Abstract
This study aimed to investigate expression of the proto-oncogene POK erythroid myeloid ontogenic factor (Pokemon) in colorectal cancer (CRC), and assess inhibitory effects of a small interference RNA (siRNA) expression vector in SW480 and SW620 cells. Semi-quantitative reverse transcription-polymerase chain reaction (PCR) and immunohistochemistry were performed to determine mRNA and protein expression levels of Pokemon in CRC tissues. Indirect immunofluorescence staining was applied to investigate the location of Pokemon in SW480 and SW620 cells. The siRNA expression vectors that were constructed to express a short hairpin RNA against Pokemon were transfected to the SW480 and SW620 cells with a liposome. Expression levels of Pokemon mRNA and protein were examined by real-time quantitative-fluorescent PCR and western blot analysis. The effects of Pokemon silencing on proliferation of SW480 and SW620 cells were evaluated with reference to growth curves with MTT assays. The mRNA expression level of Pokemon in tumor tissues (0.845 ± 0.344) was significantly higher than that in adjacent tumor specimens (0.321 ± 0.197). The positive expression ratio of Pokemon protein in CRC (87.0%) was significantly higher than that in the adjacent tissues (19.6%). Strong fluorescence staining of Pokemon protein was observed in the cytoplasm of the SW480 and SW620 cells. The inhibition ratios of Pokemon mRNA and protein in the SW480 cells were 83.1% and 73.5% at 48 and 72 h, respectively, compared with those of the negative control cells with the siRNA. In the SW620 cells, the inhibition ratios of Pokemon mRNA and protein were 76.3% and 68.7% at 48 and 72 h, respectively. MTT showed that Pokemon gene silencing inhibited the proliferation of SW480 and SW620 cells. Overexpression of Pokemon in CRC may have a function in carcinogenesis and progression. siRNA expression vectors could effectively inhibit mRNA and protein expression of Pokemon in SW480 and SW620 cells, thereby reducing malignant cell proliferation.
Highlights
Colorectal cancer (CRC), one of the leading causes of cancer mortality in industrialized areas and second to lung cancer, accounts for nearly one million new cases worldwide and half a million deaths each year (Bolocan et al, 2012)
Several studies demonstrated that the POK erythroid myeloid ontogenic factor (Pokemon) gene is an important proto-oncogene extensively present in human cancers, including breast, lung, colon, prostate, bladder, and hepatocellular carcinomas (HCCs) (Maeda et al, 2005a, b; Zhao et al, 2008; Aggarwal et al, 2010; Aggarwal et al, 2011a)
Transgenic mouse models confirmed that the overexpression of Pokemon can cause malignant lymphoma and the deficiency of Pokemon genes suppresses the proliferation of liver cancer cells in vitro (Maeda et al, 2005a; Lin et al, 2012; Liu et al, 2012)
Summary
Colorectal cancer (CRC), one of the leading causes of cancer mortality in industrialized areas and second to lung cancer, accounts for nearly one million new cases worldwide and half a million deaths each year (Bolocan et al, 2012). Understanding the molecular basis of CRC is important for proper management of this disease. CRC is a common malignant tumor with complex, heterogeneous genetic and biochemical backgrounds (Remo et al, 2012). The carcinogenesis and development of CRC involve complex processes with multiple factors and stages. The overexpression of certain oncogenes and/or loss of certain anti-oncogenes may be involved in tumorigenesis of CRC. The detailed molecular mechanisms of oncogenes and anti-oncogenes related to CRC remain unclear
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