Department of Surgery, University of Chicago, Illinois, USA.
PLoS One. 2012;7(8):e44326. doi: 10.1371/journal.pone.0044326. Epub 2012 Aug 31.
The most feared complication following intestinal resection is anastomotic leakage. In high risk areas (esophagus/rectum) where neoadjuvant chemoradiation is used, the incidence of anastomotic leaks remains unacceptably high (≈ 10%) even when performed by specialist surgeons in high volume centers. The aims of this study were to test the hypothesis that anastomotic leakage develops when pathogens colonizing anastomotic sites become in vivo transformed to express a tissue destroying phenotype. We developed a novel model of anastomotic leak in which rats were exposed to pre-operative radiation as in cancer surgery, underwent distal colon resection and then were intestinally inoculated with Pseudomonas aeruginosa, a common colonizer of the radiated intestine. Results demonstrated that intestinal tissues exposed to preoperative radiation developed a significant incidence of anastomotic leak (>60%; p<0.01) when colonized by P. aeruginosa compared to radiated tissues alone (0%). Phenotype analysis comparing the original inoculating strain (MPAO1- termed P1) and the strain retrieved from leaking anastomotic tissues (termed P2) demonstrated that P2 was altered in pyocyanin production and displayed enhanced collagenase activity, high swarming motility, and a destructive phenotype against cultured intestinal epithelial cells (i.e. apoptosis, barrier function, cytolysis). Comparative genotype analysis between P1 and P2 revealed a single nucleotide polymorphism (SNP) mutation in the mexT gene that led to a stop codon resulting in a non-functional truncated protein. Replacement of the mutated mexT gene in P2 with mexT from the original parental strain P1 led to reversion of P2 to the P1 phenotype. No spontaneous transformation was detected during 20 passages in TSB media. Use of a novel virulence suppressing compound PEG/Pi prevented P. aeruginosa transformation to the tissue destructive phenotype and prevented anastomotic leak in rats. This work demonstrates that in vivo transformation of microbial pathogens to a tissue destroying phenotype may have important implications in the pathogenesis of anastomotic leak.
肠切除术后最令人恐惧的并发症是吻合口漏。在高危部位（食管/直肠），使用新辅助放化疗时，即使由高容量中心的专业外科医生进行手术，吻合口漏的发生率仍然高得不可接受（≈10%）。本研究的目的是检验以下假设，即在定植于吻合口部位的病原体在体内转化为表达组织破坏表型时，吻合口漏会发生。我们开发了一种新的吻合口漏模型，其中大鼠接受术前放疗，如癌症手术中那样，然后进行远端结肠切除，然后用铜绿假单胞菌进行肠内接种，铜绿假单胞菌是辐射肠道的常见定植菌。结果表明，与单独接受辐射的组织相比，暴露于术前辐射的肠组织在被铜绿假单胞菌定植时吻合口漏的发生率显著增加（>60%；p<0.01）。对原始接种株（MPAO1-称为 P1）和从漏吻合口组织中回收的菌株（称为 P2）进行表型分析表明，P2 在绿脓菌素产生和胶原酶活性增强、高群集运动以及对培养的肠上皮细胞（即凋亡、屏障功能、细胞溶解）的破坏表型方面发生了改变。对 P1 和 P2 之间的比较基因型分析发现 mexT 基因中的单个核苷酸多态性（SNP）突变导致无功能的截短蛋白的终止密码子。将 P2 中的突变 mexT 基因替换为原始亲本株 P1 中的 mexT 基因，使 P2 恢复为 P1 表型。在 TSB 培养基中未检测到 20 代自发转化。使用新型毒力抑制化合物 PEG/Pi 可防止铜绿假单胞菌向组织破坏性表型转化，并可防止大鼠吻合口漏。这项工作表明，体内微生物病原体向组织破坏性表型的转化可能对吻合口漏的发病机制有重要影响。
