腹壁-盲肠损伤粘连模型的建立及mXG温敏水凝胶的防粘连机制研究

Modeling of Abdominal Wall-Cecum Injury Adhesion and the Anti-adhesion Mechanism of mXG

目的构建腹壁-盲肠损伤粘连模型并探究改性木葡聚糖(mXG)温敏水凝胶对腹壁-盲肠损伤粘连的防治效果。方法选用SD大鼠构建腹壁-盲肠损伤粘连模型,随机分为空白对照组(Control)、商业化壳聚糖膜对照组(Film)和mXG温敏凝胶组(Hydrogel),每组16只。Hydrogel组,将1 mL 4%(m/V)mXG溶液涂抹至腹壁和盲肠的创面,待凝胶形成(3 min)后关腹;Film组,关腹前将2 cm×3 cm大小的壳聚糖防粘连膜覆盖于腹壁创面;Control组,关腹前将1 mL生理盐水注射于腹壁和盲肠的创面。术后7 d和14 d,开腹检查腹壁与盲肠的粘连情况,采用双盲设计进行粘连评分。取同一部位粘连组织或创面组织,进行HE、Masson和Van Gieson染色后,进行防粘连效果组织学评价,免疫组织化学染色检测转化生长因子-β1(TGF-β1)和结缔组织生长因子(CTGF)的表达情况。另取12只SD大鼠予以造模和mXG温敏水凝胶干预,术后第1周和6周取心、肝、脾、肺、肾等主要脏器使用HE染色进行组织学检查,进行体内毒性评价。结果动物模型大体观察发现,Hydrogel组的大鼠基本未发生粘连,Film组发生轻度粘连,Control组发生了严重的粘连,差异有统计学意义(P<0.05)。组织学镜下观察发现,7 d时Hydrogel组创面恢复良好,未看到变异的组织,有一定程度的炎性细胞浸润;14 d时创面组织中的炎性细胞明显减少,新生间皮层下仅含有少量胶原纤维的愈合组织;而7 d、14 d,其余两组均出现不同程度粘连的病理表现。免疫组织化学染色发现,Hydrogel组TGF-β1和CTGF的阳性表达一致且较其它两组弱,差异有统计学意义(P<0.05)。体内毒性检测发现,使用mXG凝胶的动物在不同时间点各脏器的结构均没有出现明显变化。结论 mXG温敏水凝胶能够在粘连形成的关键时期起到良好的物理隔离作用,有效预防盲肠-腹壁粘连的发生。

Objective To construct the adhesion model of abdominal wall-cecum injury and explore the prevention and treatment effect of modified xyloglucan(mXG)thermosensitive hydrogel on abdominal wall-cecal injury adhesion.Methods SD rats were used to construct the abdominal wall-cecal injury adhesion model.Model mice were randomly divided into blank control group(Control),commercial chitosan membrane Control group(Film)and mXG thermosensitive hydrogel group(Hydrogel),each group contained 16 rats.In the Hydrogel group,1 mL 4%(m/V)mXG solution was smeared on the wound surface of abdominal wall and the cecum,then closed the abdomen after gel was formed(3 min).In the Film group,2 cm×3 cm chitosan anti-adhesion Film was applied onto the wound surface of the abdominal wall before abdominal closure.In the Control group,1 mL normal saline was applied onto the wound surface of abdominal wall and the cecum before abdominal closure.On 7 and 14 dafter the operation,rats’ abdominal cavity was opened by surgery to examine and score the adhesion grade between the abdominal wall and the cecum,with double-blind design.Meanwhile,the adhesion tissue or wound tissue was taken and stained by HE,Masson and Van Gieson to histological evaluate the anti-adhesion effect.The expression of transforming growth factor-β1(TGF-β1)and connective tissue growth factor(CTGF)was determined by immunohistochemical staining as well.Another group of 12 SD rat models were subjected to mXG thermosensitive hydrogel intervention.At the 1 and 6 weeks postoperation,rats main organs such as heart,liver,spleen,lung and kidney were taken for histological examination with HE staining for the purpose of evaluation the toxicity of mXGin vivo.Results Adhesion grade evaluation results showed that Film group rats occurred mild adhesion,Control group rats occurred severe adhesion,while in Hydrogel group hardly rats occured adhesion,and the differences were statistically significant(P<0.05).Histological results showed that the Hydrogel group rats recovered well at 7 d after surgery.In healing wound tissue,no mutated tissue was observed,but a certain degree of inflammatory cell infiltration was still existed.At 14 dafter surgery,the inflammation cells in the wound were significantly reduced,and the healing tissue containing only a small amount of collagen fibers under the neonatal mesothelial layer.But the other two groups showed different degrees of adhesion at the 7 and 14 dpost surgery.Immunohistochemical staining showed that the expression of TGF-β1 and CTGF in the Hydrogel group were both weaker than those in the other groups,and the difference was statistically significant(P<0.05).In vivo toxicity tests did not show significant changes in the structure of the organs of mXG gel intervention rats at different time points.Conclusion mXG thermosensitive hydrogel plays a good role in physical isolation during the key period of adhesion formation and effectively prevent the occurrence of cecum-abdominal adhesion.