ABSTRACT
MicroRNAs have been implicated in the regulation of several cellular signaling pathways of colorectal cancer (CRC) cells. Although emerging evidence proves that microRNA (miR)-106a is expressed highly in primary tumor and stool samples of CRC patients; whether or not miR-106a mediates cancer metastasis is unknown. We show here that miR-106a is highly expressed in metastatic CRC cells, and regulates cancer cell migration and invasion positively in vitro and in vivo. These phenotypes do not involve confounding influences on cancer cell proliferation. MiR-106a inhibits the expression of transforming growth factor-ß receptor 2 (TGFBR2), leading to increased CRC cell migration and invasion. Importantly, miR-106a expression levels in primary CRCs are correlated with clinical cancer progression. These observations indicate that miR-106a inhibits the anti-metastatic target directly and results in CRC cell migration and invasion.
Subject(s)
Cell Movement/genetics , Colorectal Neoplasms/genetics , Gene Expression Regulation, Neoplastic , MicroRNAs/genetics , Protein Serine-Threonine Kinases/genetics , Receptors, Transforming Growth Factor beta/genetics , Blotting, Western , Cell Line, Tumor , Cell Proliferation , Cell Survival/genetics , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Disease Progression , Female , HT29 Cells , Humans , Kaplan-Meier Estimate , Male , Neoplasm Invasiveness , Neoplasm Metastasis , Neoplasms, Experimental/genetics , Neoplasms, Experimental/metabolism , Neoplasms, Experimental/pathology , Protein Serine-Threonine Kinases/metabolism , RNA Interference , Receptor, Transforming Growth Factor-beta Type II , Receptors, Transforming Growth Factor beta/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Transplantation, HeterologousABSTRACT
AIM: To evaluate the effect of nigericin on colorectal cancer and to explore its possible mechanism. METHODS: The human colorectal cancer (CRC) cell lines HT29 and SW480 were treated with nigericin or oxaliplatin under the conditions specified. Cell viability assay and invasion and metastasis assay were performed to evaluate the effect of nigericin on CRC cells. Sphere-forming assay and soft agar colony-forming assay were implemented to assess the action of nigericin on the cancer stem cell properties of CRC cells undergone epithelial-mesenchymal transition (EMT). RESULTS: Compared with oxaliplatin, nigericin showed more toxicity for the HT29 cell line (IC50, 12.92 ± 0.25 µmol vs 37.68 ± 0.34 µmol). A similar result was also obtained with the SW116 cell line (IC50, 15.86 ± 0.18 µmol vs 41.02 ± 0.23 µmol). A Boyden chamber assay indicated that a significant decrease in the number of HT29 cells migrating through polyvinylidene fluoride membrane was observed in the nigericin-treated group, relative to the vehicle-treated group [11 ± 2 cells per high-power field (HPF) vs 19.33 ± 1.52 cells per HPF, P < 0.05]. Compared to the control group, the numbers of HT29 cells invading through the Matrigel-coated membrane also decreased in the nigericin-treated group (6.66 ± 1.52 cells per HPF vs 14.66 ± 1.52 cells per HPF, P < 0.05). Nigericin also reduced the proportion of CD133+ cells from 83.57% to 63.93%, relative to the control group (P < 0.05). Nigericin decreased the number of spheres relative to the control group (0.14 ± 0.01 vs 0.35 ± 0.01, P < 0.05), while oxaliplatin increased the number of spheres relative to the control group (0.75 ± 0.02 vs 0.35 ± 0.01; P < 0.05). Nigericin also showed a decreased ability to form colonies under anchorage-independent conditions in a standard soft agar assay after 14 d in culture, relative to the control group (1.66 ± 0.57 vs 7 ± 1.15, P < 0.05), whereas the colony numbers were higher in the oxaliplatin group relative to the vehicle-treated controls (14.33 ± 0.57 vs 7 ± 1.15, P < 0.05). We further detected the expression of E-cadherin and vimentin in cells treated with nigericin and oxaliplatin. The results showed that HT29 cells treated with nigericin induced an increase in E-cadherin expression and a decrease in the vimentin expression relative to vehicle controls. In contrast, oxaliplatin downregulated the expression of E-cadherin and upregulated the expression of vimentin in HT29 cells relative to vehicle controls. CONCLUSION: This study demonstrated that nigericin could partly reverse the EMT process during cell invasion and metastasis.
Subject(s)
Colorectal Neoplasms/drug therapy , Epithelial-Mesenchymal Transition/drug effects , Gene Expression Regulation, Neoplastic , Nigericin/pharmacology , AC133 Antigen , Anti-Bacterial Agents/pharmacology , Antigens, CD/metabolism , Cadherins/metabolism , Cell Line, Tumor , Cell Movement , Cell Survival , Flow Cytometry/methods , Glycoproteins/metabolism , Humans , Neoplasm Invasiveness , Neoplasm Metastasis , Neoplastic Stem Cells/cytology , Organoplatinum Compounds/pharmacology , Oxaliplatin , Peptides/metabolism , Vimentin/metabolismABSTRACT
AIM: To elucidate the effects of hyperthermic CO2 pneumoperitoneum on human gastric AGS cells. METHODS: Based on a newly devised in vitro study model, we evaluated the anti-cancer effects of HT-CO2 (42-44 degrees C for 2-4h) on human gastric cancer cells, and also the corresponding mechanisms. RESULTS: HT-CO2 (42-44 degrees C for 2-4h) severely inhibited cell proliferation as assessed by Cell Counting Kit-8 assay, while inducing apoptosis in a temperature- and time-dependent manner demonstrated by annexin-V/PI flow cytometry and morphological analysis (Hoechst/PI fluorescence). In addition, it was found that HT-CO2 (42-44 degrees C for 2-4h) promoted the up-regulation of Bax by western blotting. Significantly, it could also suppress gastric cancer cell invasion and metastasis by in vitro invasion and motility assay. CONCLUSION: In conclusion, HT-CO2 had an efficacious cytotoxic effect on gastric cancer cells through Bax-induced mitochondrial apoptotic signaling. Our studies indicate that it may serve as a potential therapy for peritoneal carcinomatosis of gastric cancer. Further investigations in vivo using animal models are now urgently needed.
Subject(s)
Adenocarcinoma/prevention & control , Apoptosis/drug effects , Carbon Dioxide/therapeutic use , Hyperthermia, Induced , Pneumoperitoneum/prevention & control , Stomach Neoplasms/prevention & control , Adult , Blotting, Western , Cell Line, Tumor , Cell Proliferation/drug effects , Female , Flow Cytometry , Humans , Mitochondria/drug effects , Mitochondria/metabolism , Proto-Oncogene Proteins c-bcl-2/metabolism , bcl-2-Associated X ProteinABSTRACT
BACKGROUND: Cancer stem-like cells (CSCs) in colorectal cancers (CRC) may account for the failure of treatments because they are resistant to many current anticancer therapies. Salinomycin, a potassium ionophore, was recently identified as a selective inhibitor of breast CSCs. METHODS: The human CRC cell lines HT29 and SW480 were treated with salinomycin and oxaliplatin. Cell viability was determined with cell counting kit 8. Fraction of CD133+ cell subpopulations was assessed by Flow Cytometric analysis. Clonogenecity and migration were determined with soft agar and Boyden chamber assays. Molecular changes were assessed by immunofluorescence staining, RT-PCR, and Western blot analysis. RESULTS: We report that salinomycin reduces the proportion of CD133+ subpopulations in human CRC HT29 and SW480 cells. Furthermore, salinomycin treatment decreases colony-forming ability and cell motility in HT29 cells. Moreover, salinomycin downregulates the expression of vimentin and induces the E-cadherin expression in HT29 cells. CONCLUSIONS: This study demonstrates the ability of salinomycin to selectively target "CD133+" cell subpopulations and decrease the malignant traits in colorectal cancer lines.