IBI302对实验性脉络膜新生血管的干预作用及机制研究

摘要

目的 观察探讨IBI302对实验性脉络膜新生血管(CNV)的干预作用及其机制.方法 采用酶联免疫吸附测定法观察IBI302与血管内皮生长因子(VEGF)-A165、VEGF-A121和胎盘生长因子(PlGF)3种VEGF家族细胞因子和C3b、C4b两种补体蛋白的亲和力.采用人脐静脉血管内皮细胞(HUVEC)增生、迁移和管腔形成实验观察IBI302的抗VEGF活性.采用补体经典途径和旁路途径介导溶血实验观察IBI302的抗补体活性.采用激光诱导恒河猴CNV模型.分为CNV模型对照组、贝伐珠单抗注射组以及IBI302 0.25、0.50、1.25mg干预组.于给药后14、28 d,荧光素眼底血管造影及光相干断层扫描检查观察各组恒河猴CNV荧光素渗漏面积及视网膜厚度变化.给药后29 d,对比分析各组恒河猴房水中VEGF水平.结果 IBI302与VEGF-A165、VEGF-A121、PlGF以及C3b、C4b均具有较高亲和力.IBI302能明显抑制VEGF诱导的HUVEC增生、迁移和管腔形成.IBI302可抑制补体经典途径介导的绵羊红细胞溶血以及补体旁路途径介导的兔红细胞溶血.给药后14、28 d,IBI302 0.25、0.50、1.25 mg干预组恒河猴CNV渗漏面积明显减小;3组之间荧光素渗漏面积比较,差异无统计学意义(P＞0.05).贝伐珠单抗注射组恒河猴CNV荧光素渗漏面积也明显减小,其荧光素渗漏面积减小率较IBI302各浓度干预组偏小,但差异无统计学意义(P＞0.05).给药后14、28 d,IBI302 0.25、0.50、1.25 mg干预组恒河猴视网膜厚度均明显下降;3组之间视网膜厚度比较,差异无统计学意义(P＞0.05).贝伐珠单抗注射组恒河猴视网膜厚度也明显下降,其视网膜厚度下降率较IBI302各浓度干预组偏小,差异有统计学意义(P＜0.05).给药后29 d,CNV模型对照组、贝伐珠单抗注射组恒河猴房水VEGF浓度分别为(94.203±17.360)、(38.644±6.521) pg/ml;贝伐珠单抗注射组恒河猴房水VEGF浓度较CNV模型对照组明显降低,差异有统计学意义(P＜0.05).IBI302各浓度干预组恒河猴房水VEGF浓度均低于定量下限31.300 pg/ml.结论 IBI302能抑制实验性CNV,其机制与双重阻断VEGF和补体生物活性有关.%Objective To investigate the inhibitory effects of IBI302 on experimental choroidal neovascularization (CNV).Methods Affinity of IBI302 to vascular endothelial growth factor (VEGF) family cytokines (including VEGF-A165,VEGF-A121 and placental growth factor PlGF) and complements (C3b,C4b) was determined by enzyme-linked immunosorbent assay (ELISA).The antagonist effect of IBI302 on VEGF was measured by proliferation,migration and tube formation tests of human umbilical vein endothelial cells (HUVEC).The anti-complement activity of IBI302 was measured by hemolysis test mediated by complement classical pathway and alternative pathway.Rhesus laser-induced CNV model was divided into 5 groups including model control group,bevacizumab group,IBI302 0.25 mg group,IBI302 0.50 mg group and IBI302 1.25 mg group.Fluorescein angiography and optical coherence tomography were performed on these monkeys at 14 and 28 days after drug delivery to observe the fluorescein leakage area and retinal thickness.The aqueous VEGF concentration was measured at 29 days after drug delivery.Results IBI302 showed good affinity to VEGF-A165,VEGF-A121 and PlGF,as well as C3b and C4b.IBI302 significantly inhibited the proliferation,migration and tube formation of HUVEC induced by VEGF-A165.IBI302 inhibited the hemolysis induced by complements obviously.At 14 and 28 days after drug delivery,the area of fluorescein leakage and retinal thickness in IBI302 0.25 mg group,IBI302 0.50 mg group,IBI302 1.25 mg group were reduced.The differences of the area of fluorescein leakage and retinal thickness in three IBI302 groups were not significant (P＞0.05).At 29 days after drug delivery,the VEGF concentration in the aqueous of rhesus monkey in bevacizumab group [(38.644 ± 6.521) pg/ml] was decreased than that in model control group [(94.203± 17.360) pg/ml],the difference was significant (P＜ 0.05).The VEGF concentration in the aqueous of rhesus monkey in three IBI302 groups were less than 31.300 pg/ml.Conclusion IBI302 inhibited experimental CNV through blocking the activity of VEGF and complement.