摘要
背景:在泌尿外科临床工作中,尤其是肾脏移植手术广泛开展以来,应用输尿管-膀胱再植吻合术重建尿液通路系统,已经成为经常运用的操作技术。然而,膀胱解剖位置深,手术显露差,输尿管-膀胱的手法吻合操作复杂而耗时,术后漏尿和狭窄等并发症的发生率高,虽然通过采用输尿管内放置支架管的方法能较好的预防术后组织水肿和吻合口漏尿形成的输尿管狭窄,但它同时也带来了置管的并发症,增加了泌尿系感染机会,影响了输尿管的蠕动功能,另外术后DJ管长期留置,管壁易形成结石,取出困难,同时管体韧性减退,脆性增加,取出时易造成DJ管断裂,并且术后有可能发生DJ管回缩、移位,导致取出困难,甚至需要二次手术取出。再者,安放的输尿管支架管需要术后进行膀胱镜下取管操作,无形之中增加了侵袭性操作的机会和风险,同时亦给患者带来了不必要的身心痛苦及经济负担。上述问题困扰着临床医生,因此,人们日益期盼一种简便,快捷,安全,无支架的新方法来完成输尿管-膀胱再植吻合。
目的:本项目拟应用钛轮钉吻合器行输尿管-膀胱粘膜机械吻合,并用钛夹关闭膀胱壁肌层建立抗反流隧道,以开辟尿路重建的新方式,评价及探讨钛轮钉吻合器行输尿管-膀胱粘膜机械吻合的安全性,可靠性,操作的难易程度和可行性,并与手工缝合法免双J管组及双J管置入组比较,评价其优势和劣势,探讨输尿管-膀胱机械吻合重建尿路的临床应用价值,为其能够应用于临床提供实验依据。
方法:应用钛轮钉吻合器遵循Lich-Gregoir技术,通过膀胱外途径行输尿管-膀胱粘膜再植术,以常规手工缝合输尿管-膀胱粘膜免置DJ管组及放置DJ管组做为对照组,每组9例。比较术中膀胱-输尿管粘膜吻合时间、抗反流隧道建立时间、一次吻合成功率、吻合口漏尿情况等,分别在术后12周通过彩色多普勒超声、逆行膀胱造影、大剂量静脉肾盂造影、Whitaker test实验、大体标本观察及HE染色观察吻合口愈合情况、各组间、各部位间组织学变化、术侧肾脏功能情况、上尿路尿动力学变化,尿路通畅情况和并发症发生情况,利用Masson染色观察吻合口部位及上尿路胶原纤维的分布变化,通过免疫组织化学染色及Western Blot技术进行各组间胶原蛋白(ColⅠ、ColⅢ)半定量分析并利用图像分析软件测定其表达水平,利用扫描电子显微镜观察吻合口粘膜覆盖情况及有无结石、结晶形成,评估该吻合技术的可行性及安全性。
结果:1、所有手术均顺利完成,Ⅰ组(钛轮钉机械吻合组)及Ⅱ、Ⅲ对照组中均没有发生术中死亡,吻合时间Ⅰ组明显少于Ⅱ、Ⅲ对照组(11.56±0.34 min versus18.11±1.06 min及22.78±0.49 min,P<0.01),手术过程中测量的各组间输尿管管径的平均值分别是Ⅰ组: 2.94±0.13mm (2.5-3.5mm之间);Ⅱ组:2.91±0.27mm(2.6-3.7mm之间);Ⅲ组: 3.12±0.17mm (2.5-3.6mm之间)。各组间输尿管管径相比无显著性差异。一次吻合成功率:钛轮钉组:88.9%;手工缝合免双J管组:100%;手工缝合置双J管组77.8%。
2、Ⅰ组及Ⅱ组中均没有发生术后观察期间死亡,Ⅲ组中有1例犬因泌尿系感染于术后20天死亡,其膀胱内滞留的DJ管表面有结石形成,其余犬只正常存活且膀胱留取的尿液细菌培养结果均呈阴性。术后12周彩色多普勒超声、大剂量静脉肾盂造影、逆行膀胱造影、Whitaker试验及大体标本观察结果显示:Ⅰ组所有实验犬吻合部位的钉环及钛夹位置良好,未发生移位,造影剂通过吻合口顺畅,肾脏、膀胱、输尿管形态良好,无输尿管梗阻及膀胱-输尿管返流发生,吻合口愈合良好,肾脏功能正常,肾盂内压力测试均在正常范围内,输尿管蠕动功能良好;Ⅱ组中有1例犬出现不完全性输尿管梗阻,狭窄部位在输尿管-膀胱吻合处;Ⅲ组有1例犬发生膀胱-输尿管返流。26例存活犬术后排尿正常,未观察到肉眼血尿发生,无切口感染、裂开,无尿囊肿形成及尿漏发生。HE染色光镜下观察结果显示各组间的组织学改变相似,均有轻度的输尿管及肾间质局部出血水肿及炎性细胞浸润,各组肾小球病变轻微,少数肾小球可见轻度充血,观察钛轮钉机械吻合组吻合口粘膜平坦,覆盖完整,对合情况良好,Ⅱ组中有2例犬吻合口管腔内粘膜略有凹凸不平,其中1例管腔较狭窄,Ⅱ组中剩余7例犬及Ⅲ组中所有犬只的吻合口管腔内粘膜与机械吻合组观察情况相似。观察钛环及钛夹周围组织结构与Ⅱ、Ⅲ组吻合口管壁厚度、粘膜炎症程度、纤维及肉芽组织增生情况无明显差异存在。Masson染色图片经IPP6.0图像分析软件分析,结果显示Ⅰ、Ⅱ、Ⅲ组间吻合口部位、上、下段输尿管、肾脏组织的胶原纤维的分布变化及平均光密度值无显著差异。应用免疫组化染色检测Ⅰ、Ⅱ、Ⅲ组间的输尿管膀胱吻合口、上、下段输尿管及肾脏组织的Ⅰ型、Ⅲ型胶原蛋白(ColⅠ、ColⅢ)表达,经图像分析技术进行半定量分析的结果与Masson染色一致。扫描电子显微镜观察钛轮钉机械吻合形成的新输尿管开口粘膜覆盖完整,管腔通畅,无结石、结晶形成。利用Western Blot技术,测定各组间胶原蛋白(ColⅠ、ColⅢ)的表达水平,经Alphalmager 2200图像分析软件测定各杂交带积分光密度值(IDV),对比其结果,显示各组间、各相应部位间无显著差异。
结论:应用钛轮钉机械吻合进行膀胱外途径的输尿管-膀胱再植术是安全可行的,该技术在保持输尿管开口畅通的情况下无需置放DJ管,且操作简便易行,缩短了手术所需时间,同时有可能会减少并发症的发生。
BACKGROUD AND OBJECTIVE
Ureteroneocystostomy is a frequently performed procedure to manage distal ureteralstenosis and vesicoureteral reflux. It is also a major method of surgical reconstruction ofurinary tract continuity in renal transplantation. Today, a variety of surgical techniques areavailable, among which the extravesicalureteroneocystostomytechnique (Lich-Gregoir methods) with their modifications, is probably the mostpopular. Despite improved surgical technique and immunosuppressive agents, urologicalcomplications still occur and account for significant morbidity and graft failure. The mostcommon urological complications are ureteral obstruction and leakage of theureteroneocystostomy. Although the use of a splint through the ureterocystostomy willresult in a significantly lower urological complication rate, routine splinting is known toincrease the number of urinary tract infections that can be graft and life-threatening.Meanwhile, dislodgement of the stent results in injury or obstruction, and removal of thisdevice requires an invasive procedure.Recently, we have utilized the titanium ring-pinstapler to reconstruct artery in clinical renal transplantation . We showed that the ring-pinsystem can be safely and easily applied to major artery reconstruction with a high patencyrates. The mechanical technique results in less bleed leakage and artery stenosis with aneverting anastomosis, providing a viable alternative to manual suturing. The purpose of thisexperiment was to compare ring-pin stapler with conventional sutures with routine uretericstenting and without routine ureteric stenting in vesicoureteral anastomosis, with specialfocus on anastomosis time, ureteral obstruction, urine leakage, vesicoureteral reflux, andstone formation during a follow-up period of 3 months. To our knowledge, this is the firstreport of a sutureless technique without routine ureteric stenting for extravesical ureteroneocystostomy.
MATERIALS AND METHODS:
Eighteen mongrel male dogs weighing 10 to 19 kg (mean, 16.5 kg) were used in our experiment. The animals were randomly divided into three groups of 9. Group I received non-sutured ureteroneocystostomy using the titanium ring pin stapler, and Group II received hand-sutured without routine ureteric stenting ureteroneocystostomy using Vicryl 5-0 adsorbable sutures, and GroupШreceived hand-sutured with routine ureteric stenting ureteroneocystostomy using 5-0 adsorbable Vicryl sutures. All experiments were performed according to our Institutional Guidelines on Animal Care and Use. The principles of laboratory animal care were followed.
Preoperative Management
Animals were not fed anything but water 24 h before the operation. The introduction of anesthesia was started via the intraperitoneal injection of the sterilized pentobarbital sodium (30mg/kg). The dog was then placed in supine position and intubated. An intravenous infusion of atropin (0.01 mg/kg) was performed, and the anesthesia was continued with intravenous administration of pentobarbital sodium. All dogs were subjected to color duplex ultrasonography to exclude congenital anomalies of urinary system before the start of the experiments. The vesical pressure was nullified by insertion of a urethral catheter (6-8 F). For infection prophylaxis, a dose of ceftriaxone sodium (2 g) was administered intravenously at the beginning of surgery.
Surgical Techniques
A midline incision was made to expose the urinary bladder. The random ureter was ligated and divided at its most distal part. The outer diameter of ureter was measured with a caliper. Mechanical dilation of the ureter end with a mosquito clamp was performed to make the ureter mouth larger when it was necessary. Before ureterovesical anastomsis, the distal ureter was spatulated and a 3 to 4cm long myotomy was made in the posterior-lateral bladder wall at a point 2 cm above the site of the normal ureterovesical junction.
Non-sutured ureteroneocystostomy
The anastomotic process of bladder mucosa and distal ureter with titanium ring pin staplers was performed . Firstly,the bladder mucosa in the distal edge of the myotomy was clawed with a crochet-hook and stretched up to form a conus about 1cm high. The mucosa conus was then passed through a titanium ring, which was mounted in a ring holder. Secondly, the circular cone was cut open and its edge was everted and impaled on the small pins. A similar procedure was performed on the ureter end. Thirdly, the two ring holders were brought together by passing the pins of the two rings through the everted edges of both of ureter and bladder mucosa to be included in the anastomosis with each other. The ring holders were then compressed with the crushing clamp to make the ureter and bladder mucosa unite tightly. Finally, the assembled rings together with the anastomosis were released as the ring holders were simultaneously unlocked. The anastomosis was secured by interlocking the small pins, which were bent by clamping when the two rings were brought together and compressed. The two edges of the muscle layer were grasped with tissue forceps and brought together. Three to four titanium clips were applied in a nearly horizontal direction to close the longitudinal line of the muscle layer over the implanted ureter. In this way, the distal ureter was embedded in a seromuscular vesical tunnel, thus creating the antireflux tunnel. In order to avoid ureter damage or occlusion by clips, the distal ureter was pressed down while the detrusor muscle layer was being closed with straight titanium clips(THICON ENDO-SURGERY, USA, LT300).
Hand-sutured without routine ureteric stenting ureteroneocystostomy
The technique has been previously described in detail by Lich and Gregoir. In brief, an incision of the bladder mucosa was made in the distal edge of the myotomy. Using interrupted 5-0 Vicryl suture, a direct anastomosis was carried out between the spatulated end of the ureter and the bladder mucosa. After that, the two longitudinal lines of muscle layer were closed togerther over the ureter with interrupted DJ 4-0 sutures to create the antireflux tunnel.
Hand-sutured with routine ureteric stenting ureteroneocystostomy
Same operational procedure has been done according to Lich and Gregoir technique above described, by way of addition, we have adopted the ureteral stenting technique correspondingly. The stent was a silastic urologic J–J stent which ranged in size from 5 to 7 Fr and in length from 12 to 14 cm. In order to removal this device postoperatively, we have the DJS to join the urethral catheter with 7 # silk suture during the operation Period. The stent was removed when the urinary bladder catheter was routinely removed approximately 2 weeks after surgery. After that, the two longitudinal lines of muscle layer were closed togerther over the ureter with interrupted DJ 4-0 sutures to create the antireflux tunnel. One of our experienced surgeons (Gang Ye) performed all suture-free procedure and another skillful surgeon (Feng Zhou) performed each hand-sutured anastomosis. Intraoperative data collection included the diameters of the ureter, ureterovesical anastomosis time, muscle layer closure time and total ureteroneocystostomy time. The absence of leakage of urine was confirmed by bladder filling. The laparotomy incision was closed in layers. The animals were allowed to recover and were then placed in standard animal care facilities. The urethral catheter (6-8 F) and DJS was left in for at least 2 weeks.
Postoperative Assessment and Follow-up
All animals were followed for 3 months and were monthly evaluated by color ultrasonography. When the follow-up was scheduled to end, all dogs were evaluated by ascending cystography and retrograde Cystography and the Whitaker test under general anesthesia. TheWhitaker test was utilized to study the intrapelvic pressure of the corresponding and the normal contralateral renal units. The urodynamic evaluation was carried out b insertion of a cannula (F5 ) into the renal pelvis. The cannula had a Y connection for infusion of warm saline (2.2 ml/min) and simultaneous recording of intrapelvic pressure profile. The recording system was DUET MULTI-P, (Medtronic,USA).
The animals were then sacrificed (still under anesthesia) using saturated potassium chloride. The urinary tract was excised and harvested. The bladder and ureter were opened without any damage of the new ureterovesical junction. The anastomosis sites were assessed for surface irregularities, bare clips or suture material or calculus formation under surgical microscope. The new ureterovesical junctions were catheterized with a ureteral catheter, and prepared for investigation with microscope. The specimens for histologic examination were fixed in formalin (the clips were removed by microdissection). The paraffin blocks were prepared, and multiple sections of the specimens stained with hematoxylin-eosin (H&E) for histologic examination. Collagen fibers observed by Masson staining in ureteral and renal tissue sections. Collagen I,Шin the upper and the lower ureteral and renal tissues was assessed by immunohistochemical method. The expression of Collagen I,Шin the upper and the lower ureter and the kidney was detected with Western Blot. The changes of the new ureter orifice open construction with the mechanical anastomosis were observed under the scanning electron microscope.
Statistical analysis
The data are presented as the mean±S.E.M. The paired t-test was used for statistical analysis with significance at the 0.05 level.
RESULTS
The ureterovesival anastomosis with the ring pin stapler was successfully completed in our studies. The antireflux tunnel construction with titanium clips also had a 100% technical success rate. The diameter of ureteral end included in the anastomosis was 2.94±0.13mm (range from 2.5-3.5mm), which showed no significant difference when compared with that in the other group. The 3.5 mm ring was applied in 2 cases, and 3mm ring in 4 and 2.5mm in 3 animals. The time required for extravesical ureteroneocystostomy in the three groups are summarized. Compared with manual suturing anastomosis, the suture-free technique with ring-pin stapler took a significantly shortest time. The time required for mechanical ureter reimplantation is only about 1/2 of that required for suturing anastomosis. All of the animals survived the surgical procedures and had an uneventful recovery, but one dog of groupШhad died of postpoertive infection. At 3-month follow-up, all ureters in Group I was patent with no sign of ureter dilation or hydronephrosis. However, 1 dog of Group II showed evidence of ureteral dilation with hydronephroses on color ultrasonography, which were subsequently confirmed by IVP studies. In our study, ascending cystography demonstrated that no vesicoureteral reflux was detected after ring-pin stapler, but one dog of suture groupШhad vesicoureteral reflux. The results of urodynamic evaluation showed that the baseline pressure of the pelviswas 1 to 2cm water. The intrapelvic pressure of the normal contralateral kidney ranged from 4 to 6 cm water (mean, 2.8 ,5.9 and 7.6 cm water for Group I , Group II and GroupШ, respectively). The intrapelvic pressure of the corresponding kidneys of the Group I and GroupШwas approximately normal. One case with hydronephrosis of group II had higher intrapelvic pressures of 21 cm water.
Necropsy examination revealed no signs of skin and urinary tract infection, urinary cyst, and fistulas occurred in any canines of Group I and Group II. One dog of GroupШwas revealed that it died of urinary tract infection by necropsy examination postoperative. All the dogs of the group I showed normal renoureteral units, while 1 dog of group II and1 dog of groupШshowed dilation of corresponding renoureteral units, which are in accordance with that finding of IVP and ascending cystography. The inner surface at the new ureterovesical junction was smooth without any bare pins in 9 specimens of the stapler group. The anastomosic site at the time of autopsy and histological examination showed the epithelial integrity overlying the staples of the new ureterovesical junction. In two specimens of Group II, the mucosal irregularities were seen at the anastomsis site, whereas in the remaining 7 specimens 9 specimens of GroupШ, the inner surface was smooth. There was no sign of calculus formation detected in any of the reimplantation sites of Group I and Group II. One dog of groupШwas detected calculus formation around of the dislodged and kinking distal DJS. Collagen fibers observed by Masson staining in ureteral and renal tissue sections. The expression of Collagen I,Шwas measured through immunohistochemical method, Western Blot in the upper and the lower ureter and the kidney of group I, II andШ. Compared with group II～Ш, the expression of Collagen I,Шof the upper and the lower ureteral and renal tissues was similar in group I (P>0.05). The scanning electron microscope showed the integral epithelial overlying the ureterovesical anastomotic site using the mechanical anastomosis without sign of calculus formation.
CONCLUSION
In conclusion, the suture-free technique with ring pin stapler is suitable for extravesical ureteroneocystostomy. This technique can maintain the new ureter orifice open, allowing good urine excrection and making use of stents unnecessary. Our technique is a quick and safe way to perform ureteroneocystostomy, and could result in a probably lower complication rate. Although this experiment demonstrates the safety, speed, and technical ease of extravesical ureteroneocystostomy using the titanium ring pin stapler system in canine’s ureteral reimplantation, there is no doubt that conventional suturing will continue to be the gold standard until futher data are available. Clearly, further investigations with a longer term of follow-up are needed to confirm the preliminary results.
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