LncRNA HOXD-AS1在斑马鱼异种移植模型对肝细胞癌细胞增殖和迁移能力的影响*

doi:10.3969/j.issn.1672-5069.2022.04.005

摘要/Abstract

摘要： 目的探讨在斑马鱼异种移植模型(zPDX)长链非编码核糖核酸同源异形基因D簇反义核糖核酸1(lncRNA HOXD-AS1)对肝细胞癌(HCC)细胞的调控作用。方法在HepG2、Hep3B和Huh7细胞,分别敲低HOXD-AS1和微小RNA(miRNA)miR-130a-3p,检测细胞HOXD-AS1水平变化,使用CCK-8检测细胞增殖,使用Transwell法检测细胞迁移能力,建立zPDX模型。结果 HepG2、Hep3B和Huh7细胞HOXD-AS1相对水平分别为正常LO2细胞的65.6±5.7(P<0.01)、4.6±0.5(P<0.01)和23.4±1.0(P<0.001)倍;在Hep3B细胞,敲低HOXD-AS1后每视野细胞数为49.6±3.9,显著低于对照组的221.8±63.3(P<0.01);在zPDX,代表增殖的CM-DiI阳性信号为对照组的56.0±12.0%(P<0.01),代表迁移的CM-DiI阳性信号为对照组的49.0±8.9%(P<0.05);在Huh7细胞,敲低HOXD-AS1后Huh7细胞每视野细胞数为54.2±15.2,显著低于对照组的226.8±26.3(P<0.01);在zPDX,代表增殖的CM-DiI阳性信号为对照组的46.5±16.8%(P<0.01),代表迁移的CM-DiI阳性信号为对照组的41.9±10.2%(P<0.01);在Huh7细胞,下调miR-130a-3p后每视野细胞数为39.0±9.2,显著大于对照组的24.4±7.2(P<0.001);在zPDX,代表迁移的CM-DiI阳性细胞显著增加,为对照组的187.8±42.7%(P<0.05);将si1-HOXD-AS1和miR-130a-3p抑制剂共转染Huh7细胞,共转染组、si1-HOXD-AS1组和NC组每视野细胞数分别为78.2±15.3、39.3±9.5和79.4±18.3,差异显著(P<0.01);在zPDX,si1-HOXD-AS1组和共转染组CM-DiI阳性细胞分别为NC组的48.9±13.5%(P<0.05)和109.4±24.9(P>0.05)。结论本研究在体外和体内证明了HOXD-AS1/miR-130a-3p ceRNA网络对HCC细胞的影响,zPDX可用于肿瘤细胞实验研究。

关键词: HepG2, Huh7, HOXD簇反义核糖核酸1, 斑马鱼异种移植物, miR-130-3p, 迁移

Abstract: Objective The aim of this study was to investigate the effect of long non-coding RNA (lncRNA) HOXD cluster antisense RNA 1 (HOXD-AS1) on proliferation and motion of HepG2,Hep3B and Huh7 cells in zebrafish patient derived xenograft (zPDX). Methods The HOXD-AS1 and miR-130A-3P were knocked down in HepG2,Hep3B and Huh7 cells, and the HOXD-AS1 mRNA was detected. The cell proliferation was detected by CCK-8 and the cell migration was by Transwell. The zPDX was established. Results The HOXD-AS1 mRNA levels in HepG2, Hep3B and Huh7 cells were 65.6±5.7(P<0.01), 4.6±0.5(P<0.01) and 23.4±1.0(P<0.001) times than in LO2 cells; the cell counts per field in Hep3B cells with knock-down of HOXD-AS1 was 49.6±3.9, significantly decreased than 221.8±63.3(P<0.01) in the control; in zPDX, the proliferation of cells was 56.0±12.0%(P<0.01) and the migration was 49.0±8.9%(P<0.05) of control; the cell counts per field in Huh7 cells with HOXD-AS1 knock-down was 54.2±15.2, significantly decreased than 226.8±26.3(P<0.01) in the control; in zPDX, the proliferation was 46.5±16.8%(P<0.01), and the migration was 41.9±10.2%(P<0.01) of control; the cell counts per field in Huh7 cells with down-regulated miR-130a-3p was 39.0±9.2, significantly increased than 24.4±7.2(P<0.001) in the control; in zPDX, the cell migration increased greatly; the cell counts per field in Huh7 cells with si1-HOXD-AS1 and miR-130a-3p co-tranfection, si1-HOXD-AS1-transfected and control were 78.2±15.3, 39.3±9.5 and 79.4±18.3, significantly different among them (P<0.01); in zPDX, the CM-DII positive cells in si1-HOXD-AS1-transfected cells and in co-transfected cells were 48.9±13.5% (P<0.05) and 109.4±24.9% (P> 0.05) of control. Conclusion This study demonstrates the effects of HOXD-AS1 / miR-130a-3p ceRNA network on HCC cells in vitro and in vivo, and the zebrafish might be used for tumor metastasis study.

Key words: HepG2, Huh7, HOXD cluster antisense RNA 1, Zebrafish patient derived xenograft, miR-130-3p, Migration

张缨, 江龙委, 秦峰, 贾绍昌. LncRNA HOXD-AS1在斑马鱼异种移植模型对肝细胞癌细胞增殖和迁移能力的影响^*[J]. 实用肝脏病杂志, 2022, 25(4): 472-475.

Zhang Ying, Jiang Longwei, Qin Feng, et al. Improvement of proliferation and migration of HepG2,Hep3B and Huh7 cells by lncRNA HOXD-AS1 in zebrafish patient derived xenograft[J]. Journal of Practical Hepatology, 2022, 25(4): 472-475.

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LncRNA HOXD-AS1在斑马鱼异种移植模型对肝细胞癌细胞增殖和迁移能力的影响^*

Improvement of proliferation and migration of HepG2,Hep3B and Huh7 cells by lncRNA HOXD-AS1 in zebrafish patient derived xenograft

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