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PUMA gene transfection increases sensitivity of pancreatic cancer cell line AsPC 1 to 5 FU induced apoptosis
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Abstract:
To investigate whether PUMA gene transfection can increase sensitivity of pancreatic cancer cells (PC) to 5 FU induced apoptosis. Methods: PUMA pCEP4 containing full length PUMA cDNA or pCEP4 was transfected into human pancreatic cancer cell line AsPC 1 by lipofectamine transfection, G418 selection was used to select positive cells. AsPC 1, AsPC 1/PUMA and AsPC 1/pCEP4 cells were separately treated with serial concentrations of 5 FU（0.01 100 μmol/L）. MTT assay was used to determine the cell survival rate in each group and IC 50 of 5 FU was calculated. TUNEL，FCM and DNA ladder observation were employed to study cell apoptosis. Western blotting was performed to detect the expression of PUMA protein. Results: The 5 FU IC 50 values of AsPC 1, AsPC 1/PUMA and AsPC 1/pCEP4 cells were (12±1.9)μmol/L,（1.6±0.4）μmol/L and (10.4±1.6) μmol/L, respectively, with the sensitivity of AsPC 1/PUMA cells increased by 7.5 folds. 5 FU induced cell apoptosis of AsPC 1 cells in a dose dependent manner, with the apoptosis of AsPC 1/PUMA cells more prominent than those of AsPC 1 and AsPC 1/pCEP4 cells. Low concentration of 5 FU (0.1 μmol/L) induced few apoptosis of AsPC 1/pCEP4 cells（\[1.14±0.28\]%) and AsPC 1 cells (\[0.9±023\]%), and induced apoptosis in AsPC 1/PUMA cells（\[6.47±1.42\]%). High concentration of 5 FU (1.0 μmol/L ) induced apoptosis in all groups, with that in AsPC 1/PUMA cells（\[34.54±9.36\]%) significantly higher than those in AsPC 1/pCEP4 cells（\[15.8±5.15\]%) and AsPC 1 cells (\[12.8±3.74\]%, both P <0.01) . FCM and electrophoresis showed the same results. Expression of PUMA protein in AsPC 1/PUMA cells was significantly higher than those in AsPC 1 and AsPC 1pCEP4 cells. Conclusion: PUMA gene transfection greatly enhances the sensitivity of AsPC 1 cells to 5 FU induced apoptosis.
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Project Supported:
Surpported by the Science Research Foundation of Public Health (No.WKJ 2004 2 001)
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