改良NADPH-d和AchE酶组织化学法诊断先天性巨结肠  

Diagnosis of Hirschsprung's disease with modified NADPH-d and AchE enzyme histochemistry

作　　者：朱利斌[1] 张华[1] 林进汉[1] 戴丽娜[1] 黄璜[1] 李仲荣[1] 

机构地区：[1]温州医科大学附属第二医院小儿外科,浙江温州325027

出　　处：《温州医学院学报》2014年第1期31-34,38,共5页Journal of Wenzhou Medical College

基　　金：浙江省卫生厅科研基金资助项目(2009A149)

摘　　要：目的:探讨进一步改良后的还原型尼克酰胺腺嘌呤二核苷酸黄递酶(NADPH-d)和乙酰胆碱酯酶(AchE)酶组织化学染色法诊断先天性巨结肠(HD)的优势,以期提高HD术前及术中诊断的准确率。方法:取HD患儿术中切除的结肠标本18例,其中女7例,男11例,平均年龄(8.6±3)月。标本分狭窄段、移行段、扩张段,其冰冻切片分别用改良的NADPH-d和AchE酶组织化学染色法进行染色,与常规HE染色进行比较,并用肠神经元特异标记物β-III Tubulin(TUJ-1)行免疫荧光及蛋白基因产物9.5(protein gene product 9.5,PGP9.5)行免疫组化检查论证改良方法的特异性。结果:NADPH-d法能快速确定扩张段近端切缘是否存在神经节细胞,诊断HD的阳性率为94.4%(17/18);AchE法对于HD狭窄段黏膜下异常增生的神经丛诊断阳性率为88.9%(16/18);两组间诊断阳性率差异无统计学意义(P>0.05)。耗时方面NADPH-d法为(7.0±1.7)min,AchE法为(30.9±2.2)min,组间比较差异有统计学意义(P<0.001)。上述两种改良方法与TUJ-1及PGP9.5对照结果一致。结论:NADPH-d法能快速确定肠管切缘是否有神经节细胞分布,适用于术中快速诊断HD;AchE法适用于术前直肠黏膜层的活检诊断。Objective： To assess the superiorities of modified NADPH-diaphorase （NADPH-d） and acetyl-cholinesterase （AchE） enzyme histochemistry staining technique to diagnose Hirschsprung＇s disease （HD）. To increase the identification of ganglion cells in the boundary of the bowel in HD and decrease the risks of reoperation. Methods： Full-thickness colon samples of 18 children with HD （girl 7 cases, boy 11 cases ）, with average age （8.6 ± 3） months, were examined. Frozen sections at three levels （narrow segment, transitional segment and dilated segment） were performed and investigated to determine the extent of aganglionosis by modified NADPH- d and Ache staining respectively, which were compared by traditional HE staining. TUJ-1 （β -Ⅲ Tubulin） and PGP9.5（Protein Gene Product 9.5）, markers of enteric neural cells, were used for identification of positive myen- teric plexus and ganglion cells by immunofluorescence or immunohistochemistry. Results： NADPH-d staining could rapidly confirm whether the normal ganglion cells existed at the proximal end of the resected bowel. AchE technique clearly stained submucosal hypertrophic nerve trucks in aganglion segments. The specifica identifica- tions of HD, NADPH-d staining was 94.4% （17/18）, and Ache staining was 88.9% （16/18）, respectively （P〉 0.05）. Time comsuming was （7.0 ± 1.7） rain in NADPH-d, and （30.9 ± 2.2） rain in AchE, respectively （P〈0.01）. The results of two modified methods mentioned above were in keeping with TUG-landPGP 9.5. Conclusion： Modified NADPH-d staining can rapidly confirmed whether the ganglion ceils distribute at proximal end of resected bowel and it is suitable for rapidly intraoperative diagnosis of HD. AchE staining is the best technique to demonstrate hypertrophic nerve trunks in the submucosa and is suitable for diagnosing samples of suction rectal biopsy.

关 键 词：先天性巨结肠 诊断 酶组织化学 结直肠外科手术 

分 类 号：R726.1[医药卫生—儿科]