Effect of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells and its molecular mechanism

Qingsheng FU; Lei JIN; Xudong ZHANG; Yingchen XU; Chunfu ZHU; Xihu QIN; Baoqiang WU

doi:10.19401/j.cnki.1007-3639.2023.03.007

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Effect of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells and its molecular mechanism

Article | 更新时间：2025-12-31

- Effect of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells and its molecular mechanism
- China Oncology Vol. 33, Issue 3, Pages: 241-249(2023)
- 作者机构：
  
  1. 蚌埠医学院研究生院，安徽蚌埠 233030
  2. 常州市第二人民医院肝胆胰外科，江苏常州 213003
- 作者简介：
- 基金信息：
- DOI：10.19401/j.cnki.1007-3639.2023.03.007
  CLC： R735.9
- Received：19 September 2022，
  
  Revised：2022-12-14，
  
  Published：30 March 2023
- 稿件说明：
移动端阅览
Qingsheng FU, Lei JIN, Xudong ZHANG, et al. Effect of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells and its molecular mechanism[J]. China Oncology, 2023, 33(3): 241-249.
DOI：

Qingsheng FU, Lei JIN, Xudong ZHANG, et al. Effect of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells and its molecular mechanism[J]. China Oncology, 2023, 33(3): 241-249. DOI： 10.19401/j.cnki.1007-3639.2023.03.007.

摘要

背景与目的：

tRNA衍生的片段（tRNA-derived fragments，tRF）是一类长度为14~30 nt的小分子非编码RNA，其影响着恶性肿瘤的发展进程。本研究旨在探讨tRF-Pro-CGG对小鼠胰腺癌细胞生物学行为的影响及其可能的分子机制。

方法：

采用实时荧光定量聚合酶链反应（real-time fluorescence quantitative polymerase chain reaction，RTFQ-PCR）检测tRF-Pro-CGG在小鼠胰腺癌细胞系pan02、LTPA，人胰腺癌细胞系Capan-2和正常胰腺细胞HPDE6-C7中的表达水平。通过慢病毒转染技术过表达pan02细胞及敲低LTPA细胞中tRF-Pro-CGG的表达，采用RTFQ-PCR和蛋白质印迹法（Western blot）检测过表达和敲低效果。采用细胞计数试剂盒-8（cell counting kit-8，CCK-8）检测细胞增殖情况。采用transwell实验检测细胞迁移和侵袭能力。采用动物模型检测tRF-Pro-CGG对胰腺癌裸鼠移植瘤生长和转移的影响。采用H-E染色观察移植瘤的组织病理学结构。采用Western blot检测移植瘤组织中Ki-67增殖指数、转移相关蛋白E-钙黏蛋白（E-cadherin）、波形蛋白（vimentin）的表达及磷脂酰肌醇3-激酶（phosphoinositide 3-kinase，PI3K）/蛋白激酶B（protein kinase B，AKT）信号转导通路蛋白的表达及磷酸化情况。

结果：

小鼠胰腺癌细胞系pan02中tRF-Pro-CGG表达最低，在pan02细胞中转染tRF-Pro-CGG mimics后，tRF-Pro-CGG的mRNA和蛋白表达水平均显著升高（

0.01），细胞增殖能力显著降低（

0.01），细胞迁移（

0.001）和侵袭能力（

0.001）显著降低，裸鼠移植瘤体积（

0.01）和重量（

0.001）均显著降低，裸鼠移植瘤组织中出现明显坏死和凋亡细胞；裸鼠移植瘤组织中Ki-67增殖指数和vimentin的表达显著降低（

0.001），而E-cadherin的表达显著升高（

0.001），PI3K、P-PI3K、AKT和P-AKT的表达显著降低（

0.001），胰腺癌肝转移例数差异无统计学意义（

0.05）。小鼠胰腺癌细胞系LTPA中tRF-Pro-CGG表达最高，在LTPA细胞中转染tRF-Pro-CGG inhibitor后，tRF-Pro-CGG的mRNA和蛋白表达水平显著降低（

0.01），细胞增殖能力显著升高（

0.01），细胞迁移（

0.001）和侵袭能力（

0.001）显著升高，裸鼠移植瘤体积（

0.01）和重量（

0.01）均显著升高，裸鼠移植瘤组织中出现少量的坏死及凋亡细胞；裸鼠移植瘤组织中Ki-67和vimentin的表达显著升高（

0.001），而E-cadherin的表达显著降低（

0.001），PI3K、P-PI3K、AKT和P-AKT的表达显著升高（

0.001），胰腺癌肝转移例数差异无统计学意义（

0.05）。

结论：

过表达tRF-Pro-CGG可以抑制小鼠胰腺癌细胞增殖、迁移和侵袭，抑制胰腺癌裸鼠移植瘤的生长，下调Ki-67增殖指数、vimentin的表达水平和PI3K/AKT磷酸化水平。tRF-Pro-CGG可能通过调节PI3K/AKT信号转导通路抑制胰腺癌的发生、发展。

Abstract

Background and purpose:

tRNA-derived fragments (tRF) are a kind of short non-coding RNA (14-30 nt) that influences the course of cancer. This study aimed to investigate the molecular pathways that might underlie the effects of tRF-Pro-CGG on the biological behavior of mouse pancreatic cancer cells.

Methods:

Real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR) was used to assess the expression of tRF-Pro-CGG in mouse pancreatic cancer cell lines pan02 and LTPA

human pancreatic cancer cell line Capan-2

and normal pancreatic cells HPDE6-C7. tRF-Pro-CGG overexpression in pan02 cells and LTPA cell suppression were achieved through lentiviral transfection

and RTFQ-PCR and Western blot were used to determine overexpression and knockdown effects. Cell counting kit-8 (CCK-8) was used to detect cell proliferation. Transwell assays were used to detect cell migration and invasion ability. The effect of tRF-Pro-CGG on the growth and metastasis of pancreatic cancer transplantation tumors in nude mice model was investigated. H-E staining was used to observe the histopathological structure of transplantation tumors. Western blot was used to detect the expression and phosphorylation of proliferation-related protein Ki-67 and metastasis-related proteins. Western blot was used to assess the expressions of cadherin

vimentin

phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) pathway protein and phosphorylation in transplanted tumor tiss

ues.

Results:

tRF-Pro-CGG expression was lowest in the mouse pancreatic cancer cell line pan02. Both mRNA and protein expression levels of tRF-Pro-CGG were significantly increased (

0.01) after transfection of tRF-Pro-CGG mimics in pan02 cells

and cell proliferation ability (

0.01)

cell migration (

0.001) and invasion ability (

0.001) were significantly reduced. A significant decrease in the volume (

0.01) and weight (

0.001) of transplanted tumors in nude mice was observed

and significant necrotic and apoptotic cells in transplanted tumor were identified. In transplanted tumor tissues of nude mice

the Ki-67 proliferatien index and expression of vimentin were significantly decreased (

0.001)

while E-cadherin was increased (

0.001). The expressions of PI3K

P-PI3K

AKT and P-AKT were significantly decreased (

0.001). There was no significant difference in the number of liver metastases from pancreatic cancer (

0.05). The mouse pancreatic cancer cell line LTPA had the greatest level of tRF-Pro-CGG expression. The mRNA and protein expression levels of tRF-Pro-CGG were significantly reduced (

0.01) after transfection of tRF-Pro-CGG inhibitor in LTPA cells. The proliferation ability of cells was significantly increased (

0.01)

the migration of cells (

0.001) and invasive ability (

0.001) were significantly increased. The volume (

0.01) and weight (

0.01) of transplanted tumors in nude mice were significantly increased

and a limited proportion of necrotic and apoptotic cells were seen in nude mice tumor tissues implanted. In the transplanted tumor tissues of nude mice

the Ki-67 proliferation index and expression of vimentin were significantly increased (

0.001)

while E-cadherin was decreased (

0.001). The expressions of PI3K

P-PI3K

AKT

and P-AKT were significantly increased (

0.001). There was no difference in the number of liver metastases from pancreatic cancer (

0.05).

Conclusion:

Overexpression of tRF-Pro-CGG reduced pancreatic cancer cell proliferation

migration and invasion in mice

slowed the formation of pancreatic cancer transplanted tumors in nude mice

and decreased Ki-67 proliferation index and expression of vimentin and PI3K/AKT phosphorylation levels. The PI3K/AKT signaling pathway may be regulated by tRF-Pro-CGG

which may suppress the development of pancreatic cancer.

关键词

Keywords

references

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