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Cytotoxicity of polymethyl methacrylate cement on primary cultured metastatic spinal cells
Ji Fang,Jieliang Shen,Wei Jiang,Wen Dong,Zhenming Hu,Zhenming Hu 대한독성 유전단백체 학회 2016 Molecular & cellular toxicology Vol.12 No.2
Polymethyl methacrylate (PMMA) bone cement has been commonly used for percutaneous injection into collapsed vertebral bodies due to malignant tumor. The purpose of this study was to investigate the possible mechanisms of PMMA’s cytotoxcity on primary cultured spinal metastastic cells (SMCs) in vitro. PMMA specimens were prepared in standard discs made of dough and polymerization stages, and the eluates were prepared following the ISO standard. Primary SMCs were obtained and isolated from 7 patients with spinal metastatic tumors undergoing vertebroplasty. Primary cultured SMCs were treated with PMMA specimens of different stages for 24 h, or co-cultured with extracted medium for successive 3 days. The temperatures in two locations from cement discs were recorded by K-type thermocouples. Furthermore, cell proliferation, apoptosis and cycles were determined by MTT and flow cytometry, respectively. The modulation of apoptotic proteins (bcl- 2, bax, caspase-3, caspase-8, Fas and PARP) and cell cycle proteins (cyclin D1, P21 and P27) were analyzed by western blot and real time PCR. The results of this study demonstrated that PMMA treatment was able to suppress SMCs proliferation, induce cell apoptosis and inhibit cell cycle arrest when compared to the control group in vitro (P<0.05). Simultaneously, the temperature recorded at the periphery of the PMMA specimen was not high enough to casue thermal injury. Furthermore, molecular markers of apoptosis including Fas, caspase-3 and caspase-9 activated, and Bcl-2/Bax dysregulated in the SMCs with PMMA stimulation. In addition, PMMA treatment showed decreased expression of cyclin D1 that induces cell cycle and increased epxression of inhibitory protein P21, with no significant difference of P27 expression. In summary, the present study confirms that PMMA cement can compromise the vitality and apoptosis of SMCs. This cytotoxic effect may be regulated by the biomarkers Fas, Bcl-2, Bax, caspase-3, caspase-9, cyclin D1 and P21, but not on account of thermal damage.
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( Liangbo Lin ),( Quanhe Qiu ),( Nian Zhou ),( Wen Dong ),( Jieliang Shen ),( Wei Jiang ),( Ji Fang ),( Jie Hao ),( Zhenming Hu ) 생화학분자생물학회(구 한국생화학분자생물학회) 2016 BMB Reports Vol.49 No.3
Bone morphogenetic protein 9 (BMP9) is a potent inducer of osteogenic differentiation of mesenchymal stem cells. The Wnt antagonist Dickkopf-1 (Dkk1) is involved in skeletal development and bone remodeling. Here, we investigated the role of Dkk1 in BMP9-induced osteogenic differentiation of MSCs. We found that overexpression of BMP9 induced Dkk1 expression in a dose-dependent manner, which was reduced by the P38 inhibitor SB203580 but not the ERK inhibitor PD98059. Moreover, Dkk1 dramatically decreased not only BMP9-induced alkaline phosphatase (ALP) activity but also the expression of osteocalcin (OCN) and osteopontin (OPN) and matrix mineralization of C3H10T1/2 cells. Furthermore, exogenous Dkk1 expression inhibited Wnt/β-catenin signaling induced by BMP9. Our findings indicate that Dkk1 negatively regulates BMP9-induced osteogenic differentiation through inhibition of the Wnt/β-catenin pathway and it could be used to optimize the therapeutic use of BMP9 and for bone tissue engineering. [BMB Reports 2016; 49(3): 179-184]
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