目的 采用转基因血管荧光斑马鱼研究芦丁、甘草酸、汉防己甲素、葛根素、柚皮素、鞣花酸、黄芩苷、大黄素、穿心莲内酯、苦参碱促进血管新生的作用。方法 选取48 hpf转基因血管荧光Fli-1品系斑马鱼3 480尾于六孔板中,每孔均处理30尾(实验组),分别水溶给予芦丁81.90~1 637.95 μmol·L^-1、甘草酸60.76~1 215.17 μmol·L^-1、汉防己甲素5.02~80.29 μmol·L^-1、葛根素120.08~2 161.49 μmol·L^-1、柚皮素3.67~734.59 μmol·L^-1、鞣花酸0.21~3.31 μmol·L^-1、黄芩苷28.00~448.07 μmol·L^-1、大黄素0.19~740.08 μmol·L^-1、穿心莲内酯35.67~570.69 μmol·L^-1、苦参碱50.33~805.28 μmol·L^-1,同时设置对照组(养鱼用水处理斑马鱼),处理24 h后观察记录斑马鱼的毒性表型和死亡情况,确定供试品对斑马鱼的最大耐受浓度(MTC)。用10个化合物的MTC与斑马鱼共培养24 h,检测肠下血管面积和肠下血管出芽数。结果 与对照组比较,芦丁组、甘草酸组、汉防己甲素组、葛根素组、柚皮素组、黄芩苷组、穿心莲内酯组、苦参碱组斑马鱼肠下血管面积像素升高,其中甘草酸组、汉防己甲素组、黄芩苷组、穿心莲内酯组差异显著(P<0.05、0.01、0.001);鞣花酸组、大黄素组斑马鱼肠下血管面积像素降低,差异不显著;芦丁组、汉防己甲素组、葛根素组、柚皮素组、鞣花酸组、苦参碱组斑马鱼肠下血管出芽数比对照组多,芦丁组差异显著(P<0.05);甘草酸组、黄芩苷组、穿心莲内酯组斑马鱼肠下血管出芽数比对照组少,大黄素组斑马鱼肠下血管出芽数与对照组相当。结论 芦丁、甘草酸、汉防己甲素、黄芩苷和穿心莲内酯对斑马鱼具有促进血管新生作用;葛根素、柚皮素、鞣花酸、大黄素和苦参碱对血管新生无明显影响。

Objective The effects of rutin, glycyrrhizic acid, tetrandrine, puerarin, naringin, ellagic acid, baicalin, emodin, androgynolide and matrine on angiogenesis were studied by transgenic fluorescein vascular zebrafish. Methods A total of 3 480 zebrafish of 48 hpf transgenic fluorescein FlI-1 strain were selected and placed in six well plates, with 30 fish in each well (experimental group). Rutin of 81.90—1 637.95 μmol·L^-1, glycyrrhizic acid of 60.76—1 215.17 μmol·L^-1, tetrandrine of 5.02— 80.29 μmol·L^-1, puerarin of 120.08—2 161.49 μmol·L^-1, naringin of 3.67—734.59 μmol·L^-1, ellagic acid of 0.21—3.31 μmol·L^-1, baicalin of 28.00—448.07 μmol·L^-1, emodin of 0.19—740.08 μmol·L^-1, androgynolide of 35.67—570.69 μmol·L^-1, matrine 50.33— 805.28 μmol·L^-1 were given respectively in aqueous solution, and the control group (zebrafish treated with fish culture water) were set up at the same time. After 24 h of treatment, the toxic phenotype and death of zebrafish were observed and recorded. The maximum tolerance concentration (MTC) of zebrafish was determined. Zebrafish were co-cultured with MTC of 10 compounds for 24 h, and the area and sprouted number of subintestinal vessels were detected. Results Compared with control group, the rutin group, the glycyrrhetinic acid group (P<0.05), the hanfangenin group (P<0.001), the gerberoside group, the naringenin group, the baicalin group (P<0.01), the andrographolide group (P<0.001), and the bitter ginseng base group had higher subintestinal area pixels in zebrafish, ellagic acid group, rhodopsin group zebrafish subintestinal vascular area pixel was lower. The number of subintestinal vascular outgrowths in zebrafish was more in the rutin group (P<0.05), hanfanganin, geranium group, naringenin group, ellagic acid group, and bitter ginseng base group than that in control group. The number of subintestinal vascular sprouts in zebrafish of glycyrrhizic acid group, baicalin group, and andrographolide group was less than that in the normal control group, and the number of subintestinal vascular sprouts in the rutin group was comparable to that in the rhodopsin group. Conclusion Rutin, glycyrrhizic acid, hanfenacin, baicalin, and andrographolide had pro-angiogenic effects on zebrafish. Puerarin, naringenin, ellagic acid, rhodopsin and bitter ginseng alkaloids had no significant effect on angiogenesis.

R285.5

重大新药创制科技重大专项(2017ZX09304003)