| BACKGROUND:Colorectal cancer (CRC) is the third most common cancer worldwide. Despite advances in surgical technique, chemotherapy and radiotherapy, recurrence and metastasis are the main problem in the management of CRC. Therefore, new therapeutic strategies are needed to improve the survival of CRC patients. Id-1(Inhibitor of differentiation/DNA binding), a member of the helix-loop-helix (HLH) transcription factor family, is up-regulated in many types of human cancer and its expression levels are correlated with poor treatment outcome and shorter survival.OBJECTIVE:To detect the expression of Id-1 in colorectal carcinoma (CRC), and to evaluate its clinical significance. To investigate the changes of the drug sensitivity in Id-1 siRNA transfected SW480 cells.METHODS:1. Id-1 expression in 56 fresh CRC tissues as well as in 107 CRC patients who had accepted resection were detected by Real time PCR, Western blot and Immunohistochemistry. The relationship between Id-1 expression and clinicopathological features, prognosis was analyzed.2. The specific Id-1 small interference RNA (siRNA) was transfected into colorectal carcinoma SW480 cells using Iipofectamine2000. Three groups of cells were used in the vitro studies, including black control group (untransfected cells), negative control group (cells transfected with the non-silencing control siRNA) and Id-1 siRNA group (cells transfected with the Id-1 siRNA). The expression of Id-1 mRNA was detected by real time reverse transcription-polymerase chain reaction. The expression of Id-1 protein was measured by western blot. Drug sensitivity of the cells to 5-fluorouracil (5-FU) was analyzed with methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay and flow cytometry. Cell migration and movement capacity was assessed using scratch healing experiment.RESULTS:1. Real time PCR and Western blot showed the expression level of Id-1 were significantly higher in CRC than intestinal mucosa distant from the tumor (P<0.01). Id-1 expression was associated with Dukes stages, grade of differentiation and lymph node metastasis (P<0.05). Similarly, immunohistochemistry showed the expression level of Id-1 was significantly higher in CRC than normal intestinal mucosa (P<0.01). The overexpression of Id-1 in CRC was correlated with Dukes stages and lymph node metastasis (P<0.05). Furthermore overexpression of Id-1 was associated with poor prognosis.2. Id-1 siRNA was transfected into the SW480 cells efficiently. The mRNA and protein of Id-1 in Id-1 siRNA group were significantly decreased compared with either negative control group or black control group (P<0.01). MTT results showed that Id-1 siRNA group had higher cell inhibitory after treated with 5-FU (P<0.01). Add 5-FU (102.4mmol/L), FCM results demonstrated that the apoptosis of Id-1 siRNA group had been increased compared with negative control group and black control group (P<0.01). Add 5-FU (1.6 mmol/L), scratch healing experiment results demonstrated that the migration and movement capacity of Id-1 siRNA group had been decreased compared with negative control group and black control group (P<0.01).CONCLUSION:1. Overexpression of Id-1 is closely associated with the progression of CRC, and might be regarded as a factor of poor prognosis in CRC.2. Id-1 siRNA led to the efficient and specific inhibition of Id-1 expression in CRC SW480 cells. Silencing of Id-1 expression can increase SW480 cells sensitization to 5-FU.
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