Zhang T W, Wang Y P, Jia C Y
Department of Burns and Plastic Surgery, the Eighth Medical Center of PLA General Hospital, Beijing 100091, China.
Department of Burns and Plastic Surgery, Xiang'an Hospital of Xiamen University, Xiamen 361102, China.
Zhonghua Shao Shang Za Zhi. 2019 Feb 20;35(2):95-103. doi: 10.3760/cma.j.issn.1009-2587.2019.02.004.
To investigate the effects of basic fibroblast growth factor (bFGF) on healing of infective wound in New Zealand rabbit after debridement. Thirty-two New Zealand rabbits (3 to 4 months old, no matter male or female) were intradermally injected with 0.1 mL of complete Freund's adjuvant on the buttocks. Six weeks later, each rabbit was injected with 0.5 mL 5×10(7) colony forming unit/mL - on both sides of the back to reproduce the model of infective wound in New Zealand rabbit. After successful modeling, the 32 rabbits were divided into growth factor (GF) group, antituberculosis drug (AD) group, combined treatment (CT) group, and blank control (BC) group according to the random number table, with 8 rabbits in each group. After a complete debridement, the wounds of rabbits in group GF were treated with recombinant bovine bFGF gel (300 IU/cm(2,) about 0.45 g for each wound), the wounds of rabbits in group AD were covered with gauze which was impregnated with 6 mL isoniazid injection and 0.15 g rifampicin powder-injection, the wounds of rabbits in group CT were covered with gauze which was impregnated with isoniazid injection and rifampicin powder-injection after being treated with recombinant bovine bFGF gel as before, the wounds of rabbits in group BC were covered with sterile gauze, with dressing change of once every two days until the wounds were completely healed. Immediately after surgery and on post surgery day (PSD) 7, 14, 21, and 28, the wounds of rabbits in each group were observed with naked eyes and photos. On PSD 7, 14, 21, and 28, the wound healing rate was calculated and the complete healing time of wound was recorded. On PSD 7, 14, 21, and 28, the tissue samples of wound edge were collected for histomorphological observation with hematoxylin and eosin staining and Masson staining. On PSD 21, the number of microvessels was counted with immunohistochemical method. On PSD 7, 14, 21, and 28, the content of hydroxyproline in wound edge was determined by enzyme-linked immunosorbent assay. The numbers of samples of above-mentioned experiments were all 8. Data were processed with analysis of variance for repeated measurement, analysis of variance of factorial design, one-way analysis of variance, least significant difference test and Bonferroni correction. (1) The rabbits in four groups all survived to the end of experiment. Immediately after surgery, edema was observed in basal wounds of rabbits in the four groups. On PSD 7, the wounds of rabbits in the 4 groups were contracted with scabs and less edema. The wounds of rabbits in groups GF and CT became redder. On PSD 14, the wounds of rabbits in the 4 groups contracted obviously. There were no obvious exudates in wounds of rabbits in groups AD and CT, while 1 wound of rabbit in group GF and 2 wounds of rabbits in group BC became red and swelling with purulent exudates. On PSD 21, wounds of rabbits in groups GF and CT were basically healed, while 2 wounds of rabbits in group BC healed slowly with purulent secretion. On PSD 28, wounds of rabbits in the 4 groups were basically healed, while 2 wounds of rabbits in group BC hardly healed with redness and swelling. (2) From PSD 7 to 28, the wound healing rates of rabbits in groups GF, AD, and CT were significantly higher than those in group BC (<0.05). On PSD 14 and 21, the wound healing rates of rabbits in groups GF and CT were significantly higher than those in group AD (<0.05). From PSD 7 to 28, the wound healing rates of rabbits in group GF were close to those in group CT (>0.05). (3) The complete healing time of wounds of rabbits in groups GF, AD, and CT was significantly shorter than that in group BC (<0.05). The complete healing time of wounds of rabbits in groups GF and CT was significantly shorter than that in group AD (<0.05). The complete healing time of wounds of rabbits in group GF was close to that in group CT (>0.05). (4) On PSD 7, a large number of inflammatory cells infiltration were observed in wound tissue of rabbits in the 4 groups and a few epithelial cells were observed in wound tissue of rabbits in groups GF, AD, and CT. On PSD 14, more epithelial cells were observed in wound tissue of rabbits in groups GF and CT, and an obvious reduction of inflammatory cells infiltration was observed in wound tissue of rabbits in groups AD and CT. On PSD 21, there was a complete wound tissue structure and distinctive nuance of dyeing in wound tissue of rabbits in groups GF and CT while thinner new epithelium in wound tissue of rabbits in groups AD and BC, and inflammatory cell infiltration was observed in wound tissue of rabbits in group BC. On PSD 28, there was a complete wound tissue structure in wound tissue of rabbits in the 4 groups, the new epithelium in wound tissue of rabbits in groups GF, AD, and CT was thicker than that in group BC. (5) On PSD 7 and 14, the quantity of collagen fibers in wound tissue of rabbits in groups GF and CT was larger than that in the other two groups. On PSD 21, a large quantity of fibroblasts and well reorganized collagen fibers were observed in wound tissue of rabbits in groups GF and CT, a moderate quantity of fibroblasts and collagen fibers in a random arrangement were observed in wound tissue of rabbits in group AD, and a little quantity of fibroblasts and collagen fibers were observed in wound tissue of rabbits in group BC. On PSD 28, the quantity of collagen fibers in wound tissue of rabbits in the 4 groups was close to that of normal skin tissue, and the collagen fibers performed more well reorganized in wound tissue of rabbits in groups GF and CT. (6) On PSD 21, the numbers of microvessels per 200-time visual field in wound edge of rabbits in groups GF (31.6±1.2), AD (27.5±1.3), and CT (32.8±1.6) were significantly higher than the number in group BC (22.3±1.7, <0.05). The numbers of microvessels in wound edge of rabbits in groups GF and CT were significantly higher than the number in group AD (<0.05). The number of microvessels in wound edge of rabbits in group GF was close to that in group CT (>0.05). (7) On PSD 7 and 28, there were no statistically significant differences in content of hydroxyproline in wound edge of rabbits in the 4 groups (=0.916, 1.752, >0.05). On PSD 14 and 21, the content of hydroxyproline in wound edge of rabbits in groups GF, AD, and CT was significantly higher than that in group BC (<0.05). The content of hydroxyproline in the wound edge of rabbits in groups GF and CT was significantly higher than that in group AD (<0.05). The content of hydroxyproline in the wound edge of rabbits in group GF was close to that in group CT (>0.05). bFGF can be used solely or combined with AD to promote infective wound healing in New Zealand rabbit after complete debridement of wound, which is better than single use of AD.
探讨碱性成纤维细胞生长因子(bFGF)对新西兰兔清创后感染伤口愈合的影响。将32只3至4月龄的新西兰兔(雌雄不限)臀部皮内注射0.1 mL完全弗氏佐剂。六周后,每只兔背部两侧各注射0.5 mL 5×10(7)菌落形成单位/mL,以复制新西兰兔感染伤口模型。造模成功后,将32只兔按随机数字表分为生长因子(GF)组、抗结核药物(AD)组、联合治疗(CT)组和空白对照(BC)组,每组8只。彻底清创后,GF组兔伤口用重组牛bFGF凝胶(300 IU/cm(2),每个伤口约0.45 g)处理,AD组兔伤口用浸有6 mL异烟肼注射液和0.15 g利福平粉针剂的纱布覆盖,CT组兔伤口先按上述方法用重组牛bFGF凝胶处理后再用浸有异烟肼注射液和利福平粉针剂的纱布覆盖,BC组兔伤口用无菌纱布覆盖,每两天换药1次,直至伤口完全愈合。术后即刻及术后第7、14、21和28天,肉眼观察并拍摄每组兔的伤口情况。在术后第7、14、21和28天,计算伤口愈合率并记录伤口完全愈合时间。在术后第7、14、21和28天,采集伤口边缘组织样本,进行苏木精-伊红染色和Masson染色,行组织形态学观察。在术后第21天,采用免疫组织化学方法计数微血管数量。在术后第7、14、21和28天,采用酶联免疫吸附测定法测定伤口边缘羟脯氨酸含量。上述各项实验样本数均为8。数据采用重复测量方差分析、析因设计方差分析、单因素方差分析、最小显著差法检验和Bonferroni校正进行处理。(1)四组兔均存活至实验结束。术后即刻,四组兔的基础伤口均出现水肿。术后第7天,四组兔的伤口均收缩结痂,水肿减轻。GF组和CT组兔的伤口颜色变红。术后第14天,四组兔的伤口明显收缩。AD组和CT组兔的伤口无明显渗出物,而GF组有1只兔的伤口及BC组有2只兔的伤口红肿并有脓性渗出物。术后第21天,GF组和CT组兔的伤口基本愈合,而BC组有2只兔的伤口愈合缓慢,有脓性分泌物。术后第28天,四组兔的伤口基本愈合,而BC组有2只兔的伤口几乎未愈合,仍红肿。(2)术后第7至28天,GF组、AD组和CT组兔的伤口愈合率显著高于BC组(<0.05)。术后第14和21天,GF组和CT组兔的伤口愈合率显著高于AD组(<0.05)。术后第7至28天,GF组兔的伤口愈合率与CT组接近(>0.05)。(3)GF组、AD组和CT组兔伤口的完全愈合时间显著短于BC组(<0.05)。GF组和CT组兔伤口的完全愈合时间显著短于AD组(<0.05)。GF组兔伤口的完全愈合时间与CT组接近(>0.05)。(4)术后第7天,四组兔伤口组织均见大量炎性细胞浸润,GF组、AD组和CT组伤口组织可见少量上皮细胞。术后第14天,GF组和CT组伤口组织可见较多上皮细胞,AD组和CT组伤口组织炎性细胞浸润明显减少。术后第21天,GF组和CT组伤口组织有完整的伤口组织结构和明显的染色差异,AD组和BC组伤口组织新生上皮较薄,BC组伤口组织有炎性细胞浸润。术后第28天,四组兔伤口组织有完整的伤口组织结构,GF组、AD组和CT组伤口组织的新生上皮比BC组厚。(5)术后第7和14天,GF组和CT组兔伤口组织中的胶原纤维数量多于其他两组。术后第21天,GF组和CT组伤口组织可见大量成纤维细胞和排列良好的重组胶原纤维,AD组伤口组织可见中等数量的成纤维细胞和随机排列的胶原纤维,BC组伤口组织可见少量成纤维细胞和胶原纤维。术后第28天,四组兔伤口组织中的胶原纤维数量接近正常皮肤组织,GF组和CT组伤口组织中的胶原纤维排列更规整。(6)术后第21天,GF组(31.6±1.2)、AD组(27.5±1.3)和CT组(32.8±1.6)兔伤口边缘每200视野微血管数量显著高于BC组(22.3±1.7,<0.05)。GF组和CT组兔伤口边缘的微血管数量显著高于AD组(<0.05)。GF组兔伤口边缘的微血管数量与CT组接近(>0.05)。(7)术后第7和28天,四组兔伤口边缘羟脯氨酸含量差异无统计学意义(=0.916,1.752,>0.05)。术后第14和21天,GF组、AD组和CT组兔伤口边缘羟脯氨酸含量显著高于BC组(<0.05)。GF组和CT组兔伤口边缘羟脯氨酸含量显著高于AD组(<0.05)。GF组兔伤口边缘羟脯氨酸含量与CT组接近(>0.05)。bFGF单独使用或与AD联合使用可促进新西兰兔伤口彻底清创后感染伤口的愈合,效果优于单用AD。