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白藜芦醇抑制脂多糖诱导破骨前体细胞Raw 264.7的激活
Resveratrol inhibits lipopolysaccharide-induced activation of osteoclast precursor Raw 264.7 cells
目的 探讨白藜芦醇 (Res) 对脂多糖 (LPS) 诱导的破骨前体细胞 Raw 264.7 细胞系释放炎性细胞因子的抑制作用。方法 采用 LPS 刺激 Raw 264.7 细胞构建炎症模型，采用抗酒石酸酸性磷酸酶 (TRAP)染色鉴定细胞，采用MTT 检测 Res 对 Raw 264.7 细胞的毒性影响，免疫荧光双标以及反转录聚合酶链反应(RT-PCR) 方法检测不同浓度 Res (1 μmol/L和 5 μmol/L) 对细胞炎性因子：肿瘤坏死因子-α (TNF-α) 和白细胞介素-1β (IL-1β) 与细胞炎性蛋白酶：诱导型一氧化氮合酶 (iNOS) 和环氧合酶-2 (COX-2)、炎性信号蛋白核因子-κB (NF-κB) 蛋白与 mRNA 的表达变化。 结果 不同浓度的 Res (1 μmol/L和5 μmol/L)在翻译水平和转录水平上明显抑制了 LPS 诱导的细胞炎性蛋白酶 iNOS 和 COX-2表达，同时了抑制细胞炎性因子 IL-1β 与炎性信号蛋白 NF-κB 的上调。 结论 Res 可能通过 NF-κB 调控 LPS 诱导的 Raw 264.7 细胞炎性细胞因子的释放进而抑制破骨前体细胞的激活，进而具有抗骨质疏松的作用。
Objective To investigate the inhibiting effects of resveratrol (Res) on the expression of potentially inflammatory cytokines by cultured osteoclast precursor Raw 264.7 cells stimulated with lipopolysaccharide (LPS). Methods Inflammatory cell model was established by LPS-stimulated Raw 264.7 cells. The Raw 264.7 cell identification test was measured by tartrate resistant acid phosphatase (TRAP) staining. The cells were treated with Res (1 μmol/L and 5 μmol/L) prior to LPS (5 mg/L) exposure. The effects on the mRNA and protein levels of inflammatory enzymes, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and inflammatory cytokines, tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β), inflammatory signaling proteins nuclear factor-κB (NF-κB) were analysed by reverse transcription-polymerase chain reaction (RT-PCR) and double-immunofluorescence labeling assay. The effects of Res on cytotoxicity determination of Raw 264.7 cells were measured by MTT assay. Results LPS-induced iNOS, COX-2 and NF-κB protein and mRNA expression levels were significantly decreased by Res. Res had an effect on the expression of TNF-α, IL-1β through transcriptional and translational inhibition. Conclusion The inhibitory effects of Res on LPS-induced inflammatory mediators in osteoclast precursor cells exert functions on its anti-osteoporosis through NF-κB.
白藜芦醇 / 脂多糖 / Raw 264.7细胞 / 炎性因子 / 反转录聚合酶链反应 / 免疫荧光双标 / 小鼠
Resveratrol / Lipopolysaccharide / Raw 264.7 cells / Inflammatory cytokines / RT-PCR / Double-immunofluorescence labeling / Mouselabeling
［1］Li NH, Zhu HM, Ou PZh, et al. Prevalence of primary osteoporosis in the middle aged and elderly population in parts of China ［J］. Chinese Journal of Orthopaedics, 2001,21(5):275-277. (in Chinese)
李宁华, 朱汉民, 区品中,等. 中国部分地区中老年人群原发性骨质疏松症患病率研究［J］. 中华骨科杂志, 2001,21(5):275-277.
［2］Li C, Li B, Dong Z, et al. Lipopolysaccharide differentially affects the osteogenic differentiation of periodontal ligament stem cells and bone marrow mesenchymal stem cells through Toll-like receptor 4 mediated nuclear factor kappaB pathway ［J］. Stem Cell Res Ther, 2014,5(3):67-80.
［3］Zhou MJ, Zhu ChY. Research advance in biology activity of resveratrol ［J］. Progress in Modern Biomedicine, 2008,8,(11): 2140-2148. (in Chinese)
周美娟,朱春燕.白藜芦醇生物学活性研究进展［J］. 现代生物医学进展, 2008,8,(11): 2140-2148.
［4］Pei LP, Hui BD, Wang DH, et al. Influence of trans-resveratrol on osseous changes in D-galactsose induced aging rats ［J］. Journal of Beijing University of Traditional Chinese Medicine, 2007,30(5):317-322. (in Chinese)
裴凌鹏, 惠伯棣, 王东晖,等.反式白藜芦醇对D-半乳糖致老大鼠骨质影响研究［J］. 北京中医药大学学报, 2007,30(5):317-322.
［5］Zong Y, Sun L, Liu B, et al. Resveratrol inhibits LPS-induced MAPKs activation via activation of the phosphatidylinositol 3-kinase pathway in murine RAW 264.7 macrophage cells ［J］. PLoS One, 2012,7(8):e44107.
［6］Arron JR, Choi Y. Bone versus immune system ［J］. Nature, 2000, 408(6812): 535-536.
［7］De Martinis M, Di Benedetto MC, Mengoli LP, et al. Senile osteoporosis:is it an immune-mediated disease ［J］? Inflamm Res, 2006, 55(10): 399-404.
［8］Garcia JG, Schaphorst KL. Regulation of endothelial cell gap formation and paracellular permeability ［J］. J Investig Med, 1995, 43(2): 117-126.
［9］Nair SP, Megji S, Wilson M, et al. Bacterially induced bone destruction: mechanisms and misconceptions ［J］. Infect Immun, 1996, 64(7): 2371-2380.
［10］Raisz LG, Alander C, Eilon G, et al. Effects of two bacterial products, muramyl dipeptide and endotoxin, on bone resorption in organ culture ［J］. Calcif Tissue Int, 1982, 34(4): 365-369.
［11］Li XH, Qi Y, Cai RL, et al. Effect of lipopolysaccharide-induced expression of inducible nitric oxide synthase by aloe-emodin in Raw264.7 cells ［J］. Chinese Pharmacological Bulletin, 2010, 26(4): 488-492. (in Chinese)
李晓红, 齐云, 蔡润兰,等. 芦荟大黄素对LPS诱导的Raw 264.7细胞NO生成及iNOS表达的影响 ［J］. 中国药理学通报, 2010, 26(4): 488-492.
［12］Altemeier WA, Matute-Bello G, Gharib SA, et al. Modulation of lipopolysaccharide-induced gene transcription and promotion of lung injury by mechanical ventilation ［J］. J Immunol, 2005, 175(5): 3369-3376.
［13］Zong Y, Dai JN, Sun J, et al. Ginsenoside Rg1 inhibiting the lipopolysaccharide-induced activation of microglial cells ［J］.Acta Anatomica Sinica, 2012,43(1): 13-18. (in Chinese)
宗一, 戴继男, 孙俊,等.三七皂苷Rg1抑制脂多糖诱导的小胶质细胞激活 ［J］.解剖学报, 2012,43(1): 13-18.
［14］Kitaura H, Kimura K, Ishida M, et al. Immunological reaction in TNF-α-mediated osteoclast formation and bone resorption in vitro and in vivo ［J］. Clin Dev Immunol, 2013,2013:181849.［15］Roggia C, Gao Y, Cenci S, et al. Up-regulation of TNF-producing T cells in the bone marrow:a key mechanism by which estrogen deficiency induces bone loss in vivo ［J］. Proc Natl Acad Sci USA, 2001,98(24):13960-13965.
［16］Yamamoto A, Miyazaki T, Kadono Y,et al. Possible involvement of IkappaB kinase 2 and MKK7 in osteoclastogenesis induced by receptor activator of nuclear factor kappaB ligand ［J］. J Bone Miner Res, 2002,17(4):612-621.
［17］Boeyens JC, Deepak V, Chua WH, et al. Effects of ω3- and ω6- polyunsaturated fatty acids on RANKL-induced osteoclast differentiation of RAW 264.7 cells: a comparative in vitro study ［J］. Nutrients, 2014,6(7):2584-2601.
［18］Huang BP, Lin CH, Chen YC, et al. Anti-inflammatory effects of Perilla frutescens leaf extract on lipopolysaccharide-stimulated RAW264.7 cells ［J］. Mol Med Rep, 2014,10(2):1077-1083.
［19］Gonzales AM, Orlando RA. Curcumin and resveratrol inhibit nuclear factor-kappa-B mediated cytokine expression in adipocytes ［J］. Nutr Metab(Lond), 2008,12(5): 17-30.
［20］Csiszar A, Smith K, Labinskyy N, et al. Resveratrol attenuates TNF-Alpha-induced activation of coronary arterial endothelial cells: role of NF-kappaB inhibition ［J］. Am J Physiol Heart Circ Physiol, 2006,291(4): 1694-1699.
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