Transcription factoys in autoimmune thiroid disease : its relevance to pathophysiology and novel therapeutic strategy

为了寻找自身免疫性甲状腺疾病的新的治疗策略，我们对脂肪分化相关蛋白（ADRP）进行了研究，ADRP最初被认为是脂肪细胞分化程序的早期标志物。这种蛋白质广泛表达，并参与脂质储存以及长链脂肪酸摄取。由于游离脂肪酸可能是炎症反应的介质，我们的目的是揭示ADRP基因表达的调控机制。在本研究中，我们发现小鼠ADRP基因的表达受PPARg的调控。因此，PPARg配体曲格列酮、吡格列酮和15-脱氧-D12，14-前列腺素J2在体外增加NMuLi小鼠肝细胞和J774.1小鼠巨噬细胞中的ADRP mRNA水平。这种增加被放线菌素D完全抑制。相反，PPARa配体，非诺贝特和苯扎贝特，未能增加mRNA水平。为了描述负责这种PPARg诱导的刺激的启动子区域，我们克隆了小鼠ADRP基因的约2.8kb的5 '侧翼区，并构建了用于评估启动子活性的荧光素酶报告质粒。当转染到NMuLi细胞中时，与对照质粒相比，所有含有启动子区的嵌合构建体都表现出显著的荧光素酶活性。曲格列酮或曲格列酮与9 -cis-RA联合使用可显著刺激含有-2090 bp或更长片段的启动子表达的启动子活性，但不能刺激含有-2005 bp或更短片段的启动子表达的启动子活性，表明-2090和-2006 bp之间的区域负责PPARg的作用。这些结果将有助于理解PPARg对ADRP表达的作用机制。我们目前正在研究ADRP在甲状腺细胞中的生理意义和自身免疫性甲状腺疾病的调节。

In order to search new therapeutic strategies for autoimmune thyroid diseases, we have worked on adipose differentiation-related protein (ADRP), which was originally identified as an early marker of adipose cell differentiation program. This protein is ubiquitously expressed and is involved in lipid storage as well as long chain fatty acid uptake. Since free fatty acid could be a mediator of inflammatory response, we aimed to disclose the regulatory mechanism of the ADRP gene expression. In the present study, we disclosed that the mouse ADRP gene expression is regulated by PPARg. Thus, PPARg ligands, troglitazone, pioglitazone and 1 5 -deoxy-D12,14-prostaglandin J2, increased ADRP mRNA levels in NMuLi mouse liver cells and J774.1 mouse macrophages in vitro. This increase was completely inhibited by actinomycin D.On the contrary, PPARa ligands, fenofibrate and bezafibrate, failed to increase the mRNA levels. In order to delineate the promoter region responsible for this PPARg-induced stimulation, we cloned an approximately 2.8kb 5'-flanking region of the mouse ADRP gene, and constructed luciferase reporter plasmids for the assessment of promoter activity. All chimeric constructs containing the promoter region, when transfected into NMuLi cells, exhibited significant luciferase activity by comparison to a control plasmid. Troglitazone or troglitazone plus 9 -cis-RA significantly stimulated promoter activity expressed by the promoter containing a fragment of-2090bp or longer, but not -2005bp or shorter, indicating that the region between -2090 and -2006bp is responsible for the PPARg action. These results will facilitate understanding of the mechanism of PPARg action on ADRP expression. We are currently examining physiological significance of ADRP in thyrocytes and regulation of autoimmune thyroid diseases.