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Archive > Volume 21 Issue 3 > 2014,21(3):314-319. DOI:10.3872/j.issn.1007-385X.2014.03.014 Prev Next
Basic Research
Effects of a hypoxia-induced, tumor-specific gene therapy vector on proliferation of human colon carcinoma cell line LoVo Human colon adenocarcinoma cell line
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Abstract:
Homebox transcription factor 2 (CDX2) has been described as a colorectal tumor suppressor. This study aimed to evaluate the effect of overexpression of the hypoxia response element enhancer (HRE) sequence-inserted CDX2 gene under the control of the human telomerase reverse transcriptase (hTERT) promoter on proliferation of human colon adenocarcinoma LoVo cells in vitro. Methods: Stable clones of LoVo cells expressing pLVX-hTERTp-CDX2-3FLAG (hC), pLVX-5HRE-hTERTp-3FLAG (5Hh) and pLVX-5HRE-hTERTp-CDX2-3FLAG (5HhC), respectively, were generated. LoVo cells stably expressing hC, 5Hh, and 5HhC, respectively, were cultured under hypoxia control (CoCl2 200 μmol/L) and normoxia, respectively. At the designated time points of culture, cell viability was assessed by MTT assays, colony-forming capacity by trypan blue staining and cell cycle progression by flow cytometry. Result: Viability was significantly decreased in hC- and 5HhC-expressing LoVo cells as compared with wild-type LoVo cells and 5Hh-expressing LoVo cells. The proliferation rates under normoxia and hypoxiarespectively were (48.62±3.32)% and (36.81±2.83)% at 5 days (P<0.05) and (56.44±2.28)% and (38.51±3.21)% at 7 days (P<0.05) in 5HhC-expressing cells. LoVo cells stably expressing hC and 5HhC, respectively, formed significantly less clones than cells stably expressing 5Hhunder a hypoxic condition (44.2±3.5 vs 90.8±9.3, P<0.05). The proportion of cells at G1 phase arrest was (63.59±0.55)% and (64.82±2.22)% in cells stably expressing hC and 5HhC respectively, significantly higher (P<0.05) than that in wild type LoVo cells and cells stably expression 5Hh (51.38±0.70 and 51.59±0.38) under normoxia. Under a hypoxic condition, as high as 71.38±3.02 5HhC-expressing LoVo cells were arrested at G1 phase. Conclusion: Overexpression of the CDX2 gene carrying the HRE sequence driven by the hTERT promoter may significantly inhibit proliferation and colony formation of human colon adenocarcinoma LoVo cells in vitro, through inducing G1 phase arrest. These effects were more significant under hypoxia.
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Project Supported:
Project supported by the Natural Science Foundation of China(No.81101874,No.81172362,No.81172359), the Science and Technology Foundation of Shaanxi Province(No.2011-K12-19), and the Science and Technology Co-ordinative Innovation Project of Shaanxi Province(No.2013KTCQ03-08)
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