【摘要】 目的:外伤后病理性瘢痕的形成与创面的愈合过程有着密切的联系,创伤愈合时期引入细胞因子或生长因子及其他影响因素,调节创伤愈合机制,即可能改变细胞外基质结构,达到预防瘢痕形成的目的。实验拟观察TGF-β3c2s2基因转染的骨髓间充质干细胞在创面修复与重塑过程中对转化生长因子β1和转化生长因子β3表达的影响。方法:实验于2005-10/2007-03年在重庆医科大学附属儿童医院外科实验室、动物实验中心完成。①实验材料:四五周龄日本大耳白兔2只,体质量300~400g,雌雄不限;健康成年日本大耳白兔20只,体质量1.7~2.5kg,雌雄不限,均由重庆医科大学动物实验中心提供,实验过程中对动物处置符合动物伦理学标准。②实验方法:选取四五周龄日本大耳白兔分离培养骨髓间充质干细胞。选取健康成年日本大耳白兔20只,每只兔耳腹侧制作2个皮肤、软骨全层缺损创面共80个,创面以自身对照为前提随机分为4组,每组20个创面:实验组:加入Ad-TGF-β3c2s2转染的骨髓间充质干细胞;空白对照组:加入消毒的DMEM/F12(不含胎牛血清);腺病毒组:加入Ad-TGF-β3c2s2腺病毒;骨髓间充质干细胞组:加入骨髓间充质干细胞。③实验评估:观察瘢痕形成情况,苏木精-伊红染色、免疫组织化学技术观察瘢痕组织的组织学形态和检测瘢痕组织中转化生长因子β1和转化生长因子β3表达和变化规律。结果:①实验组在整个过程中无明显高出周围皮肤的瘢痕形成,其他组上皮化后,创面均逐渐形成不同程度的瘢痕,伤后45d瘢痕增生程度达高峰,明显高出皮肤表面;持续至90d左右。②实验组和腺病毒组结构较接近正常皮肤,比正常皮肤胶原排列致密,略厚;空白对照组、骨髓间充质干细胞组伤后45d可见浅层组织结构排列紊乱,胶原纤维粗大呈交错网织状排列。③实验组伤后21,45,90d转化生长因子β1表达明显低于其他组和正常皮肤(P<0.01),其表达随时间逐渐下降,转化生长因子β3表达高于其他组和正常皮肤(P<0.01),各组转化生长因子β3的表达随时间逐渐下降。结论:TGF-β3c2s2基因转染的骨髓间充质干细胞移植于局部创面,对创面修复过程中转化生长因子β1和转化生长因子β3表达有显著影响,通过这种影响减轻创面修复后增生性瘢痕的形成。更多还原

【Abstract】 AIM: Forming of pathological scar is associated with wound healing after trauma. Cytokine, growth factor or other influential factors can prevent scar forming by regulating wound healing and adjusting extracellular matrix. The study investigated effect of bone marrow mesenchymal stem cells (BMSCs) with TGF-β3c2s2 gene on the reparation and reconstruction of wound by detecting expressions of transforming growth factor beta 1 (TGF-β1) and TGF-β3. METHODS: Experiments were conducted in Surgical Laboratory and Center of Animal Experiment, Children’s Hospital, Chongqing Medical University from October 2005 to March 2007. Two Japanese flap-eared rabbits aged 4-5 weeks (300-400 g) and twenty healthy adult Japanese flap-eared rabbits (1.7-2.5 kg), irrespective of gender, were provided by Center of Animal Experiment, Chongqing Medical University. Animal intervention was accorded with the animal ethical standards. BMSCs were collected from Japanese flap-eared rabbits aged 4-5 weeks. Eighty wounds were generated on the gastroside of the ear of 20 healthy adult Japanese flap-eared rabbits, and were randomized into four groups in every rabbit. Wounds in the experimental group were treated with Ad-TGF-β3c2s2-transfected BMSCs. Wounds in the blank control group were treated with sterile DMEM/F12 (without fetal bovine serum). Wounds in the adenovirus group were treated with Ad-TGF-β3c2s2. Wounds in the BMSCs group were treated with BMSCs. Forming of scar was observed. Histology of scar was observed by hematoxylin-eosin staining. Expressions of TGF-β 1 and TGF-β3 were detected by immunohistochemical method. RESULTS: There was no prominence scar formed in the experimental group during the whole procedure. The wound of each control group gradually formed the different degree scars after epithelization. The hyperplasty of scars reached peak on 45 days after wounding and lasted about 90 days. The structure of the scars of the experimental group and the adenovirus group were similar to normal skin, but more dense and thick than normal skin. Disorder structure and overlapping arrangement, enlargement collagen fibers were showed in the scars of the blank control group and the BMSCs group at day 45. Expression of TGF-β1 protein in experimental group was decreased than other groups and normal skin at days 21, 45 and 90 (P < 0.01), and reduced over time. But expression of TGF-β3 was increased than other groups and normal skin (P < 0.01) at days 21, 45 and 90, and reduced over time. CONCLUSION: BMSCs with TGF-β3c2s2 gene transplantation into wound can inhibit the forming of hypertrophic scar by affecting expressions of TGF-β1 and TGF-β3.更多还原