Homocysteine Suppresses the Expression of the Collagen Cross-linker Lysyl Oxidase Involving IL-6, Fli1, and Epigenetic DNA Methylation
Homocysteine Suppresses the Expression of the Collagen Cross-linker Lysyl Oxidase Involving IL-6, Fli1, and Epigenetic DNA Methylation
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DOI：
10.1074/jbc.m110.166181
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发表时间：
2011-02-18
影响因子：
4.8
通讯作者：
Varga, Franz
中科院分区：
文献类型：
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作者：
Thaler, Roman;Agsten, Marlies;Varga, Franz
血清同型半胱氨酸(Hcys)水平升高是几种慢性疾病的危险因素，包括心血管疾病、动脉粥样硬化和慢性肾功能衰竭，并影响骨骼发育、质量和动态平衡。Hcys直接通过抑制胶原交联酶赖氨酰氧化酶(Lox)和抑制其mRNA的表达来影响稳定的骨基质的形成。本研究的目的是探讨这一过程中涉及的机制。通过基因芯片、定量RT-PCR、免疫印迹和酶联免疫吸附试验，我们鉴定了成骨前细胞中依赖于hcys刺激的白介素6(IL-6)和依赖于IL-6/Janus kinase2(JAK2)信号转导通路的基因。此外，还观察到表观遗传DNA甲基化所必需的基因(DNA(胞嘧啶-5)甲基转移酶和解旋酶淋巴特异性(Hells))的上调。进一步的研究表明，Hcys通过IL-6/JAK2增加了转录因子FLI1(Friend白血病病毒整合1)的表达，我们发现该转录因子对IL-6依赖的Dnmt1刺激是必不可少的。对富含CpG的Lox近端启动子的CpG甲基化分析显示，在Hcys处理细胞后，CpG甲基化状态增加，这表明Hcys依赖的Lox抑制是表观遗传起源。抑制IL-6/JAK2途径或CpG甲基化可逆转Hcys对Lox表达的抑制作用。总之，我们证明了Hcys刺激成骨细胞中IL-6的合成，这是已知的通过破骨细胞影响骨代谢的。此外，IL-6通过JAK2、FLI1和Dnmt1的刺激结果通过表观遗传的CpG甲基化下调Lox的表达，揭示了一种新的负面影响骨基质形成的机制。
Elevated homocysteine (Hcys) serum levels represent a risk factor for several chronic pathologies, including cardiovascular disease, atherosclerosis, and chronic renal failure, and affect bone development, quality, and homeostasis. Hcys influences the formation of a stable bone matrix directly through the inhibition of the collagen cross-linking enzyme lysyl oxidase (Lox) and, as we have shown recently, by repressing its mRNA expression. The aim of this study was to investigate the mechanisms involved in this process. Through evaluation of gene arrays, quantitative RT-PCR, immunoblots, and ELISA, we identified a Hcys-dependent stimulation of interleukin 6 (IL-6) and genes involved in IL-6/Janus kinase 2 (JAK2)-dependent signal transduction pathways in pre-osteoblastic MC3T3-E1 cells. Moreover, up-regulation of genes essential for epigenetic DNA methylation (DNA (cytosine-5)-methyltransferases and helicase lymphoid-specific (Hells) was observed. Further investigations demonstrated that Hcys increased via IL-6/JAK2 the expression of Fli1 (Friend leukemia virus integration 1), a transcription factor, which we found essential for IL-6-dependent Dnmt1 stimulation. CpG methylation analysis of CpG-rich Lox proximal promoter revealed an increased CpG methylation status after treatment of the cells with Hcys indicating an epigenetic origin for Hcys-dependent Lox repression. Inhibition of the IL-6/JAK2 pathway or of CpG methylation reversed the repressive effect of Hcys on Lox expression. In conclusion, we demonstrate that Hcys stimulates IL-6 synthesis in osteoblasts, which is known to affect bone metabolism via osteoclasts. Furthermore, IL-6 stimulation results via JAK2, Fli1, and Dnmt1 in down-regulation of Lox expression by epigenetic CpG methylation revealing a new mechanism negatively affecting bone matrix formation.