摘要
目的探讨局部应用NGF联合胰岛素对糖尿病大鼠烫伤创面血管中凋亡相关因子Bcl-2、Bax表达的影响及创面愈合的机制。方法取75只清洁级雄性Wistar大鼠,体重200～220 g,随机分为正常对照组(A组)、糖尿病对照组(B组)、胰岛素治疗组(C组)、NGF治疗组(D组)、NGF联合胰岛素治疗组(E组),每组15只。B、C、D、E组大鼠采用两步给药法腹腔注射链脲佐菌素(streptozotocin,STZ)建立糖尿病模型,STZ剂量分别为第1天10 mg/kg,第3天50 mg/kg;A组给予相同剂量柠檬酸缓冲液。模型制备后1个月,采用水蒸气烫伤法于各组大鼠背部制备2个深Ⅱ度烫伤创面。烫伤模型制备后,A、B组创面外敷3层生理盐水纱布;C组创面外敷3层浸润5 U胰岛素诺和灵30R的纱布,并每日腹部皮下注射诺和灵30R 4～6 U/kg;D组创面外敷3层浸润5 mL NGF溶液(25 U/mL)的纱布;E组联合C、D组方法处理。观察大鼠一般情况,伤后7、11、15、21 d大体观察各组创面愈合情况并计算创面愈合率,伤后3、7、11、15、21 d取创面组织行组织学及免疫组织化学染色观察,检测创面Bcl-2、Bax、CD34的表达并计算微血管密度。结果各组大鼠均存活至实验完成。随时间延长,各组创面逐渐缩小,其中E组创面愈合速度、皮肤角化、毛发生长及肉芽组织和胶原纤维生长均优于其余各组。伤后各时间点E组创面愈合率均高于其余各组,差异有统计学意义(P<0.05)。伤后随时间延长,各组CD34、Bcl-2表达逐渐增强,至15 d达高峰,21 d表达减弱;E组各时间点表达均强于其他各组(P<0.05)。伤后3 d各组均未见Bax表达,7 d后开始见新生血管内皮细胞Bax表达,且随时间延长表达逐渐增强,其中E组表达强度均低于其余各组(P<0.05)。结论局部联合应用NGF和胰岛素可通过抑制创面血管内皮细胞凋亡、增加创面血管生成,促进糖尿病大鼠创面愈合。
Objective To explore the possible mechanism of nerve growth factor （NGF） mixed insul in on the angiogenesis of burn wounds and the effect on the expressions of Bcl-2 and Bax in diabetic rats. Methods A total of 75 SPF male Wistar rats, weighing 200-220 g, were selected randomly and divided into nomal control （group A, n=15）, the rats with diabetic control （group B, n=15）, insul in treatment （group C, n=15）, NGF treatment （group D, n=15）, NGF and insul in treatment （group E, n=15） groups. In groups B, C, D, and E, streptozotocin was given by intraperitoneal injection at dose of 10 mg/kg on the 1st day and 50 mg/kg on the 3rd day to prepare the diabetic rat models. In group A, citric acid buffer at the same
dose was given. After 1 month of diabetic models, second degree scald was made on the back of the rats, and then wounds were treated with 3-layer normal sal ine gauze in groups A and B, with 3-layer gauze containing 5 U Novol in 30R and subcutaneous injection of Novol in 30R （4-6 U/kg） everyday in group C, with 3-layer gauze containing 5 mL NGF （25 U/mL） in group D, and with a combination of groups C and D in group E. At 7, 11, 15, and 21 days, the wound heal ing rate was calculated; at 3, 7, 11, 15, and 21 days, the expressions of Bcl-2, Bax, and CD34 were determined and the microvascular density was measured by immunohistochemistry staining. Results All rats survived till experiment was finished. The area of wounds became smaller gradually with time. Group E was better than other groups in the wound heal ing rate （P 〈 0.05）, the skin keratosis, the hair growth, and the granulation tissue and collagen fibers growth. With time, the expressions of CD34 and Bcl-2 increased gradually, reached the peak at 15 days and decreased at 21 days; the expression was stronger in group E than in other groups （P 〈 0.05）. At 3 days, Bax did not express; at 7 days, Bax began to express in new vascular endothel ial cells and the expression increased gradually with time; the expression was weaker in group E than in other groups （P 〈 0.05）. Conclusion A combination of NGF and insul in local appl ication can enhance the angiogenesis of the burn wound in diabetic rats and accelerate wound heal ing by increasing the expression of Bcl-2 and decreasing the expression of Bax and restraining apoptosis of the wounds vascular endothel ial cells of diabetic rats.
出处
《中国修复重建外科杂志》
CAS
CSCD
北大核心
2011年第3期354-360,共7页
Chinese Journal of Reparative and Reconstructive Surgery
