ObjectiveTo investigate effect of Notch pathway regulating by inhibiting expression of forkhead box protein A1 (FOXA1) on proliferation and invasion of colon cancer SW480 cells. MethodsThe colon cancer tissues and their corresponding paracancerous tissues of 45 patients with colon cancer admitted to the First Affiliated Hospital of Henan University of Science and Technology from June 2019 to February 2021 were selected. The immunohistochemistry and real-time fluorescent quantitative PCR (qRT-PCR) methods were used to detect the expressions of FOXA1 protein and mRNA in the tissues, respectively. In addition, SW480 cells were divided into control group (untreated), shRNA-NC group (transfected with shRNA-NC), sh-FOXA1 group (transfected with sh-FOXA1), sh-FOXA1+sodium valproate group (Add 8 mmol/L Notch pathway activator sodium valproate after transfection with sh-FOXA1). Then the qRT-PCR, MTT, clone formation test, and Transwell methods were used to detect the expressions of FOXA1 mRNA, proliferation, clonogenic ability, invasion and migration of cells in each group. Western blot method was used to detect the proliferation (c-Myc, cyclinD1), invasion and migration [matrix metalloproteinase (MMP）9, MMP2], epithelial-mesenchymal transition (Vimentin, N-cadherin, E-cadherin) and Notch pathway (Notch-1, Hes-1) related protein expressions of cells in each group. Results① In the clinical cases, the expression levels of FOXA1 protein and mRNA in the colon cancer tissues were higher than those in the corresponding paracancerous tissues (protein: 0.085±0.028 vs. 0.034±0.010, t=11.036, P<0.001; mRNA: 1.62±0.34 vs. 1.00±0.09, t=11.671, P<0.001). ② In the cell experiment, compared with the control group and shRNA-NC group, the cell survival rate, and numbers of cloned cells, invasion and migrating cells were significantly reduced (P<0.05), correspondingly, the related proteins expression levels of c-Myc, cyclinD1, MMP9, MMP2, Vimentin, N-cadherin, Notch-1, Hes-1 were significantly reduced (P<0.05) and the protein expression level of E-cadherin was significantly increased (P<0.05) in the sh-FOXA1 group, which were reversed after adding the Notch pathway activator sodium valproate (P<0.05). ConclusionFOXA1 highly expresses in colon cancer tissues and colon cancer cells and it might promote the proliferation, invasion and migration of SW480 cells by activating the Notch pathway.
ObjectiveTo summarize the diagnosis and treatment of a primary gastric colon cancer, and to explore its safety and feasibility.MethodThe clinical data of a patient with gastric cancer and sigmoid colon cancer who admitted to The Affiliated Yantai Yuding Hospital of Qingdao University in October 2017 was analyzed.ResultsThe patient underwent laparoscopic radical gastrectomy plus π anastomosis and laparoscopic radical resection of colon cancer. The operation time was 330 min and the intraoperative blood loss was 120 mL. There were no complications such as stomach cramps and sputum after operation and he was successfully discharged on the 9th day after surgery. Postoperative pathological staging: gastric cancer (pT3N3M0, ⅢB) and sigmoid colon cancer (pT2N0M0, Ⅰ B).ConclusionsMultiple primary cancer of the simultaneous gastric colon should be diagnosed before operation. Laparoscopic minimally invasive treatment for gastric cancer with sigmoid colon cancer is safe and feasible, and can benefit patients.
ObjectiveTo study the effects of ATP citrate lyase (ACLY) gene on proliferation, apoptosis, invasion, and lipid metabolism of colon cancer cells.MethodsColon cancer cells HCT116 were transfected with lentiviral knockdown ACLY gene in vitro and divided into three groups according to cell treatment: HCT116 cells with ACLY gene knockdown as knockdown group, empty vector transfected cells as negative control group, and untreated colon cancer HCT116 cells as blank control group. After the stable new cell line was screened with puromycin, the expression of ACLY protein was detected by Western blot method, the lipid production of cells was detected by triglyceride test kit, the proliferation ability of cells was detected by CCK-8 method, the apoptosis rate was detected by flow cytometry, and the migration ability of cells was detected by cell scratch test.ResultsThe cell survival rate of the knockdown group was lower than those of the blank control group and the negative control group at 120 h, but there was no significant difference among the three groups at 24 h and 48 h. Compared with the negative control group and the blank control group, the apoptosis rate in the knockdown group increased, the 24 h migration ability and the level of intracellular triglyceride decreased.ConclusionACLY gene knockdown can inhibit the proliferation, apoptosis, and migration of colon cancer cells, and its mechanism may be related to the decrease of lipid synthesis ability of colon cancer cells.
Objective To investigate the expression of Bloom syndrome (BLM) helicase and tumor infiltrating dendritic cells (TIDC) in colon cancer tissues and their relationship with the prognosis of patients after surgery. Methods Onehundred and sixty-eight patients with colon cancer who underwent surgical resection in our hospital from June 2014 to August 2016 were selected as the research objects. The specimens of surgically resected colon cancer tissues and adjacent tissues archived by the pathology department were obtained, and the expression of BLM helicase and TIDC density were detected by immunohistochemistry. Pearson was used to analyze the correlation between BLM helicase expression and TIDC density, and the relationship between the expression of BLM helicase and TIDC density and the clinicopathological features of colon cancer was analyzed by using χ2 test or Wilcoxon rank test. The influencing factors of postoperative survival of patients with colon cancer were analyzed by Cox proportional hazards regression model. Results The relative expression of BLM helicase in colon cancer tissues was higher than that in adjacent tissues (1.49±0.33 vs. 1.02±0.17), while the TIDC density was lower than that in adjacent tissues [(9.53±2.36)% vs. (12.36±2.37)%], the differences were statistically significant (P<0.05). Pearson correlation analysis showed that there was a negative correlation between the expression of BLM helicase and TIDC density (r=–0.588, P<0.05). The expression of BLM helicase and TIDC density were correlated with tumor differentiation, clinical stage and lymph node metastasis (P<0.05). That is, those with high expression of BLM helicase and low density of TIDC had low degree of tumor differentiation, late clinical grade, and higher ratio of lymph node metastasis. Sixty-three cases died (37.5%) during the follow-up period (16–60 months, medium follow up 45 months). Log-rank analysis showed that the 5-year cumulative survival rate of the BLM helicase-low expression group was higher than that of the high expression group, and that of the TIDC-low density group was lower than that of the high density group (P<0.05). Cox regression analysis showed that the high expression of BLM helicase, low density of TIDC, low degree of tumor differentiation, late stage and lymph node metastasis were risk factors affecting the postoperative survival of patients with colon cancer (P<0.05). Conclusion The abnormal expression of BLM helicase and TIDC density in colon cancer tissues are related to the degree of differentiation and lymph node metastasis, which are risk factors affecting the long-term survival of patients with colon cancer.
Urokinase plasminogen activator receptor (uPAR) is a membrane protein which is attached to the cellular external membrane. The uPAR expression can be observed both in tumor cells and in tumor-associated stromal cells. Thus, in the present study, the human amino-terminal fragment (hATF), as a targeting element to uPAR, is used to conjugate to the surface of superparamagnetic iron nanoparticle (SPIO). Flowcytometry was used to examine the uPAR expression in different tumor cell lines. The specificity of hATF-SPIO was verified by Prussian blue stain and cell phantom test. The imaging properties of hATF-SPIO were confirmed in vivo magnetic resonance imaging (MRI) of uPAR-elevated colon tumor. Finally, the distribution of hATF-SPIO in tumor tissue was confirmed by pathological staining. Results showed that the three cells in which we screened, presented different expression characteristics, i.e., Hela cells strongly expressed uPAR, HT29 cells moderately expressed uPAR, but Lovo cells didn't express uPAR. In vitro, after incubating with Hela cells, hATF-SPIO could specifically combined to and be subsequently internalized by uPAR positive cells, which could be observed via Prussian blue staining. Meanwhile T2WI signal intensity of Hela cells, after incubation with targeted probe, significantly decreased, and otherwise no obvious changes in Lovo cells both by Prussian blue staining and MRI scans. In vivo, hATF-SPIO could be systematically delivered to HT29 xenograft and accumulated in the tumor tissue which was confirmed by Prussian Blue stain compared to Lovo xenografts. Twenty-four hours after injection of targeting probe, the signal intensity of HT29 xenografts was lower than Lovo ones which was statistically significant. This targeting nanoparticles enabled not only in vitro specifically combining to uPAR positive cells but also in vivo imaging of uPAR moderately elevated colon cancer lesions.
ObjectiveTo investigate the most appropriate culture time with the action of EGF in colon cancer stem cells enrichment by suspension culture.MethodsDLD-1 cells were cultured in serum-free medium containing 20 ng/mL EGF to generate spheroid cells. The time gradient was set to 10 d, 20 d, 30 d and 40 d, the cell proportion of CD133+, CD44+ and CD133+CD44+ were confirmed by flow cytometery. The ability of self-renewal was detected by the sphere forming assay and the limited dilution assay, and the in vitro tumorigenicity of the cells was detected by the colony formation assay.ResultsIn the 30 d group, the proportion of CD133+ and CD133+ CD44+ cells were significantly higher than those in the other groups (allP<0.05), the CD44+ cell was higher than that in the 20 d group (P<0.05), but there was no significant difference with the other two groups (P>0.05). The results of the limited dilution assay and the colony formation assay, the number of spheres in the 30 d or 40 d group was the highest among the 4 groups, and there was no statistical difference between the 30 d group and 40 d group (P>0.05), with statistically significant difference between the 30 d, 10 d and 20 d groups (all P<0.05). The results of the sphere forming assay and the self-renewal ability of 30 d group was significantly higher compared with other groups (all P< 0.05).ConclusionThe cancer stem cells could be enriched more efficiently by suspension culture using 20 ng/mL EGF for 30 days.
Objective To investigate the relationship between preoperative hemoglobin, albumin, lymphocyte and platelet (HALP) score, and clinicopathologic features of colon cancer, and to analyze the predictive value of HALP score for postoperative liver metastasis. Methods The clinical data of 163 patients with colon cancer admitted to the 909th Hospital of Joint Logistic Support Force (Dongnan Hospital of Xiamen University) from January 2018 to December 2019 were retrospectively analyzed. According to the occurrence of postoperative liver metastasis, the patients were divided into metastatic group (n=35) and non-metastatic group (n=128). The correlation between preoperative HAPL score and clinicopathologic features of colon cancer was analyzed. The predictive value of HALP score for postoperative liver metastasis of colon cancer was analyzed by using receiver operating characteristic (ROC) curve. The risk factors of liver metastasis after colon cancer surgery were analyzed by using univariate and multivariate logistic analysis. Kaplan-Meier risk curve was drawn, and log-rank test was used to analyze the predictive value of different HALP score for postoperative liver metastasis. Results HALP score were decreased in patients with maximum tumor diameter ≥5 cm, preoperative carcinoembryonic antigen (CEA) ≥5 μg/L, serous membrane and extrasserous infiltration, lymph node metastasis and vascular invasion, and the difference was statistically significant (P<0.05). Multivariate logistic regression analysis showed that HALP score [OR=1.467, 95%CI (1.253, 1.718), P<0.001], maximum tumor diameter [OR=3.476, 95%CI (1.475, 5.358), P=0.013], preoperative CEA level [OR= 6.197, 95%CI (2.436, 6.248), P=0.005], and lymph node metastasis [OR=2.593, 95%CI (1.667, 6.759) , P=0.003] were risk factors for postoperative liver metastasis of colon cancer. ROC curve analysis showed that the area under the curve of HALP score for predicting liver metastasis after colon cancer surgery was 0.908 (0.841, 0.974), the maximum value of the Youden index was 0.738, the optimal cut-off value of the HALP score was 35.5, the sensitivity was 0.852, the specificity was 0.886. Kaplan-Meier risk curve showed that the risk of early postoperative liver metastasis in the low HALP score group was higher than that in the high HALP score group (χ2=8.126, P=0.004). Conclusion Low HALP score in patients with colon cancer is associated with adverse prognosisi related pathological features, and is an influential factor for postoperative liver metastasis of colon cancer, and has predictive value for patients with postoperative liver metastasis of colon cancer.
ObjectiveTo understand the function of long non-coding RNA (lncRNA) colon cancer associated transcript-1 (CCAT1) and summarize its relationship with gastric cancer.MethodThe published literatures on the studies of lncRNA CCAT1 function and its relationship with gastric cancer were reviewed and analyzed.ResultsThe lncRNA CCAT1 exerted the negative regulation on the genes by binding to microRNAs (miR) as a competitive endogenous RNA, mediating chromatin circulation between the c-MYC promoter and its upstream enhancer, and promoted the expression of c-MYC gene. The recent studies had found that the CCAT1 could bind to the miR-219-1 and miR-490, thereby promoting the progress of gastric cancer. The expression of lncRNA CCAT1 in the gastric cancer tissues increased, which was obviously different from that in the paracancer tissues and normal tissues. The high expression of lncRNA CCAT1 was related to the tumor size, lymphatic metastasis and TNM stage.ConclusionsThe specific mechanism, intracellular signal transduction pathway and interaction mechanism between CCAT1 and other molecules involved in the progress of gastric cancer still need to be further explored. With the in-depth study of lncRNA, especially CCAT1, it may provide a broader prospect for the diagnosis and treatment of gastric cancer as a target of CCAT1.
Objective To investigate the inhibition effect of silence of a disintegrin and metalloproteinase 17 (ADAM17) gene on proliferation and apoptosis of HT29 colon cancer cells and its possible mechanism. Methods HT29cells were divided into 3 groups: cells of interference group were transfected with recombinant lentivirus vector, cells of negative control group were transfected with negative recombinant lentivirus vector, and cells of blank control group were treated with PBS. The expression of ADAM17 mRNA was detected by real-time PCR, the expressions of ADAM17 protein, caspase3, protein kinase B (Akt), glycogen synthase kinase-3β (GSK3β), phospho-protein kinase B (P-Akt), phospho-glycogen synthase kinase-3β (P-GSK3β) protein were detected by Western blot method, the cell proliferation was detected by MTT assay, and the apoptosis rate was detected by Annexin V-FITC/PI cell death detection kit. Results Compared with the control group and the negative control group, the interference group was related to low expressions of ADAM17 mRNA and its protein, low optical density value at the same time point (24, 48, and 72 h), high apoptosis rate, high expression level of caspase3 protein, but low expression levels of P-Akt and P-GSK3β protein (P<0.05). Conclusion Silent ADAM17 gene could significantly induces apoptosis and inhibits the proliferation of HT29 cells, which maybe via inhibiting Akt/GSK3β signaling pathway.
ObjectiveTo investigate the regulatory role of DAB2IP in proliferation effect of colon cancer cells by salinomycin. MethodsCell counting kit 8 (CCK8) assay was used to detect median inhibitory concentration (IC50) of salinomycin on HT29 and SW480 cells. Colon cancer cells with stable knock-down of DAB2IP (HT29-shDAB2IP) and control cells (HT29-shcon) were constructed by lentivirus plasmid. And colon cancer cells with stable over-expression of DAB2IP (SW480-DAB2IP) and control cells (SW480-con) were constructed using pCI plasmid. The proliferation effect of salinomycin on stable knock-down or over-expression of DAB2IP by CCK8 assay in the colon cancer cell was identified. The colon cancer stem cells makers CD44, CD24, and CD133 were investigated using real-time PCR. ResultsThe salinomycin had obvious inhibitory effects on the proliferations of HT29 and SW480 cells, the IC50 value was 20.0 μmol/L and 10.0 μmol/L, respectively. The stable knock-down of DAB2IP could significantly enhance the inhibitory effect of salinomycin on the proliferation in HT29 cells (P<0.05) and stable over-expression of DAB2IP could significantly decrease the inhibitory effect of salinomycin on the proliferation in SW480 cells (P<0.05). Further the result of real-time PCR detection showed that the expressions of cancer stem cells markers CD44, D24, and CD133 were significantly increased after stable knock-down of DAB2IP in the HT29 cells (P<0.05), while the expressions were significantly decreased after stable over-expression of DAB2IP in the SW480 cells (P<0.05). ConclusionsFrom initial results of this study, salinomycin could suppress proliferation of colon cancer cells. DAB2IP might weaken proliferative inhibitory effect of salinomycin by inhibiting expressions of cancer cells stem in colon cancer cells.