贝伐单抗结膜下注射对青光眼滤过术后滤过泡瘢痕化的抑制作用

Inhibitory effects of bevacizumab on filtering bleb scarring following trabeculectomy in rabbits

背景青光眼滤过手术术后滤过道的瘢痕化是导致手术失败的主要原因。传统的抑制滤过道瘢痕化的方法是丝裂霉素C的应用，但存在较多的并发症。研究表明贝伐单抗具有抑制新生血管和纤维增生的作用，其对青光眼滤过术后滤过泡瘢痕化是否有抑制作用受到关注。目的观察贝伐单抗结膜下注射对兔眼小梁切除术后滤过泡纤维瘢痕形成的抑制效果。方法按随机数字表法将7～9周龄新西兰大白兔40只随机分为4个组，各组兔右眼均行常规小梁网切除术。贝伐单抗单次注射组兔眼术毕结膜下注射贝伐单抗0．05ml（25mg／m1），贝伐单抗多次注射组兔眼分别于术毕及术后3、7d注射贝伐单抗，每次均注射0．05ml，丝裂霉素c组兔眼术毕局部涂用丝裂霉素c，生理盐水组兔眼术毕以同样的方法注射0．05ml生理盐水。所有兔眼术后每隔1日用Sehiotz眼压计测量眼压，行裂隙灯显微镜检查以观察滤过泡形态及其表面的血管分布，并用卡尺测量和计算滤过泡面积。分别于术后14d和28d摘取实验眼行滤过泡组织病理学检查，采用免疫组织化学法检测滤过道组织中血管内皮细胞标志物CD31的表达以计算微血管数目。结果各组兔眼术后眼压值的总体比较差异无统计学意义（F=0．88，P=0．47）。与贝伐单抗单次注射组、丝裂霉素c组和生理盐水组兔眼滤过泡的形态比较，术后7d贝伐单抗多次注射组兔眼滤过泡高度隆起且弥散，表面血管稀疏。贝伐单抗多次注射组兔眼滤过泡生存时间为27d，而贝伐单抗单次注射组、丝裂霉素C组均为19d，生理盐水组为13d。术后14d各组兔眼滤过道胶原纤维百分比分别为（49．18±1．54）％、（26．41±1．23）％、（50．68±1．87）％和（70．63±1．81）％，贝伐单抗单次注射组、贝伐单抗多次注射组、丝裂霉素c组滤过道胶原纤维百分比均低于生理盐水组，贝伐单抗多次注射组低于贝伐单抗单次注射组，差异均有统计学意义（P〈0．05）；术后28d贝伐单抗多次注射组胶原纤维百分比为（66．82±1．53）％，其他3个组出现瘢痕化。术后14d贝伐单抗多次注射组兔眼滤过道组织中微血管数目明显低于贝伐单抗单次注射组、丝裂霉素C组和生理盐水组，差异均有统计学意义（均P〈0．05）。术后28d贝伐单抗多次注射组兔眼滤过道组织中微血管数且为3．51±O．31，均高于贝伐单抗注射组、丝裂霉素组和生理盐水组，差异均有统计学意义（均P〈0．05）。结论青光眼滤过术后结膜下注射贝伐单抗有助于维持功能滤过泡的形态，抑制滤过道瘢痕化，提高手术的成功率。

Background The primary reason to trabeculectomy failure is fibrosis of conjunctiva and episclera because of progressive fibroblast proliferation and collagen deposition of the filtration bleb. Conventional methods of inhibiting bleb scarring was intraoperative application of mitomycin C （ MMC ） , but many complications occured after surgery. Researches showed that bevacizumab was an antifibrotic agent, and whether it can suppress scarring of filtering bleb after trabeculectomy is concerned. Objective The aim of this study was to evaluate the antifibrotic efficacy of bevacizumab after trabeculectomy in rabbits. Methods Forty New Zealand rabbits were randomly divided into four groups. Trabeculectomy was performed on the right eyes of each rabbits. The rabbits received subconjunctival injection of 0.05 ml bevacizumab （25 mg/ml） at the end of operation in the bevacizumab single injection group. The same dose of bevacizumab was respectively injected at the end of operation as well as 3 days and 7 days after operation in the bevacizumab repitition injection group,and 0. 05 ml normal saline solution was used in the same way in the normal saline group. In the MMC group, MMC cotton patch with 0.2 mg/ml was placed under the Tenon caplsule and scleral flap for 3 minutes during operation. The intraocular pressure （ IOP）, bleb area and shape were evaluated during the 28-day period. The animals were sacrificed on postoperative day 14 and 28, respectively for the histopathologic examination of bleb. The expression of CD31 in the bleb was detected by immunohistochemistry for the calculation of microvessels. All experiments were performed in accordance with the ethics code for animal experimentation and approved by the Institutional Review Board of Tianjin Eye Hospital. Results No significant difference was found in the postoperative IOP among the groups （F = 0.88, P = 0.47）. Compared with the bevacizumab single injection group, MMC group and normal saline group,the shape of bleb was higher and much diffuse with sparse vessels 7 days after operation in the bevacizumab repitition injection group. The survival time of bleb was 27 days, 19 days and 13 days in the bevacizumab repitition injection group,the bevacizumab single injection group,MMC group and normal saline group, respectively. The percentage of collagen deposition area was （49. 18±1.54）% ,（26.41±1.23）%,（50.68±1.87）% and （70. 63±1.81 ）% at day 14 postoperative in the bevaeizumab single injection group, bevacizumab repitition injection group, MMC group and normal saline group, respectively,with the Largest area in the normal sal_ine group, and percentage of collagen deposition area was significantly reduced in the bevacizumab repitition injection group compared with the bevacizumab single injection group （all at P〈0. 05 ）. The percentage of collagen deposition area was （66.82_± 1.53 ）% at day 28 postoperative in the bevaeizumab repitition injection group, while complete scarring was seen in other 3 groups. The number of microvessels was least at postoperative day 14 in the bevacizumab repitition injection group compared with the bevaeizumab single injection group, MMC group and normal saline group （all at P 〈 0. 05 ）. The number of mierovessels was more in postoperative day 28 in the bevaeizumab repitition injection group （3.51 ±0.31 ） compared with other groups （all at P 〈 O. 05 ）. Conclusions Subconjunctival injection of bevacizumab following trabeculectomy can improve the successful rate of surgery by remaining the survival time of filtering bleb,inhibiting the bleb scarring in rabbits.