL修复术后爆破压检测
术后第7、14、21天处理组与对照组爆破压分别为：23.54±1.41、34.14+1.86、36.70±+1.52与19.10±0.57、25.96±1.17、34.00±2.15,术后第7、14天处理组均优于对照组(t=6.54,P<0.001；t=8.31,P<0.001),术后第21天2组无显著差别(t=2.29,P=0.051)。
3.2 AL愈合部位羟脯氨酸含量检测
术后第7、14、21天处理组与对照组羟脯氨酸含量分别为：20.46±±0.76、25.08±±1.07、26.10+0.61与14.20±+0.94、20.22±±1.22、26.70±0.67,术后第7、14天处理组羟脯氨酸含量较对照组高(t=0.41,P<0.001；t=0.82,P<0.001),术后第21天2组无显著差别(t=0.85,P=0.179)
3.3 AL愈合部位MVD检测
术后第7天处理组与对照组MVD分别为：11.20+0.84与8.80+0.84,第7天MVD处理组高于对照组(t=4.54,P=0.002)。
结论
1.采用聚氨酯制作的CSD结构简单、性能优良、操作方便、安全可靠,有临床应用价值。
2.应用CSD技术一期修复结肠AL或穿孔是安全、可靠且有效。
本研究创新之处
1自主研究设计的CSD能经肛门自由进出能到达人体结直肠各处,在腹腔镜下
对结直肠穿孔及AL进行一期修补。
2 CSD治疗效果安全、可靠,能有效取代传统一期修补造瘘,二期结肠回纳手术。
Background and objective
Incidence rate of colorectal perforation is about 3-8% of the total number of colorectal cancer, spontaneous colon perforation in all diseases is about 1.2-4.6% including various tumors. Spontaneous colon perforation occur at opposite mesentery edge of the sigmoid colon or the junction of the sigmoid colon and rectum,the rate is more than 50% due to special physiological and anatomical factors at sigmoid colon. Spontaneous perforation of the colon happens to mainly the 60 years old persons or elders,because the case history is difficult to describe clearly,and with the lack of specific clinical manifestations of colon perforation, early diagnosis is less than 10%. Since a large number of bacteria are in colon contents, serious diffuse peritonitis happens after diagnosis, a substantial absorption of bacterial toxins,the formation of infection antibodies and lead to multiple organ failure, so the mortality can reach 35-47%.
The rate of various forms colorectal anastomosis leakage(AL) is about 3-9%, TME is 2-19%, the mortality rate of AL is 12.9%, AL happens to mainly the elders and the mortality is about 35-47%.
The cases of colorectal perforation or AL repair are large each year in China. The traditional treatment for these patients is taking on the principle of full-thickness interruption suture and seromuscular layer strengthened,and the colostomy in proximal colon through abdominal wall,2-3 months later the colostomy is closed and returned back to the abdomen (except cancer).This approach is characteristic of longer duration, large pain, especially the huge scar left on the abdominal wall,the complications including colon adhesion,the high cost. tremendous psychological pressure and pain due to twice surgery. How to safely and effectively simplify the treatment of this disease, shortening the course, reducing costs and seeking to a minimally invasive, beautiful and low complications surgical treatment? In recent years, minimally invasive gastrocolon surgery with laparoscopy as the main body and clinical application of artificial absorption biological material are increasing. Therefore, colon sac duct (CSD)can effectively solve the problems we face.
Purpose:
1 Summarizing successful experience of the previous clinical surgery for repairing colorectal perforation and AL, analyzing traditional surgical critical successive factors and shortcomings,human anus which has a natural channel leading to out-of-body and practical researchs I designed and made medical equipment-CSD that can reach the any parts of the body and effectively repair colorectal perforation and AL through the anus,furthermore tested the performance of CSD objectively to verify safety, reliability and validity for experimental and clinical application.
2 Through experimental studies with Tibet mini-pig colon I verified that CSD can used as a safe and reliable new technology for repairing colorectal perforation and AL effectively,in particular when colorectal perforation or anastomosis leakage is more than 48 to 72 hours,there is severe abdominal infection. Traditional surgery as the control, and further verified if CSD has a new scientific technology, security, reliability, validity and minimally invasive advantage.
Contents and methods
1 CSD prototyping
1.1 Making the default program
Making three preset programs, reviewing literature in preliminary experiments. About 35kg Tibet mini pig was dissected to understand its colorectal lumen diameter size as basis of the colon sac design.making CSD three preset programs, and drawing projects map and three-dimensional color pictures.
1.2 Material selection
With the help of polymer material engineers, comparing elastic modulus, tensile strength, break elongation rate of several medical grade polymer material widely used in clinic and with good tissue compatibility, I selected the silastic, polyurethane for CSD samples at last.
1.3 Sample production
Looking for samples production units, according to our design that provide three-dimensional color pictures,engineering production line graph,We made the mold, repeatedly revised the mold as needed to produce the three preset CSD samples.
1.4 Determine the final sample
CSD structure is as follows up:mainly with the outer capsule (1) and inner duct (2),2 circular grooves at both ends of outer capsule (3) for fixing and isolateing, there is a closed annular space between the outer capsule and the inner duct (4), with hot fusion technology to bond the two into one capsule for filling capsule with saline/gas,The inflatable duct is at one side of the capsule (5) and communicates annular space. Outer capsule texture is flexibile, inner duct texture is hard, The outer capsule contracts at both ends, extending to the inner duct wall. This design model can be used for repairing colorectal perforation or AL in 1st stage, which can block the pressure of the perforation or leakage passing the wound, the wound can also completely isolated from the colon contents.These is good for healing.
1.5 sample performance test
The CSD,inflatable duct and precision pressure gauges were connected by three-way duct,then aerateed to observe if the CSD and inner duct were deformed,circular grooves were convex, sac leaked or broke,at the same time recording the pressure readings on pressure gauge.
2 CSD repairing colon perforation and AL in animal experiments
2.1 Experimental Design
Tibet mini-pigs were randomly divided into 2 groups, treatment group and control group,15 per group, repairing colon AL of treatment group with CSD, the control group with traditional suture and colostomy in a proximal colon in 1st stage,then put colostomy back in 2nd stage.Comparing operation time, postoperative abdominal adhesions,safety,reliability, validity.at the at 7th,14th,21st days later,detecting the anastomotic leak condition:Comparing of the healing of AL intensity:burst pressure, the tissue micro vessel density(MVD) and hydroxyproline contents in healing parts, assessing practicality, reliability, security, and minimally invasive effects of CSD repairing colorectal perforation and AL.
2.2 Making animal model
Animal models were made under the surveillance of laparoscopic transanal using experimental Tibet mini-pigs, weight 30-50kg. anesthesia wtih Ketamine combined with pentobarbital. Weighing before anesthesia, intramuscular injection luminal sodium 100mg, atropine 0.5mg at the hip. Anesthesia was induced by injection of ketamine 6mg/kg by ears, when the pig became quiet, endotracheal was dealed (No.7.0) and fixed. Maintaining pentobarbital or propofol by intravenous infusion during surgery.
Connecting lines and piping, opening the cold light, modulating laparoscope, CO2 pneumoperitoneum machines, high frequency electroscalpel, laparoscopic black and white and clarity adjustment. Conventional establishment of pneumoperitoneum at upper-right navel, when intra-abdominal pressure reached 10-12mmHg state, the 10mmTrocar puncture, laparoscopic vision duct were placed, observing the organs in the abdominal cavity to exclude intra-abdominal abnormalities. Inserting vision duct, from the anus,under the supervision of the laparoscope arriving about 25-30cm from the anus, at the opposite edge of mesenteric piercing the colon wall, formating about 1.0×1.2cm perforation, then colon perforation model was made. About 25-30cm from the anus at the colon, cutting off the colon with electric coagulation hook then suturing the two ends of colon with silk, but 1/3 circle colon was left for non-closure at contralateral margin of mesenterium, then colon AL model was made.
2.3 Application of CSD for the colon perforation or AL in repairing in 1st stage.
Re-operation at 48 hours after the first surgery, in the application of laparoscope repairing the leakage and perforation with CSD. Anesthesia, position and disinfection, sterile towel were the same foregoing. Through original suture incision separating the abdominal wall layers gradually with curved forceps, entering the abdominal cavity, there was attention to prevent colon injury due to adhesion, Inserting lmm Trocar into the abdominal cavity, and placing 10mm vison duct, carefully observing the intra-abdominal adhesions, avoid damaging colon. At proper position in the left and right lower abdomen under laparoscopic monitoring cutting a incision about 5mm, and implanting 5mm Trocar and the corresponding laparoscopic instruments. Separating intra-abdominal adhesions and clearing the pus and abdominal stool left gently, looking for perforation or leak. If you had difficulty in finding them, rinsing gently with warm water and sucking carefully. After finding perforation or leakage,longitudinally seaming with code 4 thread and circular needle, hanging, cutting a small abnormal tissue for pathology. Another members expanding the anal with fingers gently and gradually to 2 finger width, then inserting the anal speculum, washing the colon with a large number of diluted povidone-iodine,Lubricating CSD and colon lumen with paraffin oil, then under laparoscopic surveillance pushing the sac duct to the place of perforation or leakage with pushing device, making sure that the perforation or leakage was in center of the sac duct, inflating with aeration the sac duct to make it tough, but not too hard, avoiding expanding and oppressing bowel, causing too large bowel wall tension, effecting lesion revascularization. The sac duct in the colon lumen duct had a certain activity. Inserting metal rod into the pushing device, the device is similar to the lift-uterus device in obstetrics and gynecology, making perforation part of the colon and its mesentery swing up and down, and a certain tension with the auxiliary,clearly revealing the avascular zone, which will help surgery operation. With the separation clamp permeating the mesocolon avascular zone on the two circular groove location of the sac duct close the colon wall, not damage the colon blood flow, fixing colon in the sac duct moderately with absorbable catgut,making the parts of the colon perforation or leakage isolate from the colon lumen completely, rinsing the peritoneal cavity with a large number of warm salt water, careful hemostasis, inventorying gauzes equipment, exhausting pneumoperitoneum, removing equipment, closing the incision, fixing the aeration duct. After 1 hour, anesthesia disappeared, giving the pig drinking water and no residue liquid diet after fasting 24 hours.
Control group of traditional surgery:anesthesia, fixing position, disinfection, posing towels, opening the abdomen from the original incision,explorating perforation site, full-thickness sutures.making a round incision with diameter about 3.0cm at about 5.0cm front left side of the incision,opening the subcutaneous tissue, fascia with electroscalpel, bluntly separating the abdominal muscles, cutting peritoneum, taking out the suitable colon section through the stoma, making the plastic sticks and rubber ducts across the mesenteric vascular-free zone to fixed the colon on the abdominal wall, making a 3.0cm longitudinal incision on the edge of the colon wall with electroscalpel, suturing the cut colon wall onto the stoma. washing abdominal cavity, careful hemostasis, remaining peritoneal drainage duct, gauze inventorying equipment, closing the abdomen and ending surgery.
2.4 The third times surgery for detecting the effect of repairing
Detecting the effect of repairing colon perforation and AL in treatment and control groups at the 7th,14th,21st days after surgery,detecting intra-abdominal adhesions and stenosis at AL with laparoscope, detecting the bursting pressure at AL, and take healing tissue samples for detecting hydroxyproline contents and MVD.
2.4.1 Detecting bursting pressure at healed AL or perforation of the colon in vivo
Detecting the location of CSD, ligating the colon with a braided belt about 5.0cm at its proximal, exhausting gas in CSD, removing it, inserting connection duct of the precision pressure gauge through the anus into the rectum 15.0-20.0cm, suturing anus with 7.0 silk to ensure that no gas permeated and the part of perforation or anastomotic leak of the colon and the pressure gauge was the same level, slowly injecting gas into the colon, the assistant closely observed and recorded pressure gauge readings, When the colon perforation or anastomotic leak site began leak, the readings was the burst pressure.
Excising tissue at healed place of colon, divided them into 5 copies.1 copy was soaked in 10% formalin for HE staining, and the remaining 4 copies were stored in-80℃freezer, respectively for detecting hydroxyproline contents and MVD.
2.4.2 Detecting hydroxyproline content of healed tissue of AL or perforation
Alkaline lysis method of the hydroxyproline assay test box:Weighing wet weight tissue, accuratly adding alkaline lysis buffer, adjust PH value, adding activated carbon in the hydrolyzate,taking out supernatant,adding reagents,water bath,cooling, centrifuged,taking out supernatant,measureing absorbance,calculating hydroxyproline contents according to a formula.
2.4.3 Detecting the MVD at healed tissue of AL or perforation
Embedding tissue sections with OCT, fixing with acetone, regular hydration, PBS wash to remove endogenous peroxidase activity,blocking with goat serum,incubating at room temperature, tilting the serum, dropping working solution with the first antibody, washing with PBS,dropping biotinylated goat anti-mouse IgG, washing with PBS, dropping streptavidin labeled with horseradish, washing with PBS, configuration DAB working solution, dropping (DAB) color reagent, washing with tap water, restaining the organization with hematoxylin, washing with PBS, dehydration, dry glass slides were mounted with neutral resin, counting microvessel.
2.5 Statistical analysis
Statistical analysis with software package SPSS 13.0, measurement data displayed with mean±standard deviation (x±s), comparison between two groups with two independent sample t-test (bilateral) analysis, P<0.05 as significant difference.
Results
1 Testing sample performance
1.1 Comparing deformation pressure of outer capsule of CSD
The outer capsule deformation pressure values of CSD made of medical silastic and polyurethane were 30.42±0.46Kpa versus 30.73±0.29 Kpa,The results showed that medical polyurethane was slightly better than the medical silastic, but no statistical difference
1.2 Comparing deformation pressure of inner duct of CSD
The inner duct deformation pressure values of CSD made of medical silastic and polyurethane were 5.96±0.22 Kpa and>100 Kpa, the CSD made of medical silastic was not enough hard, easily lead to stenosis and closure.but the inner duct of CSD made of medical polyurethane do not deform When the pressure value is more than 100 Kpa. So the CSD made of medical polyurethane is better than that made of medical silastic.
1.3 Comparing leak pressure of CSD
The leakage gas pressure values of CSD made of medical silastic and polyurethane were 59.23±1.25 Kpa versus 60.01±1.45 Kpa, medical polyurethane was slightly better than the medical silastic, but no statistical difference.
2 Testing the healing parts of colon perforation
2.1 Measuring the bursting pressure after colon perforation repaired
The 7th,14th,21st days after surgery, bursting pressure in the treatment group and control group:26.20+2.76,36.12+1.67,38.58+1.46 versus 21.28+0.80,27.72±1.07,35.92+2.01. The bursting pressure in treatment groups was higher than that of the control group at 7th,14th,21st days after surgery (t=3.83,P=0.005;t=9.46,P <0.001;t=2.39,P=0.044)
2.2 Measuring the hydroxyproline contents in the healing site after colon perforation repaired
The 7th,14th,21st days after surgery, the hydroxyproline contents in the treatment group and control group:21.42±0.76,25.96±0.87,27.50±0.59 versus 15.14±0.97,21.08±1.08,26.90±0.56, It was higher in treatment group than that of the control group at 7th,14th days after surgery (t=11.39;t=7.82,P<0.001),no significant difference between 2 groups at 21st day (t=1.65,P=0.139)
2.3 Measuring the MVD in the healing site after colon perforation repaired
The 7th day after surgery,the MVD in the treatment group and control group:12.40+0.89 versus 10.40+1.14.It was higher in treatment group than that of the control group at the 7th after surgery (t=3.09,P=0.015)
3 Testing the healing parts of AL
3.1 Measuring the bursting pressure after colon AL repaired
The 7th,14th,21st days after surgery, the bursting pressure in the treatment group and control group:23.54+1.41,34.14±1.86,36.70±1.52 versus 19.10±0.57,25.96±1.17,34.00±2.15. The bursting pressure in treatment groups is higher than that of the control group at the 7th,14th days after surgery (t=6.54, P<0.001;t=8.31,P<0.001), No significant difference between 2 groups at 21st day(t=2.29,P=0.051).
3.2 Measuring the hydroxyproline contents in the healing site after colon AL repaired.
The 7th,14th,21st days after surgery,hydroxyproline contents in the treatment group and control group:20.46±0.76,25.08±1.07,26.10±0.61 versus 14.20±0.94, 20.22±1.22,26.70±0.67, It was higher in treatment group than that of the control group at 7th,14th days after surgery(t=0.41,P<0.001;t=0.82,P<0.001), no significant difference between 2 groups at 21st day(t=0.85, P=0.179).
3.3 Measuring the MVD in the healing site of AL repaired
The 7th day after surgery, MVD in the treatment group and control group:11.20±0.84 versus 8.80±0.84. It was higher in treatment group than that of the control group at 7th day after surgery (t=4.54,P=0.002).
Conclusion
1. The CSD made of polyurethane has simple structure, excellent performance, easy operation, safety, reliability,so has clinical value.
2. Repairing colon AL or perforation with the CSD in 1st stage is safe, reliable and effective.
The innovation
1 The CSD can reach every place of colorectal through the anus, repair colorectal perforation and AL with laparoscope in 1st stage.
2 Treatment effect of CSD is safe, reliable, and can effectively replace the traditional surgery-repair and stoma in 1st stage, put back the colon in 2nd stage.
引文
[1]Runkel N S, Schlag P, Schwarz V, et al. Outcome after emergency surgery for cancer of the large intestine[J]. Br J Surg,1991,78(2):183-188.
[2]汪建平,唐远志,董文广.结直肠癌并急性结肠梗阻的外科处理——附225例临床分析[J].中国胃肠外科杂志.,1999(2)：79-81.
[3]Laurent C, Nobili S, Rullier A, et al. Efforts to improve local control in rectal cancer compromise survival by the potential morbidity of optimal mesorectal excision[J]. J Am Coll Surg,2006,203(5):684-691.
[4]Mazeh H, Samet Y, Abu-Wasel B, et al. Application of a novel severity grading system for surgical complications after colorectal resection[J]. J Am Coll Surg,2009,208(3):355-361.
[5]Kanellos I, Vasiliadis K, Angelopoulos S, et al. Anastomotic leakage following anterior resection for rectal cancer[J]. Tech Coloproctol,2004,8 Suppl 1:s79-s81.
[6]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer [J]. Br J Surg,2005,92 (2):211-216.
[7]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses [J]. Surg Gynecol Obstet,1982,154 (5):657-661.
[8]Wullstein C, Koppen M, Gross E. Laparoscopic treatment of colonic perforations related to colonoscopy[J]. Surg Endosc,1999,13(5):484-487.
[9]喻德洪.现代肛肠外科学[M].北京：人民军医出版社,1997.
[10]Chambers W M, Mortensen N J. Postoperative leakage and abscess formation after colorectal surgery [J]. Best Pract Res Clin Gastroenterol,2004,18 (5):865-880.
[11]Law W L, Chu K W. Anterior resection for rectal cancer with mesorectal excision:a prospective evaluation of 622 patients[J]. Ann Surg,2004,240 (2):260-268.
[12]Alves A, Panis Y, Mathieu P, et al. Mortality and morbidity after surgery of mid and low rectal cancer. Results of a French prospective multicentric study[J]. Gastroenterol Clin Biol,2005,29(5):509-514.
[13]Buchs N C, Gervaz P, Secic M, et al. Incidence, consequences, and risk factors for anastomotic dehiscence after colorectal surgery:a prospective monocentric study[J]. Int J Colorectal Dis,2008,23(3):265-270.
[1]Runkel N S, Schlag P, Schwarz V, et al. Outcome after emergency surgery for cancer of the large intestine[J]. Br J Surg,1991,78(2):183-188.
[2]汪建平,唐远志,董文广.结直肠癌并急性结肠梗阻的外科处理——附225例临床分析[J].中国胃肠外科杂志,1999(2)：79-81.
[3]Wullstein C, Koppen M, Gross E. Laparoscopic treatment of colonic perforations related to colonoscopy[J]. Surg Endosc,1999,13(5):484-487.
[4]Carpio G, Albu E, Gumbs M A, et al. Management of colonic perforation after colonoscopy. Report of three cases[J]. Dis Colon Rectum,1989,32(7):624-626.
[5]Anderson M L, Pasha T M, Leighton J A. Endoscopic perforation of the colon: lessons from a 10-year study[J]. Am J Gastroenterol,2000,95(12):3418-3422.
[6]Thompson S K, Chang E Y, Jobe B A. Clinical review:Healing in gastrointestinal anastomoses, part I[J]. Microsurgery,2006,26(3):131-136.
[7]Christensen H, Langfelt S, Laurberg S. Bursting strength of experimental colonic anastomoses. A methodological study[J]. Eur Surg Res,1993,25 (1):38-45.
[8]王一帆.可降解内支架结肠吻合技术及其在特殊腹腔环境中应用的研究[D].杭州：浙江大学,2008.
[9]Hendriks T, Mastboom W J. Healing of experimental intestinal anastomoses. Parameters for repair[J]. Dis Colon Rectum,1990,33(10):891-901.
[10]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses[J]. Surg Gynecol Obstet,1982, 154(5):657-661.
[1]Laurent C, Nobili S, Rullier A, et al. Efforts to improve local control in rectal cancer compromise survival by the potential morbidity of optimal mesorectal excision[J]. J Am Coll Surg,2006,203(5):684-691.
[2]Mazeh H, Samet Y, Abu-Wasel B, et al. Application of a novel severity grading system for surgical complications after colorectal resection[J]. J Am Coll Surg,2009,208(3):355-361.
[3]Kanellos I, Vasiliadis K, Angelopoulos S, et al. Anastomotic leakage following anterior resection for rectal cancer[J]. Tech Coloproctol,2004,8 Suppl 1:s79-s81.
[4]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer[J]. Br J Surg,2005,92 (2):211-216.
[5]Thompson S K, Chang E Y, Jobe B A. Clinical review:Healing in gastrointestinal anastomoses, part I[J]. Microsurgery,2006,26(3):131-136.
[6]Iqbal C W, Chun Y S, Farley D R. Colonoscopic perforations:a retrospective review[J]. J Gastrointest Surg,2005,9(9):1229-1235.
[7]Christensen H, Langfelt S, Laurberg S. Bursting strength of experimental colonic anastomoses. A methodological study[J]. Eur Surg Res,1993,25(1): 38-45.
[8]王一帆.可降解内支架结肠吻合技术及其在特殊腹腔环境中应用的研究[D].杭州：浙江大学,2008.
[9]Hendriks T, Mastboom W J. Healing of experimental intestinal anastomoses. Parameters for repair[J]. Dis Colon Rectum,1990,33(10):891-901.
[10]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses [J]. Surg Gynecol Obstet,1982,154 (5):657-661.
[1]Lipska M A, Bissett I P, Parry B R, et al. Anastomotic leakage after lower gastrointestinal anastomosis:men are at a higher risk[J]. ANZ J Surg,2006, 76(7):579-585.
[2]Luna-Perez P, Rodriguez-Ramirez S E, Gutierrez D L B M, et al. [Multivariate analysis of risk factors associated with dehiscence of colorectal anastomosis after anterior or lower anterior resection for sigmoid or rectal cancer] [J]. Rev Invest Clin,2002,54(6):501-508.
[3]赵广法,师英强,莫善兢.直肠全系膜切除术后吻合口漏的危险因素分析和对策[J].肿瘤,2004(6).
[4]李文明.直肠癌Dixon术后吻合口漏的影响因素与防治[D].济南：山东大学,2010.
[5]Vignali A, Fazio V W, Lavery I C, et al. Factors associated with the occurrence of leaks in stapled rectal anastomoses:a review of 1,014 patients[J]. J Am Coll Surg,1997,185(2):105-113.
[6]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer[J]. Br J Surg,2005,92 (2):211-216.
[7]Komen N, Dijk J W, Lalmahomed Z, et al. After-hours colorectal surgery:a risk factor for anastomotic leakage[J]. Int J Colorectal Dis,2009,24(7):789-795.
[8]Averbach A M, Chang D, Koslowe P, et al. Anastomotic leak after double-stapled low colorectal resection[J]. Dis Colon Rectum,1996,39 (7):780-787.
[9]王巍,续哲莉.中下段直肠癌保肛术153例临床分析[J].实用肿瘤学杂志,1995(1)：54-55.
[10]Carlsen E, Schlichting E, Guldvog I, et al. Effect of the introduction of total mesorectal excision for the treatment of rectal cancer[J]. Br J Surg,1998,85 (4):526-529.
[11]Enker W E, Stearns M J, Janov A J. Peranal coloanal anastomosis following low anterior resection for rectal carcinoma[J]. Dis Colon Rectum,1985,28 (8):576-581.
[12]余龙龙,陈震宏,陈栋.直肠癌前切除术后吻合口瘘的危险因素及对策[J]. 中国中西医结合外科杂志,2009(1)：29-31.
[13]Karliczek A, Benaron D A, Baas P C, et al. Intraoperative assessment of microperfusion with visible light spectroscopy in esophageal and colorectal anastomoses[J]. Eur Surg Res,2008,41(3):303-311.
[14]Ellebaek P M, Qvist N, Bisgaard C, et al. Peritoneal microdialysis. Early diagnosis of anastomotic leakage after low anterior resection for rectosigmoid cancer[J]. Scand J Surg,2009,98(3):148-154.
[15]Amrani L, Menard C, Berdah S, et al. From iatrogenic digestive perforation to complete anastomotic disunion:endoscopic stenting as a new concept of "stent-guided regeneration and re-epithelialization"[J]. Gastrointest Endosc, 2009,69(7):1282-1287.
[2]汪建平,唐远志,董文广.结直肠癌并急性结肠梗阻的外科处理——附225例临床分析[J].中国胃肠外科杂志.,1999(2)：79-81.
[3]Laurent C, Nobili S, Rullier A, et al. Efforts to improve local control in rectal cancer compromise survival by the potential morbidity of optimal mesorectal excision[J]. J Am Coll Surg,2006,203(5):684-691.
[4]Mazeh H, Samet Y, Abu-Wasel B, et al. Application of a novel severity grading system for surgical complications after colorectal resection[J]. J Am Coll Surg,2009,208(3):355-361.
[5]Kanellos I, Vasiliadis K, Angelopoulos S, et al. Anastomotic leakage following anterior resection for rectal cancer[J]. Tech Coloproctol,2004,8 Suppl 1:s79-s81.
[6]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer [J]. Br J Surg,2005,92 (2):211-216.
[7]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses [J]. Surg Gynecol Obstet,1982,154 (5):657-661.
[8]Wullstein C, Koppen M, Gross E. Laparoscopic treatment of colonic perforations related to colonoscopy[J]. Surg Endosc,1999,13(5):484-487.
[9]喻德洪.现代肛肠外科学[M].北京：人民军医出版社,1997.
[10]Chambers W M, Mortensen N J. Postoperative leakage and abscess formation after colorectal surgery [J]. Best Pract Res Clin Gastroenterol,2004,18 (5):865-880.
[11]Law W L, Chu K W. Anterior resection for rectal cancer with mesorectal excision:a prospective evaluation of 622 patients[J]. Ann Surg,2004,240 (2):260-268.
[12]Alves A, Panis Y, Mathieu P, et al. Mortality and morbidity after surgery of mid and low rectal cancer. Results of a French prospective multicentric study[J]. Gastroenterol Clin Biol,2005,29(5):509-514.
[13]Buchs N C, Gervaz P, Secic M, et al. Incidence, consequences, and risk factors for anastomotic dehiscence after colorectal surgery:a prospective monocentric study[J]. Int J Colorectal Dis,2008,23(3):265-270.
[1]Runkel N S, Schlag P, Schwarz V, et al. Outcome after emergency surgery for cancer of the large intestine[J]. Br J Surg,1991,78(2):183-188.
[2]汪建平,唐远志,董文广.结直肠癌并急性结肠梗阻的外科处理——附225例临床分析[J].中国胃肠外科杂志,1999(2)：79-81.
[3]Wullstein C, Koppen M, Gross E. Laparoscopic treatment of colonic perforations related to colonoscopy[J]. Surg Endosc,1999,13(5):484-487.
[4]Carpio G, Albu E, Gumbs M A, et al. Management of colonic perforation after colonoscopy. Report of three cases[J]. Dis Colon Rectum,1989,32(7):624-626.
[5]Anderson M L, Pasha T M, Leighton J A. Endoscopic perforation of the colon: lessons from a 10-year study[J]. Am J Gastroenterol,2000,95(12):3418-3422.
[6]Thompson S K, Chang E Y, Jobe B A. Clinical review:Healing in gastrointestinal anastomoses, part I[J]. Microsurgery,2006,26(3):131-136.
[7]Christensen H, Langfelt S, Laurberg S. Bursting strength of experimental colonic anastomoses. A methodological study[J]. Eur Surg Res,1993,25 (1):38-45.
[8]王一帆.可降解内支架结肠吻合技术及其在特殊腹腔环境中应用的研究[D].杭州：浙江大学,2008.
[9]Hendriks T, Mastboom W J. Healing of experimental intestinal anastomoses. Parameters for repair[J]. Dis Colon Rectum,1990,33(10):891-901.
[10]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses[J]. Surg Gynecol Obstet,1982, 154(5):657-661.
[1]Laurent C, Nobili S, Rullier A, et al. Efforts to improve local control in rectal cancer compromise survival by the potential morbidity of optimal mesorectal excision[J]. J Am Coll Surg,2006,203(5):684-691.
[2]Mazeh H, Samet Y, Abu-Wasel B, et al. Application of a novel severity grading system for surgical complications after colorectal resection[J]. J Am Coll Surg,2009,208(3):355-361.
[3]Kanellos I, Vasiliadis K, Angelopoulos S, et al. Anastomotic leakage following anterior resection for rectal cancer[J]. Tech Coloproctol,2004,8 Suppl 1:s79-s81.
[4]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer[J]. Br J Surg,2005,92 (2):211-216.
[5]Thompson S K, Chang E Y, Jobe B A. Clinical review:Healing in gastrointestinal anastomoses, part I[J]. Microsurgery,2006,26(3):131-136.
[6]Iqbal C W, Chun Y S, Farley D R. Colonoscopic perforations:a retrospective review[J]. J Gastrointest Surg,2005,9(9):1229-1235.
[7]Christensen H, Langfelt S, Laurberg S. Bursting strength of experimental colonic anastomoses. A methodological study[J]. Eur Surg Res,1993,25(1): 38-45.
[8]王一帆.可降解内支架结肠吻合技术及其在特殊腹腔环境中应用的研究[D].杭州：浙江大学,2008.
[9]Hendriks T, Mastboom W J. Healing of experimental intestinal anastomoses. Parameters for repair[J]. Dis Colon Rectum,1990,33(10):891-901.
[10]Shikata J, Shida T, Satoh S, et al. The effect of local blood flow on the healing of experimental intestinal anastomoses [J]. Surg Gynecol Obstet,1982,154 (5):657-661.
[1]Lipska M A, Bissett I P, Parry B R, et al. Anastomotic leakage after lower gastrointestinal anastomosis:men are at a higher risk[J]. ANZ J Surg,2006, 76(7):579-585.
[2]Luna-Perez P, Rodriguez-Ramirez S E, Gutierrez D L B M, et al. [Multivariate analysis of risk factors associated with dehiscence of colorectal anastomosis after anterior or lower anterior resection for sigmoid or rectal cancer] [J]. Rev Invest Clin,2002,54(6):501-508.
[3]赵广法,师英强,莫善兢.直肠全系膜切除术后吻合口漏的危险因素分析和对策[J].肿瘤,2004(6).
[4]李文明.直肠癌Dixon术后吻合口漏的影响因素与防治[D].济南：山东大学,2010.
[5]Vignali A, Fazio V W, Lavery I C, et al. Factors associated with the occurrence of leaks in stapled rectal anastomoses:a review of 1,014 patients[J]. J Am Coll Surg,1997,185(2):105-113.
[6]Peeters K C, Tollenaar R A, Marijnen C A, et al. Risk factors for anastomotic failure after total mesorectal excision of rectal cancer[J]. Br J Surg,2005,92 (2):211-216.
[7]Komen N, Dijk J W, Lalmahomed Z, et al. After-hours colorectal surgery:a risk factor for anastomotic leakage[J]. Int J Colorectal Dis,2009,24(7):789-795.
[8]Averbach A M, Chang D, Koslowe P, et al. Anastomotic leak after double-stapled low colorectal resection[J]. Dis Colon Rectum,1996,39 (7):780-787.
[9]王巍,续哲莉.中下段直肠癌保肛术153例临床分析[J].实用肿瘤学杂志,1995(1)：54-55.
[10]Carlsen E, Schlichting E, Guldvog I, et al. Effect of the introduction of total mesorectal excision for the treatment of rectal cancer[J]. Br J Surg,1998,85 (4):526-529.
[11]Enker W E, Stearns M J, Janov A J. Peranal coloanal anastomosis following low anterior resection for rectal carcinoma[J]. Dis Colon Rectum,1985,28 (8):576-581.
[12]余龙龙,陈震宏,陈栋.直肠癌前切除术后吻合口瘘的危险因素及对策[J]. 中国中西医结合外科杂志,2009(1)：29-31.
[13]Karliczek A, Benaron D A, Baas P C, et al. Intraoperative assessment of microperfusion with visible light spectroscopy in esophageal and colorectal anastomoses[J]. Eur Surg Res,2008,41(3):303-311.
[14]Ellebaek P M, Qvist N, Bisgaard C, et al. Peritoneal microdialysis. Early diagnosis of anastomotic leakage after low anterior resection for rectosigmoid cancer[J]. Scand J Surg,2009,98(3):148-154.
[15]Amrani L, Menard C, Berdah S, et al. From iatrogenic digestive perforation to complete anastomotic disunion:endoscopic stenting as a new concept of "stent-guided regeneration and re-epithelialization"[J]. Gastrointest Endosc, 2009,69(7):1282-1287.