Subjects: Medicine, Pharmacy >> Preclinical Medicine submitted time 2017-12-27 Cooperative journals: 《南方医科大学学报》
Abstract： Objective To investigate the antitumor effect of lycorine on renal cell carcinoma ACHN cells and explore the possible mechanism. Methods We used flow cytometry to examine the effect of lycorine on ACHN cell cycle and apoptosis. The cell proliferation, migration and invasion were assessed with MTS assay, wound healing assay, and Transwell assay, respectively. Colony forming assay was performed, and the mRNA and protein levels of Bax, Bcl-2, survivin, caspase-3, cyclin D1 and CDK4 were measured with qRT-PCR and Western blotting. Results Lycorine obviously inhibited the proliferation of ACHN cells with an IC50 of 24.34 μmol/L. Lycorine also induced apoptosis of ACHN cells, caused cell cycle arrest at G0/G1 phase, and suppressed the colony forming ability of the cells in a dose-dependent manner. The migration and invasion of ACHN cells were significantly inhibited by 5 μmol/L lycorine. Lycorine up-regulated the mRNA levels of CDK4, Bax, caspase-3 while down-regulated the levels of survivin, Bcl-2 and Cyclin D1; the protein levels of CDK4 and Bax were increased and cyclin D1, Bcl-2 and surviving expressions were decreased, but caspase-3 expression showed no significant changes following the treatment. Conclusion Lycorine has obvious antitumor effect against ACHN cells, suggesting its value as a new therapeutic agent for renal cell carcinoma.
Subjects: Medicine, Pharmacy >> Preclinical Medicine submitted time 2017-12-07 Cooperative journals: 《南方医科大学学报》
Abstract： Objective To investigate the effect of stable knockdown of DNA methyltransferase 3b (DNMT3b) on the proliferation and apoptosis of bladder cancer cells. Methods Lentivirus expressing DNMT3b siRNA or the negative control siRNA was infected in human bladder cancer BIU-87 cells. MTT assay and flow cytometry were used to detect cell proliferation and apoptosis, respectively. The inhibitory effect of DNMT3b knockdown on xenograft tumors in nude mice was observed. Real-time PCR and Western blotting were carried out to investigate the expression level of cell apoptosis related genes. Methylation specific PCR was used to examine the methylation in the promoter region of the cell apoptosis related genes. Results The results of real-time PCR and Western blotting showed that DNMT3b mRNA and protein level were stably knocked down in BIU-87 cells. Stable DNMT3b knockdown suppressed BIU-87 cell growth and the tumor formation ability of the cells in nude mice. DNMT3b knockdown promoted the apoptosis of BIU-87 cells, increased the mRNA and protein expression of the cell growth and apoptosis related genes including DAPK, Bax and RASSF1A, and significantly decreased the methylation of these genes. Conclusion Stable DNMT3b knockdown can affect the methylation of the cell growth and apoptosis related genes to regulate their expression, which might be a possible mechanism for suppressed cell growth and enhanced apoptosis of BIU-87 cells.
Subjects: Medicine, Pharmacy >> Preclinical Medicine submitted time 2017-12-07 Cooperative journals: 《南方医科大学学报》
Abstract： Objective To investigate the role of microRNA-34a (miR-34a) in regulating the cell cycles of bladder cancer cell line J82 and explore the underlying mechanism. Methods J82 cells were transfected with a miR-34a mimic or an inhibitor to induce miR-34a overexpression or silencing. The RNA level of miR-34a in the transfected cells was detected by real-time PCR, and CD44 expressions at the mRNA and protein levels were detected by real-time PCR and Western blotting. Luciferase reporter assay was used to detect the activation of 3'UTR of CD44, and flow cytometry was performed to analyze the cell cycle changes. Results The expression level of miR-34a was significantly increased and CD44 expression significantly lowered in cells transfected with miR-34a mimic; miR-34a inhibitor transfection caused reverse effects on miR-34a and CD44 expressions. MiR-34a mimics downregulated while miR-34a inhibitor enhanced the activation of 3'UTR of CD44 with corresponding changes in the expressions of some cell cycle-related proteins. MiR-34a mimics and miR-34a inhibitor induced opposite changes in J82 cell cycle, which were partly reversed by CD44. Conclusion MiRNA-34a regulates cell cycles by targeting CD44 in human bladder carcinoma cell line J82.
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