Effect of Compound <italic style="font-style: italic">Phyllanthus urinaria</italic> Ⅱ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice

XU Yujie; QIN Wanli; LI Xiaohui; LI Changqing

doi:10.13422/j.cnki.syfjx.20230724

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Effect of Compound Phyllanthus urinaria Ⅱ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice

更新时间：2023-11-07

- Effect of Compound Phyllanthus urinaria Ⅱ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 29, Issue 23, Pages: 28-35(2023)
- 作者机构：
  
  1.广州中医药大学青蒿研究中心，广州 510405
  2.广州中医药大学中药学院，广州 510405
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20230724
  CLC： R2-0;R22;R242;R2-031;R285.5
- Published：05 December 2023，
  
  Published Online：31 January 2023，
  
  Received：08 October 2022，
- 稿件说明：
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徐玉洁,覃万莉,李小翚等.叶下珠复方Ⅱ号对肝癌Hep3B细胞裸鼠移植瘤生长和miR-122及IGF-1R表达的影响[J].中国实验方剂学杂志,2023,29(23):28-35.

XU Yujie,QIN Wanli,LI Xiaohui,et al.Effect of Compound Phyllanthus urinariaⅡ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(23):28-35.
徐玉洁,覃万莉,李小翚等.叶下珠复方Ⅱ号对肝癌Hep3B细胞裸鼠移植瘤生长和miR-122及IGF-1R表达的影响[J].中国实验方剂学杂志,2023,29(23):28-35. DOI： 10.13422/j.cnki.syfjx.20230724.

XU Yujie,QIN Wanli,LI Xiaohui,et al.Effect of Compound Phyllanthus urinariaⅡ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(23):28-35. DOI： 10.13422/j.cnki.syfjx.20230724.

摘要

目的

观察叶下珠复方Ⅱ号对Hep3B2.1-7人肝癌细胞（简称Hep3B细胞）裸鼠移植瘤生长的抑制作用并探讨其机制。

方法

建立肝癌Hep3B细胞裸鼠移植瘤模型，并随机分为模型组、叶下珠复方Ⅱ号高、低剂量组（57.5、28.75 g·kg

-1

）和5-氟尿嘧啶（5-FU）组（0.025 g·kg

-1

），每组8只。叶下珠复方Ⅱ号高、低剂量组均灌胃给药，每日1次；模型组予同体积生理盐水灌胃；5-FU组采用腹腔注射，隔日1次。给药28 d后处死取移植瘤，以免疫组化（IHC）法检测瘤体组织细胞核抗原（PCNA）表达，以脱氧核糖核酸断裂原位末端标记（TUNEL）法检测瘤体组织细胞凋亡，以实时荧光定量聚合酶链式反应（Real-time PCR）检测移植瘤组织miR-122和胰岛素样生长因子-1受体（IGF-1R）的mRNA表达，以蛋白免疫印迹法（Western blot）检测瘤体组织CCAAT增强子结合蛋白

（C/EBP

）、肝细胞核因子-4

（HNF-4

）和IGF-1R的蛋白表达。

结果

叶下珠复方Ⅱ号高、低剂量组和5-FU组的抑瘤率分别为74.90%、63.62%和64.15%，与模型组比较，5-FU组、叶下珠复方Ⅱ号各组的移植瘤瘤重均显著降低（

0.01），5-FU组、叶下珠复方Ⅱ号各组肿瘤生长体积显著降低（

0.01），5-FU组、叶下珠复方Ⅱ号各组移植瘤组织中细胞增殖PCNA阳性细胞变少、染色变浅，5-FU组、叶下珠复方Ⅱ号各组移植瘤组织凋亡细胞均明显增加（

0.05，

0.01），5-FU组、叶下珠复方Ⅱ号各组miR-122 mRNA表达均显著增加（

0.01），5-FU组、叶下珠复方Ⅱ号各组IGF-1R mRNA表达显著下调（

0.01），5-FU组、叶下珠复方Ⅱ号各组裸鼠移植瘤组织C/EBP

及HNF-4

的蛋白表达均显著上调（

0.01），叶下珠复方Ⅱ号高剂量组的IGF-1R蛋白表达明显下调（

0.05）；与叶下珠复方Ⅱ号低剂量组比较，叶下珠复方Ⅱ号高剂量组的凋亡细胞显著增多（

0.01），叶下珠复方Ⅱ号高剂量组miR-122 mRNA表达显著上调（

0.01），叶下珠复方Ⅱ号高剂量组的C/EBP

和HNF-4

蛋白表达均显著上调（

0.01）。

结论

叶下珠复方Ⅱ号对Hep3B细胞裸鼠移植瘤生长具有明显的抑制作用，作用机制可能与增强转录因子HNF-4

和C/EBP

的表达从而增强miR-122的表达以抑制其靶基因IGF-1R的表达有关。

Abstract

Objective

To investigate the inhibitory effects and mechanism of the compound

Phyllanthus urinaria

Ⅱ （CPU Ⅱ）on the growth of transplanted hepatocellular carcinoma Hep3B2.1-7 （Short for Hep3R） cells in nude mice.

Method

After the establishment of a xenograft model of hepatocellular carcinoma Hep3B cells in mice， the model mice were randomly divided into a model group， a high-dose CPU Ⅱ group （57.5 g·kg

-1

）， a low-dose CPU Ⅱ group （28.75 g·kg

-1

）， and a 5-fluorouracil （5-FU） group （0.025 g·kg

-1

）， with eight mice in each group. The mice in the high- and low-dose CPU Ⅱ groups were treated with drugs by gavage， once per day， and those in the model group were treated with the same volume of normal saline. The mice in the 5-FU group were treated by 5-FU by intraperitoneal injection， once every other day. After 28 days of administration， mice were sacrificed， and transplanted tumors were collected. Immunohistochemistry （IHC） was used to detect the expression of proliferating cell nuclear antigen （PCNA） of tumor tissues. Terminal-deoxynucleotidyl transferase-mediated nick end labeling （TUNEL） was used to detect cell apoptosis of tumor tissues. The mRNA expression of miR-122 and insulin-like growth factor 1 receptor （IGF-1R） in tumor tissues was detected by Real-time quantitative PCR （Real-time PCR）. The protein expression of CCAAT/enhancer-binding protein

（C/EBP

）， hepatocyte nuclear factor-4

（HNF-4

）， and IGF-1R in tumor tissues was detected by Western blot.

Result

The tumor suppression rates of the high- and low-dose CPU Ⅱ groups and the 5-FU group were 74.90%， 63.62%， and 64.15%， respectively. Compared with the model group， the CPU Ⅱ groups and the 5-FU group showed reduced weight （

0.01） and volume of tumors （

0.01）， decreased PCNA positive cells， shallow staining， increased apoptosis cells of transplanted tumor tissues （

0.05，

0.01）， increased expression of mRNA expression of miR-122 （

0.01）， down-regulated mRNA expression of IGF-1R （

0.01）， and up-regulated protein expression of C/EBP

and HNF-4

in nude mouse transplanted tumor tissues （

0.01）. The expression of IGF-1R protein in the high-dose CPU Ⅱ group was down-regulated （

0.05）. Compared with the low-dose CPU Ⅱ group， the high-dose CPU Ⅱ group showed increased apoptotic cells （

0.01）， up-regulated mRNA expression of miR-122 （

0.01）， and increased expression of C/EBP

and HNF-4

proteins （

0.01）.

Conclusion

CPU Ⅱ has an obvious inhibitory effect on the growth of transplanted hepatocellular carcinoma Hep3B cells in nude mice. The mechanism of action is related to enhancing the expression of transcription factors HNF-4

and C/EBP

， thereby promoting the expression of miR-122 and inhibiting the expression of its target gene IGF-1R.

关键词

叶下珠复方Ⅱ号肝癌胰岛素样生长因子1受体肝细胞核因子4αCCAAT增强子结合蛋白α

Keywords

compound Phyllanthus urinaria Ⅱ（CPU Ⅱ）liver cancerinsulin-like growth factor 1 receptorhepatocyte nuclear factor-4 αCCAAT/enhancer-binding protein α

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⁰

Effect of Compound Phyllanthus urinaria Ⅱ on Growth and Expression of miR-122 and IGF-1R of Hepatocellular Carcinoma Hep3B Cells in Nude Mice

DOI：10.13422/j.cnki.syfjx.20230724

摘要

Abstract

关键词

Keywords

references