A disturbed regulation of multi-differentiation in human mesenchymal stem cells by dioxin and its mechanism

芳烃受体(AhR)是一种配体激活的转录因子，介导2，3，7，8-四氯二恶英(TCDD)及其相关化合物的一系列毒理学和生物学效应。AHR基因敲除的小鼠在肝脏和脊柱弯曲处表现出脂肪堆积。因此，我们研究了TCDD对人骨髓间充质干细胞向脂肪细胞和骨细胞分化的影响。TCDD可抑制人骨髓间充质干细胞向脂肪细胞的分化。另一方面，TCDD促进了细胞向骨细胞的分化。这些结果表明，脂肪组织质量丢失(消瘦综合征)、骨发育不良和骨软骨病是由于人骨髓间充质干细胞向多功能细胞分化的调节紊乱所致。此外，我们还检测了对TCDD高度敏感的3T3-L1细胞AhR的表达水平。促肾上腺皮质激素受体蛋白水平为…3T3-L1细胞在成脂过程中受抑制程度较高。蛋白酶体抑制剂对3T3-L1细胞前脂肪细胞和脂肪细胞的作用不影响AhR蛋白水平。在3T3-L1细胞中，AhR基因的表达水平与AhR蛋白水平相似，在成脂过程中被耗尽。接下来，为了了解AhR在脂肪形成过程中被耗尽的机制，我们分析了3T3-L1细胞脂肪分化过程中AhR启动子的活性。为了确定负责对AhR转录的分化抑制作用的AhR启动子区域的序列，将一系列与CAT报告基因连锁的缺失构建体导入3T3-L1细胞。前脂肪细胞和脂肪细胞的CAT活性比较表明，-399/-339和-296/-217序列是AhR启动子活性下调的核心贡献者，依赖于分化。使用核组分的凝胶位移分析表明，该因子与-399/-339和-296/-217序列结合。前脂肪细胞的结合活性高于脂肪细胞。因此，反式作用因子的下调可能导致脂肪分化过程中AhR基因转录的抑制。较少

Arylhydrocarbon receptor(AhR) is a ligand-activated transcription factor that mediates a spectrum of toxicological and biological effects of 2,3,7,8-tetrachloro-p-dioxin(TCDD) and related compounds. AhR knockout mice exhibited a lipid accumulation in the liver and the curvature of the spinal column. Therefore, we investigated the effect of TCDD treatment on the differentiation of human mesenchymal stem cells to adipocytes and osteocytes. The differentiation to adipocytes in human mesenchymal stem cells was depressed by TCDD treatment. On the other hand, the differentiation to osteocytes in the cells was accumulated by TCDD. These results suggest that the loss of adipose tissue mass (wasting syndrome), osteodysplasia and osteochondrosis are induced by the disturbed regulation of the differentiation in the human mesenchymal stem cells to multi-functional cells.Furthermore, we examined the expression level of AhR in 3T3-L1 cells which have highly sensitivity to TCDD. AhR protein level was … More depressed in 3T3-L1 cells during adipogenesis. The treatment of proteosome inhibitors to preadipocytes and adipocytes in 3T3-L1 cells was no affected to the level of AhR protein. In the expression of AhR mRNA, the level was depleted in 3T3-L1 cells during adipogenesis as similarly the AhR protein level. Next, to understand the mechanism by which the AhR is depleted during adipogenesis, we analyzed the AhR promoter activity during adipose differentiation in 3T3-L1 cells. To identify the sequence of the AhR promoter region responsible for differentiation-dependent suppression of AhR transcription, a series of deletion constructs linked to the CAT reporter were transfected into 3T3-L1 cells. A comparison of CAT activity between preadipocytes and adipocytes revealed that the sequences of -399/-339 and -296/-217 are core contributor differentiation-dependent downregulation of AhR promoter activity. Gel shift analysis using the nuclear fraction showed the presence of the factor bound to the sequences of -399/-339 and -296/-217. The binding activity was higher in preadipocytes than in adipocytes. Consequently, the downregulation of the trans-acting factor may result in the suppression of AhR gene transcription during adipose differentiation. Less