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Digestive Diseases and Sciences

, Volume 61, Issue 4, pp 1107–1120 | Cite as

Regulatory Effects and Mechanism of Adenovirus-Mediated PTEN Gene on Hepatic Stellate Cells

  • Junyan An
  • Libo Zheng
  • Shurui Xie
  • Fengrong Yin
  • Xiaoxia Huo
  • Jian Guo
  • Xiaolan Zhang
Original Article

Abstract

Background

Tension homology deleted on chromosome ten (PTEN) is important in liver fibrosis.

Aims

The purpose of this study was to evaluate the PTEN gene effects and mechanism of action on hepatic stellate cells (HSCs).

Methods

The rat primary HSCs and human LX-2 cells were transfected by an adenovirus containing cDNA constructs encoding the wild-type PTEN (Ad-PTEN), the PTEN mutant G129E gene (Ad-G129E) and RNA interference targeting the PTEN sequence PTEN short hairpin RNA (PTEN shRNA), to up-regulate and down-regulate PTEN expression, respectively. The HSCs were assayed with a fluorescent microscope, real time PCR, Western blot, MTT, flow cytometry and Terminal-deoxynucleoitidyl transferase mediated nick end labeling. In addition, the CCl4 induced rat hepatic fibrosis model was also established to check the in vivo effects of the recombinant adenovirus with various levels of PTEN expression.

Results

The data have shown that the over-expressed PTEN gene led to reduced HSCs activation and viability, caspase-3 activity and cell cycle arrest in the G0/G1 and G2/M phases, as well as negative regulation of the PI3K/Akt and FAK/ERK signaling pathways in vitro. The over-expressed PTEN gene improved liver function, inhibited proliferation and promoted apoptosis of HSCs both in vitro and in vivo.

Conclusions

These data have shown that gene therapy using the recombinant adenovirus encoding wild-type PTEN inhibits proliferation and induces apoptosis of HSCs, which is a potential treatment option for hepatic fibrosis.

Keywords

PTEN Hepatic stellate cell Hepatic fibrosis Gene therapy 

Abbreviations

HSCs

Hepatic stellate cells

ECM

Extracellular matrix

α-SMA

Alpha-smooth muscle actin

PTEN

Tension homology deleted on chromosome ten

BDL

Bile duct ligation

shRNA

Short hairpin RNA

GFP

Green fluorescent protein

EGFP

Enhanced green fluorescent protein

H&E

Hematoxylin and eosin

MT

Masson’s trichrome

TUNEL

Terminal-deoxynucleoitidyl transferase mediated nick end labeling

FCM

Flow cytometry

FAK

Focal adhesion kinase

ERK

Extracellular signal-regulated kinase

PI3K

Phosphoinositol-3-kinase

Akt

Serine–threonine protein kinase B

ALT

Alanine aminotransferase

AST

Aspartate aminotransferase

Notes

Acknowledgments

This work was supported by the National Natural Science Foundation of China (Grant 30872513), Natural Science Foundation of Hebei Province (Grant C2010000565), and Hebei Provincial Science and Technology Department (Grant 09966108D). The authors would like to thank the foundations for their support. We appreciate Gregory X Shen for his valuable revision of written English and owe many thanks to Hong Zhang and Jinbo Guo for their photo contributions.

Compliance with ethical standards

Conflict of interest

The authors have declared no conflict of interest.

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Junyan An
    • 1
  • Libo Zheng
    • 1
  • Shurui Xie
    • 1
  • Fengrong Yin
    • 1
  • Xiaoxia Huo
    • 1
  • Jian Guo
    • 1
  • Xiaolan Zhang
    • 1
  1. 1.Department of Gastroenterology, The Second Hospital of Hebei Medical University, Hebei Key Laboratory of GastroenterologyHebei Institute of GastroenterologyShijiazhuangChina

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