重组融合多肽hEGF-AWRK6的纯化及对小鼠烫伤感染创面愈合的影响

Purification of recombinant fusion polypeptide hEGF-AWRK6 and effect on wound healing and infection of burn model mice

摘要:

旨在研究重组融合多肽hEGF-AWRK6（EK）对烫伤模型小鼠感染创面的治疗功效。采用大肠杆菌表达系统表达、纯化融合多肽EK，抑菌实验检测EK抑菌活性；构建小鼠II度烫伤和铜绿假单胞菌感染模型，实验组创面滴注EK（30 mg/L），以PBS、庆大霉素（30 mg/L）、烫伤膏（10 mg/L）为对照，计算烫伤后创面愈合率及菌落数；伤后10 d，取各组小鼠的伤口及周边皮肤进行HE染色及胶原蛋白的Western blotting检测，分析创面病理组织结构。实验结果纯化得到了具有抑菌活性的重组表达融合肽EK；伤后6 d始，EK组小鼠创面菌落数少于对照组，差异极显著（P<0.01）；EK组小鼠烫伤愈合率显著高于PBS对照组（P<0.01）；与对照组相比，EK组小鼠创面真皮层细胞排列规则，再上皮化较快，毛囊生长较多，Ⅰ型胶原蛋白表达显著增加。结果表明EK具有抑制小鼠烫伤创面感染、促创面愈合功效，具有开发成为治疗烧伤药物的潜力。

Abstract:

To test the therapeutic effect of recombinant fusion polypeptide hEGF-AWRK6 (EK) on burn infection of model mice. EK6 was expressed and purified with Escherichia coli expression system, and the Ⅱ degree burns and Pseudomonas aeruginosa infection model mouse were established. Experiment group was treated with EK (30 mg/L), and the control group was treated with PBS, gentamicin (30 mg/L), burn ointment (10 mg/L). The wound healing rate and colony count were calculated. Wound and surrounding skin were taken for HE staining and collagen western-blot analysis, and the wound pathological changes were observed after 10 days of drug delivery. The results showed that fusion peptide EK was successfully expressed and purified with significant antibacterial activities against Pseudomonas aeruginosa. Compared to the control group, the colony count (CFU) of the wound surface in EK mouse had a remarkable decrease (P<0.01) and healing rate had a significant increase in group EK6 (P<0.01). Pathological analysis result showed that compared to the control group, wound dermal cells in group EK arranged regularly, had more hair growth and a faster epithelization. These results indicated that the fusion peptide EK would be a good candidate for the drug development for the treatment of burning wounds.