负压封闭引流治疗小鼠创面铜绿假单胞菌感染的效果及机制

目的 观察VSD对感染创面中铜绿假单胞菌生长的影响,并探讨其可能机制。 方法 选取健康成年雄性C57BL/6小鼠40只,按照随机数字表法分为对照组和治疗组,每组20只。无菌条件下切除各小鼠背部1 cm×1 cm的全层皮肤,将细菌荧光素酶目的基因luxCDABE标记的野生型铜绿假单胞菌菌株PAO1 -lux涂抹于创面,包扎创面24 h,制成铜绿假单胞菌感染小鼠模型。治疗组小鼠创面行VSD治疗（负压为-16.625 kPa）,对照组创面常规换药。分别于治疗前和治疗24 h时,用小动物活体成像系统检测2组小鼠创面PAO1-lux荧光强度,激光多普勒血流成像仪检测创面血流量,以实时荧光定量RT-PCR检测创缘组织IL-1β、血管内皮生长因子(VEGF)的mRNA表达水平。观察治疗24 h时2组小鼠创缘组织病理学特点。对实验数据行t检验。 结果(1)治疗前,治疗组小鼠创面PAO1 -lux荧光强度与对照组相近(t=0.03,P=0.98);治疗24 h时,治疗组的荧光强度为(2.69±0.75)光子·秒-1·厘米-2·单位角度-1(photons· s-1- cm-2·sr-1),明显低于对照组的(5.18±0.96)photons·s-1·cm-2·sr-1,t =3.54,P=0.02。(2)治疗前,治疗组小鼠创面血流量与对照组相似(t =0.50,P=0.64);治疗24 h时,治疗组创面血流量为(96±9)灌注单位,明显高于对照组的(70±11)灌注单位,t=3.13,P=0.04。(3)治疗前,2组小鼠创缘皮肤组织中IL-1β、VEGF mRNA表达水平接近（t=0.19,P=0.86;t=0.07,P=0.95）;治疗24h时,治疗组IL-1β、VEGF mRNA表达水平分别为4.72±0.37、2.68±0.39,均明显高于对照组的2.24±0.50、1.22±0.13,t值分别为6.90、6.12,P值均为0.00。(4)治疗24 h时,治疗组创缘皮肤组织内炎性细胞浸润数量较对照组增加约77％。 结论 与常规换药相比,VSD治疗在小鼠全层皮肤缺损早期即能明显降低创面铜绿假单胞菌含量。其机制可能与增加创面局部血流量、提高创面组织炎性细胞数量、促进IL-1β和VEGF的mRNA表达有关。

Efficacy of vacuum sealing drainage in mice infected with Pseudomonas aeruginosa and its mechanism

Objective To observe the effect of vacuum sealing drainage (VSD) on the proliferation of Pseudomonas aeruginosa (PA) in infected wound, and to explore its possible mechanism. Methods Full-thickness skin wounds each with area of 1 cm × 1 cm were produced on the back of 40 C57 BL/6 mice,and then they were contaminated with wild type PA strains PAO1 marked with target gene of bacterial luciferase luxCDABE (PAO1-lux), they were dressed for 24 hours to reproduce PA infection model. Then mice were divided into experiment [E, with treatment of VSD ( pressure value at - 16. 625 kPa)] and control (C, with treatment of conventional dressing change) groups according to the random number table, with 20 mice in each group. The fluorescence intensity of PAO1-lux and blood flow in wound was respectively measured by in vivo optical imaging system and laser Doppler perfusion imager before treatment and at post treatment hour (PTH) 24. The expression levels of IL-1β and vascular endothelial growth factor (VEGF) mRNA in wound edge were determined by real-time fluorescence quantitative RT-PCR before treatment and at PTH 24. The specimens of wound edge tissue were collected for observation of pathological change at PTH 24.Data were processed with t test. Results There were no obvious difference in fluorescence intensity of PAO1-lux and blood flow in wound between E and C groups before treatment ( with t value respectively 0. 03,0.50, P values all above 0.05 ). The fluorescence intensity of PAO1-lux and blood flow in wound in E group at PTH 24 [(2.69 ±0.75) photons · s-1 · cm-2 · sr-1and (96 ±9) PU] was respectively lower and higher than that in C group [(5.18 ±0.96) photons · s-1 · cm-2 · sr-1 and (70 ± 11 ) PU, with t value respectively 3.54, 3.13, P values all below 0. 05]. The expression levels of IL-1 β and VEGF mRNA in both groups before treatment were similar ( with t value respectively 0.19, 0. 07, P values all above 0.05 ). The expression levels of IL-1β and VEGF mRNA in E group at PTH 24 was respectively 4.72 ± 0.37, 2.68 ± 0. 39, all markedly higher than those in C group ( 2.24 ± 0.50, 1.22 ± 0. 13, with t value respectively 6. 90, 6.12, P values all equal to 0.00). The number of inflammatory cell infiltrating the wound edge in E group at PTH 24 was increased by nearly 77％ as compared with that in C group. Conclusions Compared with conventional dressing change, VSD can reduce the amount of Pseudomonas aeruginosa in full-thickness skin defect wound at the early stage, it may be related with an increase in blood flow and number of inflammatory cells in wound tissue, promoting expression of IL-1β and VEGF mRNA.