Long non-coding RNA GClnc1 promotes progression of colorectal cancer by inhibiting p53 signaling pathway.

OBJECTIVE: The aim of this study was to investigate whether long non-coding RNA (lncRNA) GClnc1 was involved in the development of colorectal cancer, and to explore its possible mechanisms. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was conducted to detect GClnc1 expression in 48 colorectal cancer tissues and normal colon tissues. The Kaplan-Meier method was used to analyze the relationship between GClnc1 expression and survival rate of patients with colorectal cancer. In addition, GClnc1 expression in colorectal cancer cell lines and normal colonic epithelial cell lines were analyzed. After knockdown and over-expression of GClnc1 in colorectal cancer cells, Cell Counting Kit-8 (CCK-8) and colony formation assay were performed to detect the viability and proliferation of cells, respectively. RNA pull-down and RNA-binding protein immunoprecipitation (RIP) were applied to examine the specific interaction between GClnc1 and p53. After over-expression of GClnc1 in colorectal cancer cells, qPCR and Western blot were performed to evaluate the expression levels of p53, p21 and BAX. Meanwhile, the Luciferase reporter gene assay was established to reveal the activity of p53 after over-expression of GClnc1. ChIP assay was applied to figure out whether GClnc1 could affect the binding ability of p53 to the promoter region of p21. After p53 or GClnc1 knock-down in colorectal cancer cells, the protein level of p53 was analyzed using Western blot. Finally, qRT-PCR, CCK-8 and colony formation assay were used to detect the levels of p21 and BAX, the viability, as well as the proliferation ability of cells, respectively. RESULTS: The expression of GClnc1 in colorectal cancer tissues was significantly higher than that of para-cancerous tissues. Meanwhile, GClnc1 expression in T3 and T4 tumors was markedly higher than that of T1 and T2. The survival analysis revealed that patients with a higher level of GClnc1 showed remarkably lower overall survival than those with lower expression of GClnc1. QRT-PCR results indicated that GClnc1 expression in colorectal cancer cells (including SW620 and HCT116) was conspicuously higher than that of normal colonic epithelial cells (NCM640). After knocking down GClnc1 in SW620 cells, the viability and proliferation abilities were conspicuously decreased. Meanwhile, the expression level of GClnc1, as well as the viability and colony formation ability of cells, were significantly increased after over-expression of GClnc1 in HCT116 cells. Subsequently, the qRT-PCR assay demonstrated that GClnc1 was mainly localized in the nucleus. RNA pull-down and RIP experiments revealed that there was a specific interaction between GClnc1 and p53. Moreover, qRT-PCR and Western blot analysis indicated that the expression level of p53 was not affected after over-expression of GClnc. However, the expressions of p21 and BAX were remarkably decreased. The Luciferase reporter gene assay revealed that GClnc1 over-expression markedly weakened the Luciferase activity of p53. Meanwhile, ChIP experiments demonstrated that GClnc1 up-regulation affected the binding condition of p53 to p21. Western blot analysis showed that knockdown of p53 reversed the increased mRNA level of p21 as well as BAX. Furthermore, p53 down-regulation significantly weakened cell viability and colony formation ability caused by knockdown of GClnc1. CONCLUSIONS: LncRNA GClnc1 was highly expressed in colorectal cancer tissues. Meanwhile, it could increase the proliferation of colorectal cancer cells by reducing the expression of p21 as well as BAX via p53 signaling pathway, thereby promoting the progression of colorectal cancer.

Intraoperative ultrasound-guided transhepatic lithotomy. A new alternative surgical procedure for the management of residual hepatic stones.

BACKGROUND: The high incidence of residual stones has been a major problem in the treatment of hepatolithiasis. Although various imaging techniques have been used to locate the stones, and many different postoperative procedures have been used as remedial modalities to remove the residual calculi, results have been far from satisfactory. OBJECTIVES: To remove obstinate hepatic stones and reduce the incidence of residual calculi. DESIGN: Prospective clinical trial. SETTING: Medical university-affiliated hospital. PATIENTS AND METHODS: Ten patients who had residual intrahepatic stones after conventional operative procedures underwent intraoperative ultrasound (IOUS)-guided transhepatic lithotomy between July 1988 and July 1995. This surgical technique includes accurately locating stones with IOUS, choosing a surgical approach path under the guidance of IOUS while avoiding critical blood vessels and uninvolved biliary tracts, dividing hepatic parenchyma to the involved biliary ducts, and removing the obstinate calculi using the real-time image of IOUS, which is able to monitor the movement of the lithotomy instruments without interruption. MAIN OUTCOME MEASURES: Clinical practical value of IOUS-guided transhepatic lithotomy in the treatment of residual hepatic stones. RESULTS: Complete removal of the stones was achieved in all 10 patients. There were no severe complications or mortality at a median follow-up of 39 months. CONCLUSIONS: IOUS-guided transhepatic lithotomy can greatly decrease the incidence of residual hepatic stones. It is accurate and safe. As a new surgical procedure, IOUS-guided transhepatic lithotomy should be an alternative modality in the management of hepatolithiasis, although the long-term benefits still need to be observed.