封闭血管内皮生长因子C对角膜淋巴管的抑制和对同种移植物存活的促进作用研究

Effect of Vascular Endothelial Growth Factor C on the Inhibition of Corneal Lymphatic Vessels and Promotion of Allograft Survival

目的:研究封闭血管内皮生长因子C对角膜淋巴管的抑制效果和同种移植物存活的影响。方法:将100只BALB/c小鼠随机分为正常对照组、移植对照组、血管内皮生长因子C抑制组,每组20只。正常对照组不作特殊处理,移植对照组、血管内皮生长因子C抑制组制作小鼠角膜移植模型,术后分别通过腹腔注射,正常组和移植组注射等剂量的生理盐水,血管内皮生长因子C抑制组使用单克隆抗VEGF-C抗体阻断VEGF-C,在术后第3,7,10,14d分别取材,角膜淋巴管显像特征性使用全组织包埋免疫荧光法,移植物的免疫排斥反应通过裂隙灯检查并进行排斥反应评分(RI),另外VEGF-C的表达通过免疫组化和即时PCR来检测。结果:正常对照组角膜无新生淋巴管;其余两组小鼠移植后有新生淋巴管从角膜缘发出,淋巴管计数(LVC)在第14d达峰值,移植组的淋巴管计数高于血管内皮生长因子C抑制组。RT-PCR和Western blotting检测结果显示,正常对照组有少量VEGF-C表达,移植对照组VEGF-C表达较正常对照组显著增多且在第14d达峰值(P<0.05);与移植对照组比较,血管内皮生长因子C抑制组VEGF-C表达降低(P<0.05),而血管内皮生长因子C抑制组VEGF-C表达高于正常对照组(P<0.05)。通过裂隙灯观察角膜通透性,分别计数不同组的小鼠移植存活量来评价封闭血管内皮生长因子C对同种移植物存活的影响。结论:封闭血管内皮生长因子C对角膜淋巴管有明显的抑制作用,能够显著增强同种移植物存活数量。

Objective:To study the effect of vascular endothelial growth factor C on the inhibition of corneal lymphatic vessels and promotion of allograft survival.Methods:100 BALB/c mice were randomly divided into normal control group,transplantation control group,and vascular endothelial growth factor C inhibition group,20 rats in each group.The normal control group did not take special treatment;the transplantation control group and the vascular endothelial growth factor C inhibition group made the mice corneal transplantation model.After the operation,equivalent dose of the normal saline was injected into the normal group and the transplantation group through intraperitoneal injection,the vascular endothelial growth factor C inhibition group used the monkron anti VEGF-C antibody to block the VEGF-C.After 3,7,10,and 14 days after surgery,the characteristic of corneal lymphangiography was obtained by whole tissue embedding immunofluorescence method,and the graft rejection reaction was examined by a slit lamp and the rejection score(RI)was performed.In addition,the expression of VEGF-C was detected by immunohistochemistry and immediate PCR.Results:There was no new lymphatic tube in the normal control group.The other 2 groups had new lymphoid tubes from the limbus,and the lymphatic vessels count(LVC)was in the peak of 14d.The LVC of the transplanted group was higher than that of the vascular endothelial growth factor C inhibition group.The results of RT-PCR and Western blotting showed that there was a small amount of VEGF-C expression in the normal control group,and the expression of VEGF-C in the transplanted control group was significantly higher than that in the normal control group and at the peak in 14d(P<0.05).Compared with the transplant control group,the expression of VEGF-C in the VEGF-C inhibition group decreased(P<0.05),while the VEGF-C expression of VEGF-C inhibition group was higher than that of the normal control group(P<0.05).The corneal permeability was observed by slit lamp,and the survival of different groups of mice was counted to evaluate the effect of closed vascular endothelial growth factor C on the survival of the allograft.Conclusion:Blocking vascular endothelial growth factor C can significantly inhibit corneal lymphatic vessels,and significantly enhance the number of survival allograft.