Regulatory Effects and Mechanism of Adenovirus-Mediated PTEN Gene on Hepatic Stellate Cells
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Tension homology deleted on chromosome ten (PTEN) is important in liver fibrosis.
The purpose of this study was to evaluate the PTEN gene effects and mechanism of action on hepatic stellate cells (HSCs).
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.
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.
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.
KeywordsPTEN Hepatic stellate cell Hepatic fibrosis Gene therapy
Hepatic stellate cells
Alpha-smooth muscle actin
Tension homology deleted on chromosome ten
Bile duct ligation
Short hairpin RNA
Green fluorescent protein
Enhanced green fluorescent protein
Hematoxylin and eosin
Terminal-deoxynucleoitidyl transferase mediated nick end labeling
Focal adhesion kinase
Extracellular signal-regulated kinase
Serine–threonine protein kinase B
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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