通过成功构建大鼠结核性创面模型，动态观察创面病理变化，同时初步探讨结核性创面中巨噬细胞极化改变。

35只雌性8周龄SD大鼠，选取结核菌纯蛋白衍化物(PPD)试验无反应或弱反应的30只大鼠，采取MP弗氏完全佐剂致敏，6周后采用随机数字表法分为实验组和阴性对照组，每组各15只。实验组背部皮内注射牛分枝杆菌减毒株(BCG)(5×10⁷ CFU/mL、0.2 mL/只)；阴性对照组背部皮内注射无菌的PBS(0.2 mL/只)。大体观察创面演变情况，于注射后2、6、11 d，行皮肤组织石蜡包埋切片(每个时相点每组取5只大鼠)，苏木精-伊红染色观察各组创面愈合的情况及愈合过程中炎性细胞的浸润变化；病原微生物学检测鉴定菌株；注射后6 d，行免疫组织化学染色，CD68标记巨噬细胞，其亚型M1型巨噬细胞iNOS标记、M2型巨噬细胞CD206标记，通过免疫组织化学方法分析M1型和M2型巨噬细胞在实验组及阴性对照组中的分布情况；数据比较采用独立样本t检验。

经致敏大鼠，皮内注射BCG后创面观察到明显的红肿、液化、坏死和破溃等改变。苏木精-伊红染色显示液化坏死渐进增强后进而有效愈合的病理过程。抗酸染色结果：阳性。免疫组织化学显示：实验组创面组织液化坏死高峰期中CD68巨噬细胞、iNOS M1型巨噬细胞、CD206 M2型巨噬细胞数量分别为(84.8±3.4)、(60.8±2.8)、(13.2±2.2)个/HP，均显著高于阴性对照组(1.4±0.5)、(0.4±0.5)、(0.6±0.5)个/HP，差异均有统计学意义(t＝93.2、95.5、28.2，P值均小于0.05)；创面组织中iNOS M1型巨噬细胞[(60.8±2.78)个/HP]显著高于CD206 M2型巨噬细胞[(13.2±2.18)个/HP]，差异有统计学意义(t＝27.6，P<0.05)。

经过致敏的大鼠注射BCG可有效诱导液化和坏死，液化坏死高峰期创面组织中M1型巨噬细胞的数量比M2型巨噬细胞明显增多，且明显多于阴性对照组。实验结果提示，结核性创面液化坏死高峰期创面组织局部微环境可诱导巨噬细胞向M1型转变，有利于发挥巨噬细胞有效杀灭M.tb的作用。

To observe the pathological changes of wound dynamically and to discuss the changes of macrophage polarization of tuberculosis wounds by successfully constructing rat models of tuberculosis wounds.

Thirty-five SD rats(female, 8 weeks old)were injected with purified protein derivative(PPD), then 30 rats were selected, which were no inflammatory reaction or weak reaction in PPD experiment. The 30 rats were divided into experimental group (n=15) and negative control group (n=15) according to the random number table method. Thirty rats had been immunized and allergized with Freund′s complete adjuvant 6 weeks previously. Bacillus Calmette-Guérin (BCG) was intradermal injected to each experiment rat with the doses of 0.2 mL, 5×10⁷CFU/mL. PBS was intradermal injected to each rat in negative control group. The changes of wound were observed broadly. At 2, 6 and 11 days after injection, paraffin-embedded skin sections was did in each group (each time took five rats from each group), and hematoxylin-eosin staining was used to observe wound healing and infiltration of inflammatory cells in each group. The pathogenic microorganism was used to detecte and identify strains. Six days after injection, immunohistochemical staining was did, CD68-labeled macrophages, subtypes of M1 macrophage iNOS markers and M2 macrophages CD206 markers were used to analyze the expression of M1 and M2 macrophages by immunohistochemistry in the experimental group and the negative control group. All data were analyzed by t-test.

Apparent red, liquefaction, necrosis and ulceration were produced in the skin of rats injected with BCG. The appearance of tuberculousis wound from ulceration to healing was observed by the use of hematoxylin-eosin staining. The result of acid-fast staining was positive. The macrophage differentiation markers were counted by immunohistochemical staining in the peak period of tuberculousis wound: CD68, iNOS, CD206 in the experiment group were (84.8±3.4)/HP, (60.8±2.8)/HP, (13.2±2.2)/HP respectively, which were significantly higher than those in the control group (1.4±0.5)/HP、(0.4±0.50)/HP、(0.6±0.50)/HP, the differences were statistically significant (t=93.2, 95.5, 28.2, with P values below 0.05). iNOS [(60.8±2.8)/HP] in the experiment group were significantly higher than CD206 [(13.2±2.2)/HP] in the peak period of tuberculousis wound, the differences were statistically significant (t=27.6, P<0.05).

After sensitization of rats injected with BCG can effectively induce liquefaction and necrosis, the number of M1 macrophages in wound tissue at the peak of liquefaction necrosis was significantly higher than that of M2 macrophages, and significantly more than the negative control group. The experimental results suggest that the local microenvironment of wound tissue at the peak of liquefaction necrosis in tuberculous wounds can induce the change of macrophages to M1 type, which is beneficial to the effect of macrophages in killing M. tb effectively.