The lentivirus-mediated u PA interference in the proliferation, apoptosis, and secretion of osteoarthritic chondrocytes was examined in this study. Cells were obtained from the cartilage tissues of New Zealand white r...The lentivirus-mediated u PA interference in the proliferation, apoptosis, and secretion of osteoarthritic chondrocytes was examined in this study. Cells were obtained from the cartilage tissues of New Zealand white rabbits. They were cultured with interleukin(IL)-1β(10 ng/m L) for 24 h and then divided into three groups: u PA-si RNA group(cells transfected with u PA-si RNA lentiviruses), blank control group(untreated cells), and negative control group(cells transfected with empty vectors). Western blotting and real-time quantitative reverse transcription-PCR(RT-QPCR) were performed to detect the protein and m RNA expression levels of u PA, MMP-1, MMP-3, MMP-9, MMP-10, MMP-13 and MMP-14 in osteoarthritic chondrocytes. Cell Counting Kit-8, flow cytometry, and colony formation assay were used to examine the proliferation and apoptosis of chondrocytes. The results showed that after u PA-si RNA transfection, the protein and mR NA expression levels of uP A, MMP-1, MMP-3, MMP-9, MMP-10, MMP-13, and MMP-14 were significantly decreased(P〈0.05 for MMP-1, MMP-9, MMP-10 and MMP-14, P〈0.01 for u PA, MMP-3 and MMP-13). Cell proliferation and colony formation rate were significantly higher and the cell apoptosis rate was significantly lower in u PA-si RNA group than in control groups(P〈0.01). The proportion of cells in G0/G1 phase was markedly increased and that in the S phase decreased, and the cell cycle was arrested at the G1/S phase in the control group. In the u PAsi RNA group, the proportion of cells in the S phase was significantly increased, resulting in a different proportion of cells in cell cycle phase(P〈0.01). It was suggested that the down-regulation of uP A gene could inhibit the expression of MMPs protein and cell apoptosis, increase the proliferation and colony formation of osteoarthritic chondrocytes.展开更多
基金supported by grants from the National Natural Science Foundation of China(Nos.81160225,812604531,and 81360451)the Xinjiang Bingtuan Special Program of Medical Science(Nos.2011BC004,2013BA020,and 2014BC003)
文摘The lentivirus-mediated u PA interference in the proliferation, apoptosis, and secretion of osteoarthritic chondrocytes was examined in this study. Cells were obtained from the cartilage tissues of New Zealand white rabbits. They were cultured with interleukin(IL)-1β(10 ng/m L) for 24 h and then divided into three groups: u PA-si RNA group(cells transfected with u PA-si RNA lentiviruses), blank control group(untreated cells), and negative control group(cells transfected with empty vectors). Western blotting and real-time quantitative reverse transcription-PCR(RT-QPCR) were performed to detect the protein and m RNA expression levels of u PA, MMP-1, MMP-3, MMP-9, MMP-10, MMP-13 and MMP-14 in osteoarthritic chondrocytes. Cell Counting Kit-8, flow cytometry, and colony formation assay were used to examine the proliferation and apoptosis of chondrocytes. The results showed that after u PA-si RNA transfection, the protein and mR NA expression levels of uP A, MMP-1, MMP-3, MMP-9, MMP-10, MMP-13, and MMP-14 were significantly decreased(P〈0.05 for MMP-1, MMP-9, MMP-10 and MMP-14, P〈0.01 for u PA, MMP-3 and MMP-13). Cell proliferation and colony formation rate were significantly higher and the cell apoptosis rate was significantly lower in u PA-si RNA group than in control groups(P〈0.01). The proportion of cells in G0/G1 phase was markedly increased and that in the S phase decreased, and the cell cycle was arrested at the G1/S phase in the control group. In the u PAsi RNA group, the proportion of cells in the S phase was significantly increased, resulting in a different proportion of cells in cell cycle phase(P〈0.01). It was suggested that the down-regulation of uP A gene could inhibit the expression of MMPs protein and cell apoptosis, increase the proliferation and colony formation of osteoarthritic chondrocytes.
文摘目的探索urokinase-type plasminogen activator(u PA)合成抑制剂阿米洛利(氨氯吡咪,Amiloride)在卵巢癌细胞迁徙、侵袭过程中的作用。方法检测阿米洛利处理后卵巢癌细胞OVCAR3的迁徙及侵袭能力变化,RT-RCR及Western Blotting检测u PA表达水平变化。结果卵巢癌细胞OVCAR3经阿米洛利处理后u PA m RNA及蛋白表达水平及细胞迁徙、侵袭能力显著下降,且呈浓度依赖性。结论 u PA合成抑制剂阿米洛利可能通过下调u PA表达水平抑制卵巢癌细胞迁徙及侵袭,是一种潜在的抑制卵巢癌发展的药物。