血管内皮生长因子DNA质粒对真皮支架修复猪Ⅲ度烧伤创面中血管化的影响

Angiogenesis of full-thickness burn wounds repaired with collagen-sulfonated carboxymethyl chitosan porous scaffold encoding vascular endothelial growth factor DNA plasmids

目的探讨血管内皮生长因子（VEGF）DNA质粒对胶原一磺化羧甲基壳聚糖真皮支架在猪Ⅲ度烧伤创面修复中血管化的影响。方法将单纯胶原-磺化羧甲基壳聚糖（A组）和负载VEGFDNA质粒（B组）胶原．磺化羧甲基壳聚糖两种不同真皮支架各移植于6只猪Ⅲ度烧伤切痂后创面，对植入支架后1、2、3周的创面及植入支架2周创面加植表皮后2周（植入支架后4周）的创面修复情况及病理切片进行观察。同时，通过采用免疫组织化学方法，对1、2、3、4周的支架及创面中表达CD31的新生血管、表达α-SMA的成熟血管和VEGF表达阳性的细胞数进行检测和观察。实验以不植入真皮支架（c组）的烧伤切痂后创面作为对照。结果A组和B组1～3周创面和2周创面植表皮后2周CD31表达的新生血管分别为18．7±3．1、25．7±2．3、36．8±2．5和26．2±2．9，24．5±3．8、32．3±2．8、39．2±2．2和27．3±3．0；α-SMA表达的成熟血管分别为11．7±1．9、20．5±1．9、35．0±4．5和24．0±2．8，20．2±3．1、33．5±3．7、38．2±4．5和26．5±2．3；VEGF表达阳性的细胞数分另0为48．7±7．9、141．7±9．1、201．5±8．6和107．0±8．2，97．3±7．9、172．3±8．1、208．7±8．3和114．0±5．8；C组1～4周创面CD31、α-SMA和VEGF的表达强度分别为6．0±2．0、9．8±3．4、19．3±2．5、18．7±2．2，3．7±2．0、8．7±1．8、13．0±2．5、14．0±2．8，3．5±2．3、10．3±3．5、23．0±5．6、21．5±5．1。A、B、C3组差异均有统计学意义（均P〈0．05）。结论外源性pDNA．VEGF导入胶原-磺化羧甲基弃聚糖真皮支架可促进创面肉芽组织表达更多的VEGF，加快支架血管化及创面修复。

Objective To investigate the effects of vascular endothelial growth factor （VEGF） DNA plasmids, encoded in collagen-sulfonated carboxymethyl chitosan porous scaffold, on the angiogenesis in full- thickness burn wounds. Methods Wounds and pathological changes were observed at Week 1,2, 3 and 4 after pure collagen-sulfonated earboxymethyl chitosan porous scaffold （group A） and eollagen-sulfonated carboxymethyl ehitosan porous scaffold encoding VEGF DNA plasmids （group B ） were transplanted onto eachar-removed wounds of full-thickness burn in 6 Bama miniature pigs respectively. Wound healing was observed by pathologic slides after epidermal grafting for 2 weeks （at Week 4） onto the surface of different dermal scaffolds transplanted on wounds for 2 weeks. At the same time, new-forming vessels expressing CD31 and mature vessels expressing α-SMA （a-smooth muscle actin） and the expression of VEGF in wounds at Week 1, 2, 3 and 4 were detected in situ by immunohistochemical staining. The burn wounds without any transplanted scaffold （ group C） were studied as the controls. Results Wounds with various scaffolds were different from those without any scaffold. Angiogenesis of the group B was better than that of the group A. Neo-forming micro-vessels expressing CD31 in the wounds of group A or B at Week 1, 2, 3 and4 were as follows： 18.7 ±3.1, 25.7±2.3, 36.8 ±2.5 ＆ 26.2 ±2.9; 24.5 ±3.8, 32.3 ±2.8,39.2±2.2 ＆ 27.3 ±3.0. Mature vessels expressing α-SMA： 11.7 ±1.9, 20.5± 1.9, 35.0 ±4.5 ＆ 24.0 ± 2. 8 ; 20.2 ±3.1, 33.5 ＋ 3.7, 58.2 ± 4. 5 ＆ 26. 5 ± 2.3. The expressions of VEGF ： 48.7 ± 7.9, 141.7±9.1, 201.5 ±8.6 ＆ 107.0 ±8.2; 97.3 ±7.9, 172.3 ±8.1, 205.7 ±8.3 ＆ 114.0 ±5.8. Expressing intensity of CD31, α-SMA and VEGF in the wounds of group C at Week 1, 2, 3 ＆ 4 ： 6.0 ± 2.0, 9.8±3.4, 19.3±2.5 ＆18.7±2.2; 3.7±2.0, 8.7±1.8, 13.0±2.5 ＆ 14.0±2.8; 3.5±2.3, 10. 5 ± 3.5, 23.0 ± 5.6 ＆ 21.5 ± 5. 1. There were significant statistical differences among three groups. Conclusion Artificial dermal scaffold encoding exogenous pDNA-VEGF can promote a quicker angiogenesis through a high expression of VEGF. Thus a full-thickness burn wound may be better repaired.