三七总皂苷经VEGF-A抑制OIR新生血管的形成

Panaxnotoginsenosides PI3K/Akt/VEGF Inhibits Formation of OIR Neovascularization

目的在氧诱导的新生小鼠视网膜病变(OIR)模型中,观察三七总皂苷经PI3K/Akt调节血管内皮生长因子(VEGF)和成纤维细胞生长因子(FGF2)对视网膜新生血管的抑制作用及相关机制。方法将80只7 d龄C57BL/6J小鼠采用双盲随机对照法平均分为正常对照组、OIR对照组、三七总皂苷(PNS)组和雷珠单抗组,每组各20只小鼠。正常对照组从第7 d到第17 d在正常空气环境中饲养,余下3组第7 d到第11 d在氧浓度为75%±1%的饲养箱中饲养,之后在正常空气中饲养至第17 d,PNS组第12 d玻璃体腔注射PNS 1μL,雷珠单抗组第12 d玻璃体腔注射雷珠单抗1μL,所有小鼠在第17 d处死。免疫组织化学染色法检测VEGF的阳性表达; HE染色计数突破视网膜内界膜的新生血管内皮细胞核数;实时荧光定量PCR检测PI3k,Akt,VEGF-A,FGF2 mRNA的表达。结果OIR组可见大量内皮细胞核突破视网膜内界膜进入玻璃体; PNS组和雷珠单抗组见少量突破的内皮细胞核; OIR组突破视网膜内界膜新生血管内皮细胞核的数量显著高于正常对照组(P<0. 05),PNS组和雷珠单抗组明显低于OIR组(P<0. 05);免疫组化结果显示OIR组VEGF在视网膜血管内皮细胞中表达增多,PNS组和雷珠单抗组VEGF的表达明显降低;与正常对照组比较,OIR组VEGF-A,FGF2,PI3K,Akt的表达上调(P<0.05),与OIR组比较,PNS组和雷珠单抗组VEGF-A,FGF2,PI3K,Akt的表达下调(P<0. 05),PNS组与雷珠单抗组VEGF-A,FGF2,PI3K,Akt的表达无明显变化(P>0.05)。结论 PNS经PI3K/Akt能够调节VEGF和FGF2抑制氧诱导的新生小鼠OIR新生血管的形成。

Objective To observe the inhibitory effect of Panaxnotoginsenosides on retinal neovascularization and related mechanisms and regulate VEGF and fgf2 through PI3 K/Akt.Methods 80 C57 BL/6 J mice aged 7 days( P7) were divided into 4 groups by double-blind randomized control: normal control group A( group A),OIR control group( group B),Panax notoginsenosides group( group C) and recombinant antibody group( group D).Group A was raised in normal air from p7-p17.The OIR model were established in the groups of B,C and D and p7-p11 was raised in a feeding box with an oxygen concentration of 75%+/-1.Group C was injected with 1 μL Panaxnotoginsenosides in the vitreous cavity of P12,while group D was injected with 1 μL licensed resistance in the vitreous cavity of P12.Then they were raised in normal air until P17.All mice were killed on day 17,and positive expression of vascular endothelial growth factor was detected by immunohistochemical staining.HE staining broke through the number of neovascular-endothelial nuclei in the retinal inner boundary membrane,and real-time PCR was used to detect the changes in mRNA expression of PI3 k,Akt,vegf-a and fgf2. Results The retinal immunohistochemistry test showed that the expression of VEGF in the group B was significantly higher than that in the group A,and that in the group C and group D was significantly reduced.The retinal placement indicated that there was a large amount of vascular flexion hyperplasia in the group B,no retinal hyperplasia in the group A,and a small amount of retinal vascular hyperplasia in the group C and group D. HE staining showed that the number of vascular endothelial nuclei in the group B that broke through the retinal membrane was significantly increased,indicating the success of modeling.The number of vascular endothelial nuclei that broke through the retinal membrane in the group C and group D was significantly reduced,with statistically significant differences( P < 0. 05),while no significant changes were found in the blank group. Real-time PCR results showed that the expression of vegf-a and fgf2 was significantly down after the injection of Panax notoginsenosides( P<0.05).Conclusion The Panaxnotoginsenosides can regulate the formation of retinal neovascularization induced by inhibiting oxygen induced by VEGF and fgf2 through PI3 K/Akt.