Molecular Biological Analysis of Wound Healing

1采用糖尿病小鼠和正常家兔，观察重组人胰岛素样生长因子-1 （rhIGF-I）和重组人胰岛素样生长因子结合蛋白-1 （rhIGFBP-1）对伤口愈合的刺激作用。在糖尿病C57BL/KsJ db/db小鼠背部和正常兔耳内侧制备直径6 mm的全层创面。将不同浓度的rhigf -1和/或rhIGFBP-1涂抹于db/db小鼠开放性创面，每天1次，连续5天；正常家兔创面后涂抹1次。创面切片于第7、8天通过测量再上皮化率（%）、肉芽组织面积（mm2）和毛细血管数量进行组织学评价。按rhIGF-I + rhIGFBP-1、rhIGF-I、rhIGFBP-1和单独使用载体的降序排列，各处理均能加速伤口修复。在db/db小鼠中，50 mg rhIGF-I和165 mg rhIGFBP-1（等摩尔比）联合使用可显著刺激小鼠的模拟组织形成（p<0.01）和毛细血管数量（p<0.05）。（Ref 1）2采用体外培养的正常人角质形成细胞，研究了紫外线B （UVB）照射对内皮素-1 （ET-1）和ET受体表达的影响。角质形成细胞在培养基中分泌ET-1，水平为2.1 pg/天/105个细胞。高达10 mJ/cm2的UVB照射使ET-1分泌增加3倍，并增强ET-1 mRNA的表达。在角化细胞中检测到ETA和ETB受体mrna，在5 mJ/cm2的UVB照射下，它们的表达上调。使用动物模型评估糖和聚维酮碘（SP）在伤口上的局部应用。与未处理的小鼠相比，SP处理的db/db小鼠伤口显示出良好的修复率。与未处理的兔相比，SP处理的正常兔伤口在这三个参数上的修复率都较好。（参考文献3）4使用体外模型研究了igf -1和IGFBP-1在伤口收缩中的作用。在无血清培养基中将人真皮成纤维细胞（1*105个细胞/ml）掺入猪I型胶原（终值0.21%）中。将成纤维细胞包埋于12孔板中的胶原凝胶从孔中浮起，然后将各种试剂添加到检测培养基中。通过测量胶原蛋白凝胶的直径来计算凝胶的表面积。高剂量（30和100 ng/ml） IGF-I分别显示6.8%和7.7%的凝胶收缩，10 ng/ml的des (1-3) IGF-I产生4.5%的凝胶收缩。同时，IGFBP-1未引起明显收缩。IGF-I和IGFBP-1在每种试剂1 ng/ml的浓度下联合使用，当每种试剂单独测试时未观察到凝胶收缩，产生14%的凝胶收缩。在实验培养基中加入抗IGF-I、抗igfbp -1或抗IGF-I受体抗体后，IGF-I和igfbp -1诱导的凝胶收缩被显著抑制（p<0.001）。（参考文献4）少

1 The stimulatory effect of recombinant human insulin-like growth factor-I (rhIGF-I) and recombinant human insulin-like growth factor-binding protein-1 (rhIGFBP-1) on wound healing was assessed using diabetic db/db mice and normal rabbits. Full thickness wounds of 6 mm diameter were prepared on the backs of diabetic C57BL/KsJ db/db mice and on the inner sides of normal rabbit ears. Various concentrations of rhIGF-I and/or rhIGFBP-1 were applied locally to the open wounds of db/db mice once daily for 5 days and to the covered wounds of normal rabbits once after wounding. Sections of the wounds were evaluated histologically on the 7th or 8th day by measuring reepithelialization (%), area of granulation tissue (mm2) and capillary numbers. Wound repair was accelerated by each of the treatments in descending order of rhIGF-I plus rhIGFBP-1, rhIGF-I,rhIGFBP-1, and vehicle alone. In db/db mice, the combination of 50 mg rhIGF-I and 165 mg rhIGFBP-1 (equimolar ratio) significantly stimulated gr … More anulation tissue formation (p<0.01) and capillary numbers (p<0.05). (Ref 1)2 The effect of ultraviolet B (UVB) irradiation on Endothelin-1 (ET-1) and ET receptor expressions was examined using cultured normal human keratinocytes. Keratinocytes secreted ET-1 in the medium at a level of 2.1 pg/day/105 cells. UVB irradiation up to 10 mJ/cm2 increased ET-1 secretion 3-fold, and potenitated expressions of mRNA for ET-1. Both ETA and ETB receptor mRNAs were detected in kerationocytes, and their expressions were up-regulated by 5 mJ/cm2 UVB irradiation. (Ref 2)3 Topical application of sugar and povidone iodine (SP) on wounds was assessed using animal models. The wounds in db/db mice treated with SP displayd good repair rates compared to those for non-treated mice. The wounds in normal rabbits treated with SP had good repair rates for all three parameters compared with those for non-treated rabbits. (Ref 3)4 Involvement of IGF-I and (IGFBP-1 in wound contraction was studied using an in vitro model. Human dermal fibroblasts (1*105 cells/ml) were incorporated into a porcine type I collagen (0.21% final) in serum-free medium. The fibroblast-embedded collagen gels in a 12-well plate were floated from the well, and various reagents were then added to the assay medium. The surface area of the gel was calculated by measuring the diameters of the collagen gel. IGF-I at high doses (30 and 100 ng/ml) revealed 6.8% and 7.7% gel contraction, respectively, and des (1-3) IGF-I at 10 ng/ml produced a 4.5% gel contraction. Meanwhile, IGFBP-1 did not induce any significant contraction. A combination of IGF-I and IGFBP-1 at 1 ng/ml of each reagent, a concentration at which gel contraction was not observed when each of the reagents was tested individually, produced a 14% gel contraction. When anti-IGF-I,anti-IGFBP-1 or anti-IGF-I receptor antibody was added to the assay medium, the IGF-I and IGFBP-1-induced gel contraction was significantly suppressed (p<0.001). (Ref 4) Less