Mechanism of astragaloside Ⅳ regulating exosome production and secretion and inhibiting metastasis of colorectal cancer

PU Yunzhou; LI Haoze; LI Ling; ZHOU Jing; JI Qing

doi:10.16305/j.1007-1334.2023.2303095

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Mechanism of astragaloside Ⅳ regulating exosome production and secretion and inhibiting metastasis of colorectal cancer

更新时间：2023-06-16

- Mechanism of astragaloside Ⅳ regulating exosome production and secretion and inhibiting metastasis of colorectal cancer
- Shanghai Journal of Traditional Chinese Medicine Vol. 57, Issue 6, Pages: 41-49(2023)
- 作者机构：
  
  1.上海中医药大学附属曙光医院肿瘤科/肿瘤研究室（上海 201203）
  2.宁波市第二医院中西医结合肝病科（浙江宁波 315010）
- 作者简介：
- 基金信息：
- DOI：10.16305/j.1007-1334.2023.2303095
  CLC：
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浦匀舟,李昊泽,李玲,等.黄芪甲苷调控肿瘤外泌体生成与分泌抑制结直肠癌转移的作用机制[J].上海中医药杂志,2023,57(6):41-49.

PU Yunzhou,LI Haoze,LI Ling,et al.Mechanism of astragaloside Ⅳ regulating exosome production and secretion and inhibiting metastasis of colorectal cancer[J].Shanghai Journal of Traditional Chinese Medicine,2023,57(6):41-49.
浦匀舟,李昊泽,李玲,等.黄芪甲苷调控肿瘤外泌体生成与分泌抑制结直肠癌转移的作用机制[J].上海中医药杂志,2023,57(6):41-49. DOI： 10.16305/j.1007-1334.2023.2303095.

PU Yunzhou,LI Haoze,LI Ling,et al.Mechanism of astragaloside Ⅳ regulating exosome production and secretion and inhibiting metastasis of colorectal cancer[J].Shanghai Journal of Traditional Chinese Medicine,2023,57(6):41-49. DOI： 10.16305/j.1007-1334.2023.2303095.

摘要

目的,2,从调控外泌体分泌角度，探讨黄芪甲苷（ASⅣ）抑制结直肠癌转移的作用及机制。,方法,2,①细胞实验：使用低、中、高剂量ASⅣ干预MC38细胞，收集细胞上清提取外泌体；透射电镜（TEM）观察外泌体形态；纳米颗粒跟踪分析仪检测外泌体密度；免疫荧光法和实时荧光定量逆转录聚合酶链式反应（RT-qPCR）法检测ASⅣ对外泌体生成、分泌相关基因中性鞘磷脂酶-2（,nSMase2,）、Ras相关蛋白27a（,Rab27a,）表达的影响；划痕和Transwell实验检测ASⅣ对肿瘤细胞迁移能力的影响。②动物实验：构建小鼠结直肠癌肝转移模型，随机分为模型组、ASⅣ组和鞘磷脂酶抑制剂（GW4869）组，28 d后处死小鼠，观察小鼠肝转移情况。苏木素-伊红（HE）染色法观察肝组织病理学变化，免疫荧光法检测肝转移灶内nSMase2、Rab27a的表达。,结果,2,①细胞实验结果显示：与对照组比较，ASⅣ干预不改变外泌体形态，但能够通过浓度和时间依赖方式抑制MC38细胞外泌体分泌（,P,<,0.05）；与对照组比较，ASⅣ能够减少MC38细胞中,nSMase2,、,Rab27a,的mRNA和蛋白表达水平（,P,<,0.05），且呈浓度依赖性；与对照组比较，ASⅣ能够抑制MC38细胞侵袭迁移能力（,P,<,0.05），且呈浓度依赖性。②动物实验结果显示：与模型组比较，ASⅣ能够显著降低小鼠结直肠癌肝转移模型肝组织质量（,P,<,0.05），减少肝转移瘤数量（,P,<,0.05），降低nSMase2、Rab27a蛋白表达（,P,<,0.05）；且ASⅣ抑制结直肠癌肝转移的作用与外泌体生成抑制剂GW4869相当（,P,>,0.05）。,结论,2,ASⅣ能够调控肿瘤外泌体生成与分泌，进而抑制结直肠癌肝转移。

Abstract

Objective,2,To investigate the effect and mechanism of Astragaloside Ⅳ （ASⅣ） in inhibiting colorectal cancer metastasis through regulating exosome secretion.,Methods,2,①Cell experiments： MC38 cells were treated with ASⅣ at low， medium and high doses， and cell supernatants were collected to extract exosomes. The morphology of exosomes was observed by transmission electron microscopy （TEM）. Nanoparticle tracking analyzer was used to detect exosome density. Immunofluorescence assay and RT-qPCR were used to detect the effect of ASⅣ on the expression of ,nSMase2 ,and ,Rab27a,， which were genes related to exosome production and secretion. The effect of ASⅣ on tumor cell migration was detected by cell scrath assay and Transwell assay. ②Animal experiments： A mouse model of colorectal cancer liver metastasis was established. The mice were randomly divided into model group， ASⅣ group and GW4869 group， and were sacrificed after 28 d to observe the liver metastasis. HE staining was used to observe the histopathological changes in the liver tissue， and immunofluorescence was used to detect the expression of nSMase2 and Rab27a in the liver metastases.,Results,2,①Cell experiments showed that compared with the condition in control group， ASⅣ intervention did not change the morphology of exosomes but could inhibit exosome secretion from MC38 cells in a concentration- and time-dependent manner （,P,<,0.05）； ASⅣ could reduce the mRNA and protein expression levels of ,nSMase2, and ,Rab27a ,in MC38 cells in a concentration-dependent manner （,P,<,0.05）； and ASⅣ could inhibit the invasion and migration of MC38 cells in a concentration-dependent manner （,P,<,0.05）. ②Animal experiments showed that compared with the condition in model group， ASⅣ significantly reduced the liver tissue mass （,P,<,0.05）， decreased the number of liver metastases （,P,<,0.05）， and inhibited the protein expression levels of nSMase2 and Rab27a （,P,<,0.05） in the mouse model of colorectal cancer liver metastasis； Moreover， the inhibitory effect of ASⅣ on liver metastasis of colorectal cancer was comparable to that of GW4869， an inhibitor of exosome production （,P,>,0.05）.,Conclusion,2,ASⅣ could inhibit colorectal cancer liver metastasis by regulating exosome production and secretion.

关键词

结直肠癌黄芪甲苷外泌体转移小鼠模型作用机制中药研究

Keywords

colorectal cancerAstragaloside Ⅳexosomemetastasismouse modelmechanism of actiontraditional Chinese herbal medicine research

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