昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路抑制斑马鱼血管新生

MA Rui-jiao; Maharajan Kannan; XIA Qing; ZHANG Shan-shan; TU Peng-fei; LIU Ke-chun; ZHANG Yun

doi:10.1007/s11655-022-3625-5

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昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路抑制斑马鱼血管新生

Original Article | Updated：2023-02-01

- 昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路抑制斑马鱼血管新生
- Kunxian Capsule Extract Inhibits Angiogenesis in Zebrafish Embryos via PI3K/AKT-MAPK-VEGF Pathway
- 中国结合医学杂志（英文版） 2023年29卷第2期页码：137-145
- Affiliations：
  
  1.Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan (250103), China
  2.Engineering Research Center of Zebrafish Models for Human Diseases and Drug Screening of Shandong Province, Jinan (250103), China
  3.State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing (100191), China
- Author bio：
  
  Dr. ZHANG Yun, E-mail: xiaohan_0818@163.com
- Funds：
  
  National Key R&D Program of China(2018YFC1707300);Shandong Provincial Natural Science Foundation(ZR2020YQ60);Jinan Talent Project for University(2021GXRC047)
- DOI：10.1007/s11655-022-3625-5
  中图分类号：
- 纸质出版日期：2023-02，
  
  网络出版日期：2022-12-15，
  
  录用日期：2022-05-18
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昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路抑制斑马鱼血管新生[J]. 中国结合医学杂志（英文版）, 2023,29(2):137-145.

MA Rui-jiao, Maharajan Kannan, XIA Qing, et al. Kunxian Capsule Extract Inhibits Angiogenesis in Zebrafish Embryos via PI3K/AKT-MAPK-VEGF Pathway[J]. Chinese Journal of Integrative Medicine, 2023,29(2):137-145.
昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路抑制斑马鱼血管新生[J]. 中国结合医学杂志（英文版）, 2023,29(2):137-145. DOI： 10.1007/s11655-022-3625-5.

MA Rui-jiao, Maharajan Kannan, XIA Qing, et al. Kunxian Capsule Extract Inhibits Angiogenesis in Zebrafish Embryos via PI3K/AKT-MAPK-VEGF Pathway[J]. Chinese Journal of Integrative Medicine, 2023,29(2):137-145. DOI： 10.1007/s11655-022-3625-5.

摘要

目的:

利用模式生物斑马鱼评价昆仙胶囊提取物的抗血管生成活性

并探讨其分子机制.

方法:

取昆仙胶囊5.0 g

用40%甲醇100 mL超声提取后冷冻干燥

即得昆仙胶囊提取物. 称取10.00 mg昆仙胶囊提取物

制备供试品溶液. 采用超高效液相色谱对昆仙胶囊提取物的主要活性成分雷公藤甲素和淫羊藿苷进行含量检测. 将血管标记绿色荧光的转基因斑马鱼Tg(flk1:GFP)在受精后20 小时脱去卵膜

分别用PTK 787和3.5、7、14和21 μg/mL 昆仙胶囊提取物处理. 给药处理24小时后

观察昆仙胶囊提取物对斑马鱼死亡率、畸形率、节间血管 (ISV) 形成以及血管生成相关基因mRNA表达水平的影响. 血管生成相关基因包括磷脂酰肌醇-3激酶 (PI3K)

蛋白激酶B (AKT)

细胞外信号调节激酶 (ERKs)

丝裂原活化蛋白激酶 (MAPK)

血管内皮生长因子 (VEGF) 和碱性成纤维细胞生长因子 (FGF-2) . 此外

昆仙胶囊提取物处理受精后72小时斑马鱼

观察给药24小时后肠下静脉 (SIV) 的发育情况.

结果:

昆仙胶囊提取物中雷公藤甲素和淫羊藿苷含量分别为0.089 mg/g和48.74 mg/g. 在斑马鱼实验中

昆仙胶囊提取物显著抑制ISV (

0.01)和SIV形成(

0.05). mRNA表达分析结果显示

昆仙胶囊提取物可显著抑制PI3K、AKT、ERKs和MAPK的mRNA水平

进一步抑制VEGF及其受体 (VEGFR和VEGFR-2) 表达

并下调FGF-2.

结论:

昆仙胶囊提取物通过调节PI3K/AKT-MAPK-VEGF通路对斑马鱼具有抗血管生成作用

具有治疗类风湿关节炎的潜力.

Abstract

Objective:

To investigate the anti-angiogenic activity of Kunxian Capsule (KX) extract and explore the underlying molecular mechanism using zebrafish.

Methods:

The KX extract was prepared with 5.0 g in 100 mL of 40% methanol followed by ultrasonication and freeze drying. Freeze dried KX extract of 10.00 mg was used as test stock solution. Triptolide and icariin

the key bioactive compounds of KX were analyzed using ultra-high performance liquid chromatography. The transgenic zebrafish Tg(flk1:GFP) embryos were dechorionated at 20-h post fertilization (hpf) and treated with PTK 787

and 3.5

14 and 21 μg/mL of KX extract

respectively. After 24-h post exposure (hpe)

mortality and malformation (%)

intersegmental vessels (ISV) formation

and mRNA expression level of angiogenic pathway genes including phosphoinositide 3-kinase (PI3K)

protein kinase B (AKT)

extracellular signal-regulated kinases (ERKs)

mitogen-activated protein kinase (MAPK)

vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF-2) were determined. Further

the embryos at 72 hpf were treated with KX extract to observe the development of sub-intestinal vein (SIV) after 24 hpe.

Results:

The chromatographic analysis of test stock solution of KX extract showed that triptolide and icariin was found as 0.089 mg/g and 48.74 mg/g

respectively

which met the requirements of the national drug standards. In zebrafish larvae experiment

KX extract significantly inhibited the ISV (

0.01) and SIV formation (P

0.05). Besides

the mRNA expression analysis showed that KX extract could significantly suppress the expressions of PI3K and AKT

thereby inhibiting the mRNA levels of ERKs and MAPK. Moreover

the downstream signaling cascade affected the expression of VEGF and its receptors (VEGFR and VEGFR-2). FGF-2

a strong angiogenic factor

was also down-regulated by KX treatment in zebrafish larvae.

Conclusion:

KX extract exhibited anti-angiogenic effects in zebrafish embryos by regulating PI3K/AKT-MAPK-VEGF pathway and showed promising potential for RA treatment.

关键词

血管新生类风湿关节炎滑膜炎症骨质疏松中药

Keywords

angiogenesisrheumatoid arthritissynovial inflammationosteoporosisChinese medicine

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