环氧化酶2在翼状胬肉新生血管形成中的作用及机制研究

发布时间：2019-01-03 12:50

【摘要】：目的翼状胬肉是我国常见眼表疾病之一，本病尚无有效的药物治疗，手术切除是目前治疗翼状胬肉的主要方法，但手术后并发症多且手术后的复发率很高，所以治疗相当棘手，因此进一步研究翼状胬肉的发病机制十分必要。本实验旨在研究翼状胬肉中COX-2与新生血管的表达情况，探讨COX-2在翼状胬肉新生血管生成和发展中的作用，为探索翼状胬肉的靶向治疗提供新的理论依据。 方法使用苏木素-伊红染色（HE染色）法及免疫组化的方法对实验标本进行研究。第一步：临床收集组织标本用于切片制作。门诊手术获得正常球结膜组织23例，翼状胬肉组织45例，术前裂隙灯检查排除参与者其他角结膜性疾病。所有组织经过固定剂固定，浓度依次递升的乙醇脱水，使用石蜡包埋，组织切片，并做好标记并保存备用。第二步：HE染色法观察翼状胬肉与正常球结膜组织的形态学差异。取切片，经脱蜡后苏木素浸染5min，将细胞核染成蓝色，盐酸酒精分色后伊红染色10s，切片脱水、透明后中性树胶封片，显微镜下观察。第三步：免疫组织化学方法分别检测翼状胬肉和球结膜组织中COX-2、VEGF、PDGF与CD31分布及表达情况。取切片，经脱蜡后置于PH6.0的抗原修复液中高热修复6min，3％去离子水封闭内源性抗原，在不同组织编号的玻片上分别加入一抗，4℃孵育过夜，冲洗后加入对应二抗，DAB显色，显微镜下观察并计数。 结果（1）HE染色后正常球结膜组织与翼状胬肉组织中形态学表现：正常球结膜组织表层上皮细胞排列整齐，基底上皮细胞边界清晰，间质血管少；翼状胬肉翼状胬肉组织表现为上皮层及基质浅层成纤维细胞增殖，，表层上皮细胞排列紊乱，细胞体积增大，基底上皮边界模糊，间质中新生血管大量增生。（2）免疫组织化学方法分别检测翼状胬肉和球结膜组织中COX-2、VEGF、PDGF与CD31分布及表达情况：23例球结膜组织中均为呈阴性，翼状胬肉组织样本中有36例COX-2表达阳性(P 0.01)； CD31在两种组织中均有表达，通过计数及统计，翼状胬肉组织中MVD为20.3±4.4，球结膜组织中MVD为9.7±2.8(P0.01)； COX-2表达阳性的翼状胬肉组织中MVD为19.06±1.84，表达阴性的翼状胬肉组织中MVD为10.44±2.98(P0.01)。翼状胬肉组织切片中34例PDGF表达阳性，36例COX-2表达阳性的胬肉组织中32例PDGF阳性；翼状胬肉组织切片中39例VEGF表达阳性，36例COX-2阳性的胬肉组织中34例VEGF表达阳性；差异有统计学意义（P0.005）。在促进翼状胬肉新生血管生成过程中COX-2的表达与血管相关因子VEGF和PDGF具有相关性(r＝0.458, r＝0.621)。 结论本实验研究发现翼状胬肉组织中MVD、VEGF、PDGF以及COX-2的表达明显强于正常球结膜组织，另外，COX-2可能协同VEGF及PDGF共同刺激翼状胬肉新生血管的发生。以上结果不仅证实COX-2在翼状胬肉的发病中发挥重要的作用，同时进一步证实翼状胬肉属于紫外线诱导相关肿瘤性疾病。
[Abstract]:Objective pterygium is one of the common ocular surface diseases in China. There is no effective drug treatment for pterygium. Surgical resection is the main method to treat pterygium at present, but there are many complications after operation and the recurrence rate after operation is very high, so the treatment is quite difficult. Therefore, it is necessary to further study the pathogenesis of pterygium. The purpose of this study was to investigate the expression of COX-2 and neovascularization in pterygium and to explore the role of COX-2 in angiogenesis and development of pterygium, and to provide a new theoretical basis for the targeted treatment of pterygium. Methods hematoxylin-eosin staining (HE staining) and immunohistochemical method were used to study the experimental specimens. Step 1: clinical tissue specimens are collected for slice making. 23 cases of normal bulbar conjunctiva and 45 cases of pterygium were obtained by outpatient operation. All tissues were fixed with fixants, then dehydrated by ethanol with increasing concentration, paraffin embedded, tissue sections, labeled and preserved. Step 2: the morphological difference between pterygium and normal bulbar conjunctiva was observed by HE staining. The nuclei were stained blue after dewaxing and stained with hematoxylin for 5 min. The nuclei were stained with eosin for 10 s after separation of hydrochloric alcohol. The distribution and expression of COX-2,VEGF,PDGF and CD31 in pterygium and bulbar conjunctiva were detected by immunohistochemistry. After dewaxing, 3% deionized water was placed in the antigenic repair solution of PH6.0 for 6 min to seal the endogenous antigen. The first antibody was added to the different tissue numbered glass slides, incubated overnight at 4 鈩

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