多聚酶链反应检测细菌DNA对大鼠空、回肠吻合口瘘的早期诊断价值

目的:探讨PCR检测大鼠外周血及腹水中细菌DNA对空肠-空肠、回肠-回肠吻合口瘘的早期诊断价值。方法:健康Wistar雌性大鼠50只，随机分成5组，每组10只：A组为假手术组;B组为空肠-空肠吻合组;C组为空肠吻合口瘘组;D组为回肠-回肠吻合组;E组为回肠吻合口瘘组。采集手术前后外周血及术后腹水，抽提DNA, 比较lacZ基因和16SrRNA基因的PCR阳性率，并观察各组的病理学情况。结果:（1）C，E组术后外周血lacZ基因PCR阳性率与B，D组无显著性差异（P>0.05）;C，E组术后外周血16SrRNA基因PCR阳性率显著高于B，D组（P<0.05）。（2）C，E组腹水lacZ基因和16SrRNA基因PCR阳性率均显著高于B，D组（P<0.05）。（3）C，E组腹水lacZ基因阳性率显著高于外周血（P<0.05）;C，E组腹水16SrRNA基因阳性率与外周血无显著性差异（P>0.05）。结论:（1）PCR检测术后外周血16SrRNA基因对空、回肠吻合口瘘的早期诊断有一定意义;（2）检测术后腹水lacZ基因和16SrRNA基因对空肠-空肠、回肠-回肠吻合口瘘的早期诊断也有一定意义。

Predictable value of PCR in detecting bacterial DNAs for early diagnosis of jejunal anastomotic leakage and ileal anastomotic leakage in rats

Abstract:Objective:To assess the value of detecting bacterial DNA in rats′ blood with PCR technique for early diagnosis of jejunojejunal anastomotic leakage and ileaoileal anastomotic leakage.Methods :Fifty healthy female Wistar rats were randomly divided into five groups:Group A(n=10), sham operation group; Group B(n=10), jejunojejunal anastomosis group; Group C(n=10), jejunojejunal anastomotic leakage group; Group D(n=10) ileaoileal anastomosis group, Group E(n=10), ileaoileal anastomotic leakage group.Group B and D rats had a complete anastomosis (end-to-end single layer anastomoses with 0# silk sutures).Group C and E rats had an anastomosis with a 5mm opening in intestinal anastomosis anterior wall. Group B and C rats had 3 cm jejunum resection at 15 cm from the Treitz ligament. Group D and E rats had 3 cm ileum resection at 15 cm proximal to the ileocecal junction. Pre-and post-operative venous blood, and postoperative ascites samples were collected. DNAs was extracted from these blood and ascites samples, and PCR techniques were used to amplify lacZ genes and 16S ribosomal RNA genes(16SrRNA genes).Results:(1) Comparing Group B with Group C, or Group D with Group E,there was no difference in positive ratio of lacZ genes in peripheral blood (PB) (P>0.05), but the positive ratio of 16SrRNA genes expression in PB in Group C and Group E was significantly higher than that in group B and Group D respectively (P<0.05). (2) In ascites samples,the positive expressing ratios of lacZ genes and 16SrRNA genes in Group C and Group E were both higher than in Group B and Group D(P<0.05); In jejunal anastomotic leakage and ileal anastomotic leakage groups, the positive ratios of lacZ genes in ascites were higher than the positive ratios in PB(P<0.05); but there were no significant differences between the the positive ratios of 16SrRNA genes in ascites and in PB (P>0.05).Conclusions:(1) Detecting 16SrRNA genes from PB with PCR has certain significance for early diagnosis of jejunal anastomotic leakage and ileal anastomotic leakage, (2) PCR might be a useful tool for early diagnosis of jejunal anastomotic leakages and ileal anastomotic leakages by detecting lacZ genes or 16SrRNA genes from ascites.