血管内皮细胞和成纤维细胞混合移植对人工真皮血管化的影响

目的观察同种血管内皮细胞和成纤维细胞移植对人工真皮血管化的促进作用。方法在27只Wistar大鼠背部造成2．5 cm×2．5 cm全层皮肤缺损创面(2处／只),将其分为血管内皮细胞组:将血管内皮细胞混入0．5 ml纤维蛋白胶中,按1．0×105／cm2的密度均匀喷洒于移植床;混合组:将血管内皮细胞和成纤维细胞混入等量纤维蛋白胶后,同前密度喷洒于移植床;对照组:按同样方法喷洒等量纤维蛋白胶。随后各组移植人工真皮,每组9只大鼠18处创面。于移植后5、10 d切取移植的真皮及周围组织行HE、血管内皮生长因子(VEGF)、Masson和墨汁灌注染色,观察新生血管生长情况。于移植后5 d行伊文思蓝灌注,以分光光度计定量检测法测定微血管形成情况。结果移植后5 d,HE、VEGF、Masson和墨汁灌注染色均可见各组移植床有新生血管长入。HE染色见血管内皮细胞组、混合组新生血管数量分别为(14．2±3．6)、(12．1±2．5)条,较对照组[(3．9±1．6)条]明显增多(P<0．05)。移植后10 d,人工真皮内及移植床均有微血管形成,且胶原组织的合成增加。移植后5 d,经伊文思蓝灌注,收集并检测血管内皮细胞组、混合组真皮组织溶出的上清液,吸光度值分别为0．167±0．058、0．155±0．046,均高于对照组的0．066±0．024(P<0．05)。结论同种血管内皮细胞和成纤维细胞移植可促进创面愈合过程中的血管新生,加速人工真皮移植后血管化过程,促进类真皮组织的成熟。

Influence of mixed grafting of vascular endothelial cells and fibroblasts on the angiogenesis of artificial dermis

OBJECTIVE: To evaluate the effects of mixed grafting of vascular endothelial cells and fibroblasts on the angiogenesis of artificial dermis. METHODS: Full-thickness skin defects with the size of 2. 5 cm x 2. 5 cm were made on the back of 27 Wistar rats, and they were randomized into endothelium group ( cultured vascular endothelial cells were mixed into 0. 5 ml fibrin glue, and evenly sprayed onto wound bed in density of 1.0 x 10(5)/cm(2) before artificial dermis grafting) , mixed grafting group( cultured vascular endothelial cells and fibroblasts were mixed into 0. 5ml fibrin glue, and evenly sprayed onto wound bed in density of 1.0 x 10(5)/cm(2) before artificial dermis grafting) , and control group( with spraying of equal amount of fibrin glue on the wound bed). Each group consisted of 9 rats and 18 wounds. On 5 and 10 post-grafting day (PGF) , the grafts and surrounding tissue were harvested for the observation of neo-vascular growth with HE staining, VEGF antibody staining, Masson's trichrome staining and India ink staining. Evans blue perfusion method was also used to measure the angiogenesis of micro-vascular vessels on 5 PGD. RESULTS: The growth of new vessels was observed in each group on 5 PGD with HE staining, VEGF antibody staining, Masson's trichrome staining and India ink staining. Under high power field, the number of new vessels in endothelium and mixed grafting groups was (14. 2 3.6) and (12.1 +/- 2. 5) , which were obviously higher than that in control group (3.9 +/- 1.6, P < 0.05). On 10 PGD, angiogenesis and increased synthesis of collagen tissue were observed in both wound bed and artificial dermis. On 5 PGD, the amount of absorbed Evens blue extracted from the grafted dermis in endothelium group (0. 167 +/-0. 058) and mixed grafting group (0. 155 +/- 0. 046) was significantly higher than that in control group (0. 066+/-0. 024, P <0. 05). CONCLUSION: Mixed grafting of homogenous vascular endothelial cells and fibroblasts is beneficial for the promotion of neovascularization, acceleration of angiogenesis and maturation of artificial dermis during the wound healing process.