[Construction of PTEN eukaryotic expression plasmid and its effects on breast carcinoma cell line MDA468].
OBJECTIVE
To investigate the effects of exogenous wild PTEN gene stably transfection on growth of breast cancer cells in vitro.


METHODS
At first, a recombinant eukaryotic expression plasmid pcDNA3.1-PTEN was constructed. Human breast cancer cell line MDA468 was transfected with pcDNA3.1-PTEN or mock transfected plasmid pcDNA3.1(-) with lipofectamine. RT-PCR, immunohistochemical staining and Western blot were used to determine target gene expression. Cell viability was tested by MTT assay. Apoptosis was determined by flow cytometry with a double-staining method using FITC-conjugated annexin V and PI.


RESULTS
The PTEN stably transfected cells demonstrated the integration of the exogenous target gene and corresponding mRNA and protein over-expression. There was a significant decline in cell viability of pcDNA3.1-PTEN transfected MDA468 cells in comparison with the mock-transfected ones (P < 0.01). The PTEN-trasfected MDA468 cells also showed an increase in the rate of apoptosis, compared with parental and mock-trasfected cells (P < 0.001).


CONCLUSIONS
Stable expression of exogenous PTEN can suppress the malignant phenotypes of the human breast cancer cell line MDA468.
Chen
Qing-yong
QY
Department of General Surgery, Xiehe Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China. chenqy11@hotmail.com
Chen
Dao-da
DD
Wang
Chun-you
CY
Zhou
You-sheng
YS
D001943:Breast Neoplasms / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D005057:Eukaryotic Cells / Q000378:metabolism; D006801:Humans; D051059:PTEN Phosphohydrolase / Q000096:biosynthesis / Q000235:genetics*; D010641:Phenotype; D010957:Plasmids / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D011994:Recombinant Proteins; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human