Effect of p27mt gene on apoptosis of the colorectal cancer cell line Lovo.
OBJECTIVE
To construct p27mt recombinant adenovirus, transfect the colorectal cell line Lovo and observe the effects of p27mt on Lovo cell apoptosis and cell cycle inhibition.


METHODS
We constructed recombinant adenovirus containing p27mt by homologous recombination in bacteria. The colorectal cancer cell line Lovo was infected with recombinant replication-defective adenovirus Ad-p27mt, and expression of p27mt was determined by Western blotting; the inhibitory effect of p27mt on Lovo cells was detected by cytometry. Cell cycle was determined by flow cytometry. DNA fragment analysis identified the occurrence of apoptosis.


RESULTS
The recombinant adenovirus which already contained p27mt target gene was successfully constructed. When multiplicity of infection was >or= 50, the infection efficiency was 100%. After transfection of Lovo cells with Ad-p27mt the cells had high p27 expression which was identified by immunoblotting assay. PI staining and flow cytometry showed that 77.96% of colorectal cancer cells were inhibited in phase G(0)/G(1), while in the Ad-LacZ group and blank control group, 27.57% and 25.29% cells were inhibited in the same phase, respectively. DNA fragment analysis, flow cytometry and TUNEL assay demonstrated that p27mt is able to induce apoptosis in colorectal cancer cells.


CONCLUSIONS
p27mt has an obvious blocking effect on colorectal cancer cell cycle, and most cells were inhibited in phase G(0)/G(1). Therefore, p27mt can induce apoptosis in colorectal cells.
Chen
Jun
J
Department of Gastroenterology, Affiliated People's Hospital of Yunyang Medical College, Shiyan, Hubei Province, China. chenjun@medmail.com.cn
Ding
Wu-Hua
WH
Xu
Shao-Yong
SY
Wang
Jia-Ning
JN
Huang
Yong-Zhang
YZ
Deng
Chang-Sheng
CS
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000235:genetics*; D002453:Cell Cycle / Q000502:physiology; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics*; D053938:DNA Fragmentation; D006801:Humans; D047428:Protein Kinase Inhibitors / Q000378:metabolism*
D047428:Protein Kinase Inhibitors; D050760:Cyclin-Dependent Kinase Inhibitor p27