The Role Of NPAS2 In DLD-1 Cells And Its Prognosis Of Colorectal Cancer

Objective This study is to clarify the role of circadian gene NPAS2 in DLD-1 and explore the prognosis of colorectal cancer. We will discuss the relationship between the occurrence and evelopment of the circadian gene with colorectal cancer. In vitro experiments, the exprements will be played out in colorectal carcinoma cell lines to analysising the relationship about the abnormal expression of NPAS2 with the biological behaviors of colon cancer, including cells growth, cell cycle, apoptosis, invasion and migration.Our study will provide a basis for gene-therapy for colorectal cancer.Methods Tumor tissues with the corresponding NATs were got from 108 CRC patients. And the quantitative measurement of NPAS2 mRNA expression was performed using Quantitative reverse transcription-PCR(QRT-PCR).Using RNA interference technology, DLD-1 cells and the corresponding transfected cells were divided into SiRNA-Con group, SiRNA-1# group, SiRNA-2#group. QRT-PCR was used to detect the transfection effects of SiRNA-1# and-2# fragments. The changes of cell growth ability were observed which was after NPAS2 blocking in DLD-1 by CCK-8 and immunocytochemical analysis. After NPAS2 blocking in DLD-1, the experiences of cell cycle was dectected by using flow cytometry. Tunnel assays was used to detect the programmed cell death. The migration ability was observed by Scratch healing assays. And the invasion ability was detected using Transwell assays.Results we detected the mRNA expressions of NPAS2 in 108 CRC patients by RT-PCR, and found that NPAS2 expression was significantly down-regulated in tumor tissues than that in NATs. Clinicopathologic analysis revealed that low expression of NPAS2 was associated with the tumor size, TNM stage and tumor distance metastasis in colorectal cancer(p < 0.05). Furthermore, we effectively down-regulated NPAS2 mRNA expression by transfecting RNA interfere fragments into DLD-1 cells, and our results in vitro demonstrated that silencing NPAS2 expression could promote cell proliferation, cell invasion and increase the wound healing ability(p < 0.05). However, down-regulating NPAS2 expression did not influence the apoptotic rate in DLD-1 cells(p > 0.05).ConclusionIn colorectal cancer tissues, the expression of circadian gene NPAS2 was down regulated, and the distressed expression of NPAS2 was related with some clinicopatholofical factors, including tumor, TNM stage and tumor metastasis. In vitro experiment, our study showed that low expression of NPAS2 can make the colon cancer cells stay more phase splitting in S phase and G2/M, and reduce G0/G1 stag nation, promoting the growth of cells. At the same time, the silencing of NPAS2 expression could enhance cell migration and cell invasion ability of colorectal cancer. But there is no relationship between the cell apoptosis with the low expression of circadian gene NPAS2. By the relevant clinical and basic researches, our study demonstrated that the circadian gene NPAS2, which could act as a tumor factor, is involved in the occurrence and development of colorectal cancer. Silencing NPAS2 promotes cell growth and invasion in DLD-1 cells and correlated with poor prognosis of colorectal cancer. NPAS2, functioned as a potential tumor suppressor gene, could serve as a promising target and potential prognostic indicator for colorectal cancer.