目的：探讨外源性色素上皮衍生因子(pigment epithelium-derived factor， PEDF)抑制大鼠角膜新生血管(corneal neovascularization， CNV)形成的作用机制，检测外源性PEDF对CNV模型中内源性血管内皮生长因子(vascular endothelial growth factor， VEGF)及PEDF、细胞凋亡受体/配体(Fas/FasL)表达的影响，以及对CNV内皮细胞凋亡的影响(脱氧核糖核苷酸末端转移酶介导的缺口末端标记法——TUNEL法测定)。

方法：大鼠随机分为3组：PEDF治疗组(A组)、生理盐水对照组(B组)、正常组(C组)。建立碱烧伤诱导的大鼠CNV模型，A组和B组分别给予PEDF+氯霉素滴眼液和生理盐水+氯霉素滴眼液点眼，采用裂隙灯观察CNV的生长情况，用免疫组化方法检测并半定量分析VEGF， PEDF，Fas及FasL在正常角膜组织和碱烧伤后4，7，10，14d角膜组织中的表达，用TUNEL法检测CNV内皮细胞的凋亡情况。

结果：正常角膜组织中可以检测到高PEDF表达和低VEGF表达，也可以检测到一定的低Fas和FasL表达。各时间点角膜新生血管的面积，A组均低于B组(P<0.05)。免疫组化显示，碱烧伤后4，7，10，14d A组VEGF的表达均低于B组(P<0.05)，PEDF的表达均高于B组(P<0.05)，A组Fas/FasL的表达明显高于B组(P<0.05)，A组TUNEL法测到的角膜新生血管内皮细胞凋亡计数和凋亡指数AI明显高于对照组(P<0.05)。两组检测到的VEGF均在第7d达到峰值，随后下降； 而PEDF和Fas及FasL在第10d达到峰值，随后下降。碱烧伤后第4～10d，VEGF/PEDF>1，CNV逐渐生长并达到峰值； 第10d后，VEGF/PEDF<1，CNV开始逐渐退化。

结论：PEDF抑制碱烧伤后CNV的作用机制可以归结为两点：(1)下调VEGF的表达，控制PEDF/VEGF的动态比值，抑制CNV生长；(2)上调Fas/FasL等凋亡受体的表达，诱导角膜新生血管内皮细胞的凋亡，促进CNV的退化。

AIM:To explore the mechanism of pigment epithelium-derived factor(PEDF)in the development of experimental rat corneal neovascularization(CNV).

METHODS: Rat corneal alkaline burn model was established in both eyes by routine method in 40 anesthetized female Sprague-Dawley rats. The rats were randomly assigned to 2 groups with 20 rats each for topical administration of recombinant PEDF combined with chloramphenicol or normal saline combined with chloramphenicol(as control). At different intervals(4, 7, 10, and 14 days)of the treatment, rats were euthanized and the corneas removed for immunohistochemistry analyses to measure expression levels of PEDF, vascular endothelial growth factor(VEGF), Fas and FasL. The eyes of ten healthy rats were used as normal control. Meanwhile, the apoptosis of neovascular endothelial cells was detected by TUNEL method on the 4^th, 7^th, 10^th and 14^th day respectively after the burn.

RESULTS: There were high levels of PEDF expression and low levels of VEGF, Fas and FasL in the normal cornea. VEGF levels were significantly induced by chemical cauterization in the groups treated with chloramphenicol combined with normal saline, demonstrating CNV. In contrast, the PEDF treatment prevented the over expression of VEGF induced by the cauterization and unregulated the expression of Fas and FasL. In CNV tissues, the positive immune reaction of VEGF was most apparent during the 7^th day and then declined thereafter. However, the most positive expression of PEDF, Fas, FasL was on the 10^th day and then declined slowly after thereafter. The ratio of PEDF/VEGF raised from <1 to >1 in the course and the hinge was on the 10^th day. Certain time correlation existed between the dynamic expressions of VEGF and PEDF and the development of CNV. The expression of Fas and FasL correlated to PEDF closely in the whole procession which may underlie a simulative relationship between them. The apoptosis of CNV endothelial cells expressed most positively on the 10^th day and it was always much more intense in the PEDF group than in the normal saline group.

CONCLUSION: The expressions of VEGF and PEDF are remarkably expressed in experimental rat CNV tissues, and the fluctuation of which is consistent with the development of CNV. The breakdown of the balance between the two factors may play a role in CNV occurrence and development. That the PEDF downregulates VEGF expression and upregulates Fas and FasL expression which induces the apoptosis of CNV endothelial cells results in the inhibition of corneal NV induced by chemical cauterization. The results suggested the possible mechanism of PEDF in the therapeutic function for CNV diseases.

广东省科技计划基金(No.2008B080703037); 广州市海珠区科技计划基金(No.2010-T-26)