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Prompt Report
hTERT promoter-induced targeting expression of tumstatin in hepatocarcinoma cells and its antiangiogenic effect
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Abstract:
Objective:To observe the specific expression of tumstatin in hepatocarcinoma HepG2 cells induced by the human telomerase reverse transcriptase (hTERT) gene promoter and its antiangiogenic effect in vitro. Methods: phTERT-tumstatin, pCMV-tumstatin (positive control), phTERT-EGFP (negative control) plasmids were constructed and transfected into HepG2 and L-02 normal liver cells. The expression of EGFP was examined by fluorescence microscope. The expression of tumstatin protein in HepG2 cells was detected by Western blotting analysis; the proliferation of HepG2 cells after stably transfected with plasmids was measured by MTS assay; the effect of conditioned medium (containing tumstatin protein or not) on proliferation of human umbilical vascular endothelial cell (HUVEC) was detected by MTS assay. The effect of tumstatin protein on cellular tube structure formation of HUVEC was examined through counting the number of tube branches. Results: phTERT-tumstatin, pCMV-tumstatin, and phTERT-EGFP plasmids were successfully constructed. The specific expression of tumstatin was only observed in hepatocarcinoma HepG2, not in normal liver L-02 cells. phTERT-tumstatin and phTERT-EGFP transfection did not affect the proliferation of HepG2 cells; conditioned medium (CM) containing tumstatin protein (CM-T) inhibited the proliferation of HUVEC cells, with the inhibition rate being (56.49±0.33)%. The cellular tube structure formation of HUVEC cells on matrigel-coated plates supplemented with CM-T was significantly inhibited compared with conditioned medium CM-N and CM-NT (\[3.33±1.53\]% vs \[24.44±3.11\]%，\[23.94±2.92\]%, P<0.01). Conclusion: phTERT gene promoter can induce targeting expression of tumstatin in hepatocarcinoma cells and inhibit cellular tube structure formation of HUVEC cells.
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Project supported by the National Natural Science Foundation of China (No. 30672405)
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