Effect of Lentivirus-mediated uPA Silencing on the Proliferation and Apoptosis of Chondrocytes and the Expression of MMPs

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 div...

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Published inJournal of Huazhong University of Science and Technology. Medical sciences Vol. 35; no. 1; pp. 111 - 116
Main Author 史晨辉 王维山 张振东 李长俊 郭风劲 李锋 陈安民
Format Journal Article
LanguageEnglish
Published Heidelberg Huazhong University of Science and Technology 01.02.2015
Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Department of Orthopedics, The First Affiliated Hospital of Medical School, Shihezi University, Shihezi 832008, China%Department of Orthopedics, The First Affiliated Hospital of Medical School, Shihezi University, Shihezi 832008, China
Department of Orthopedic Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA%Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Metabolic Bone Diseases Unit, Division of Endocrinology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York 10032, USA%Department of Orthopedics, The First Affiliated Hospital of Medical School, Shihezi University, Shihezi 832008, China
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ISSN1672-0733
1993-1352
DOI10.1007/s11596-015-1398-1

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Summary: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.
Bibliography: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.
42-1679/R
si RNA; u PA; chondrocytes; MMPs
Chen-hui SHI , Wei-shan WANG, Zhen-dong ZHANG, Chang-jun LI, Feng-jing GUO , Feng LI, An-ming CHEN ( 1Department of Orthopedics, Tongji Hospital Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China; 2Department of Orthopedics, The First Affiliated Hospital of Medical School Shihezi University, Shihezi 832008, China ;3Metabolic Bone Diseases Unit, Division of Endocrinology, Department of Medicine, College of Physicians and Surgeons, Columbia University, New York 10032, USA; 4Department of Orthopedic Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA)
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ISSN:1672-0733
1993-1352
DOI:10.1007/s11596-015-1398-1