激光共焦显微镜与组织病理学对翼状胬肉组织中微血管密度研究的比较

背景 研究表明,翼状胬肉是眼表的一种异常细胞增生性疾病,血管因素在它的生长和复发中起重要作用.多种抗新生血管治疗方法被应用于翼状胬肉,但活体观察翼状胬肉血管情况的研究尚少见.目的 比较角膜激光共焦显微镜与免疫组织化学染色对翼状胬肉组织中微血管密度研究的结果. 方法 采用前瞻性病例对照研究方法,收集原发性翼状胬肉患者20例20眼为翼状胬肉组,手术中获取翼状胬肉组织；另收集年龄和性别匹配的行内眼手术及斜视手术的患者20例20眼为正常球结膜组,手术中获取正常鼻侧球结膜组织作为对照,翼状胬肉组患者均于术前用海德堡角膜激光共焦显微镜采集5幅翼状胬肉头部的微血管图像,同时采集正常球结膜组正常球结膜微血管图像.对两组患者手术中获得的标本进行常规组织病理学检查和CD31免疫组织化学染色,计算组织标本中微血管密度值,对翼状胬肉组患者激光共焦显微镜所测的微血管密度值以及手术标本中的微血管密度值分别与正常球结膜组进行比较,并分别将两组用两种方法测得的血管密度值进行相关分析.结果 角膜激光共焦显微镜下翼状胬肉基质中的微血管密度( MVD)值为( 8929±2993) μm/mm2,正常球结膜中的微血管密度值为( 4202±692) μm/mm2,二者比较差异有统计学意义(t=6.881,P＜0.01).翼状胬肉的组织标本中CD31染色阳性的微血管数为(21.00±4.06)个/高倍视野,正常球结膜组为(6.07±1.75)个/高倍视野,二者比较差异有统计学意义(t=12.312,P＜0.01).翼状胬肉组和正常球结膜组激光共焦显微镜活体测得的MVD值与免疫组织化学离体测得的MVD值均呈显著正相关(翼状胬肉组:r=0.649,P＜0.01；正常球结膜组:r=0.572,P＜0.01).结论 角膜激光共焦显微镜活体动态观察翼状胬肉组织微血管的方法优于离体的免疫组织化学检测法,角膜激光共焦显微镜对翼状胬肉的血管形态进行活体研究有助于进一步研究翼状胬肉组织的生物学行为及其机制.

Analysis of microvessel density in pterygium tissue with corneal laser confocal microscopyc in vivo and immunohistochemistry in vitro

Background Pterygium is an ocular surface disease of abnormal cell proliferative kind and angiogenesis plays an important role in its development and recurrence.Several anti-angiogenic therapies have been used to treat pterygium,but there very few studtes for the in vivo observation of the microvessles in pterygium.Objective This study was to observe angiogenesis in pterygium with a high-resolution confocal microscope in vivo and to perform immunohistochemical study in vitro. Methods A prospective case-controlled study was designed.Twenty eyes of 20 consecutive patients with primary pterygia and 20 age- and sex-matched patients with inner eye diseases and strabismus with normal conjunctiva were enrolled in this study.An in vivo confocal microscopy imaging system (Heidelberg Retina Tomograph Ⅱ Rostock Cornea Module) was used to collect microvascular pictures from the anterior part of pterygia and normal nasal conjunctiva of controls,and then immunochemistry was performed to examine the expression of CD31 in microvessel in vitro.The vascular density values were compared between these two groups.The correlation of vascular density values between in vivo Heidelberg Retina Tomograph and in vitro immunohistochemistry was calculated.Written informed consent was obtained from pationts before any examination and surgery. Results Under the in vivo confocal microscope,the microvessel density was (8929±2993) μm/mm2 and (4202 ±692)μm/mm2,respectively,in pterygium and the normal conjunctiva group with a statistically significant difference between them (t =6.881,P＜0.01 ).Immunochemistry revealed that the expression of CD31 to measure vascular density was ( 21.00 ± 4.06/400 × field ) and ( 6.07 ± 1.75/400 × field ) in pterygium and the normal conjunctiva group,showing significant difference (t =12.312,P＜0.01 ).Positive correlations were found in the vascular density values between in vivo corneal laser confocal microscopy examination and in vitro immunochemistry examination in both the pterygium group and normal conjunctiva group (pterygium group:r=0.649,P＜0.01 ;normal conjunctiva group: r=0.572,P＜0.01 ) Conclusions In vivo confocal microscopy imaging is superior to in vitro immunochemistry in evaluating the microvessel of pterygium.The results of this study offer a new way index for further investigation of the biological behavior of pterygium and its mechanism.