Functional interaction between peroxisomes and cholesterol metabolisms in glial cells

1.肾上腺脑白质营养不良(X-ALD)是一种以中枢神经系统进行性脱髓鞘为特征的遗传性疾病。X-ALD是由于ABCD1GEM的突变所致。它编码一个一半大小的过氧体ABC蛋白，即肾上腺脑白质营养不良蛋白(ABCD1)，由745个氨基酸组成。ABCD1含有一个跨膜结构域和一个三磷酸腺苷结合结构域(S)，并被认为在二聚后起作用。在X-ALD患者的错义突变中，超过70%的突变ABCD1未被免疫印迹分析检测到。我们检测了9个错义突变的ABCD1突变的细胞内命运。我们发现，不仅突变ABCD1的功能障碍，而且突变ABCD1的错误定位和降解都与X-ALD有关。此外，我们还首次发现突变的ABCD1能够被蛋白水解酶迅速降解。ABCD1功能障碍导致超长链脂肪酸(VLCFA)在整个体液中积聚，尤其是在大脑中。ABCD1一直被认为是…更多的是作为VLCFA或VLCFA-CoA的转运体，但其确切功能仍不清楚。为了研究ABCD1在胶质细胞脂质代谢中的作用，我们制备了ABCD1基因敲除的胶质母细胞瘤细胞和ABCD1基因敲除的小鼠原代星形胶质细胞，并分析了这些细胞中的VLCFA和胆固醇代谢。这项研究表明，ABCD1功能障碍导致胶质细胞脂肪酸伸长酶上调，以及过氧体VLCFAβ氧化减少。这些VLCFA代谢的中断可能导致了VLCFA在X-ALD脑内的蓄积。此外，在ABCD1基因敲除的胶质母细胞瘤细胞中，胆固醇水平显著降低。在ABCD1基因敲除的THP-1巨噬细胞中，胆固醇进入巨噬细胞的比例减少，载脂蛋白AI依赖的胆固醇外流增加。此外，培养上清液中ApoE的分泌和胆固醇合成明显增加。尽管ABCD1与胆固醇代谢的功能相互作用尚不清楚，但ABCD1在神经胶质细胞和巨噬细胞中可能具有维持细胞内胆固醇稳态的作用。胶质细胞胆固醇代谢紊乱和VLCFA代谢紊乱可能与X-ALD的神经退行性变有关。较少

1. Adrenoleukodystrophy (X-ALD) is an inherited disorder characterized by progressive demyelination of the central nervous system. X-ALD is due to the mutations in the ABCD1 gem. It encodes a half-size peroxisomal ABC protein, adrenoleukodystrophy protein (ABCD1), which consists of 745 amino acids. ABCD1 contains a transmembrane and an ATP-binding domain(s), and is supposed to work after dimerization. Among missense mutations in X-ALD patients, more than 70% of the mutant ABCD1s were not detected by immunoblot analysis. We examined intracellular fate of 9 mutant ABCD1s with missense mutation. We have found that not only dysfunction of mutant ABCD1 but also mistargeting as well as degradation of mutant ABCD1s would be associated with X-ALD. Furthermore, we found for the fast time that mutant ABCD1s were degraded rapidly by proteasomes.2. Dysfunction of ABCD1 leads to the accumulation of very long chain fatty acids (VLCFA) in total body fluids, especially in brain. ABCD1 has been thought … More to be a transporter of VLCFA or VLCFA-CoA, but the precise function is still unclear. To investigate the roles of ABCD1 in lipid metabolisms in glial cells, we prepared ABCD1-knockdown glioblastoma cells and ABCD 1-knockout mouse primary astrocytes, and analyzed the VLCFA and cholesterol metabolisms in these cells. This study shows that dysfunction of ABCD1 results in the up-regulation of fatty acid elongases as well as the reduction of peroxisomal VLCFA β-oxidation in glial cells. Disruption of these VLCFA metabolisms might result in the VLCFA accumulation in X-ALD brain. Furthermore, cholesterol level was significantly decreased in ABCD1-knockdown glioblastoma cells.In ABCD1-knockdown THP-1 macrophage, incorporation of cholesterol into macrophage was decreased and ApoAI-dependent cholesterol efflux was increased. In addition, ApoE secretion into medium and cholesterol synthesis was significantly increased. These results suggest that cholesterol efflux was inc eased by the dysfunction of ABCD1 in THP-1 macrophage.Although the functional interaction between ABCD1 and cholesterol metabolisms remains to be determined, ABCD1 might have a role for maintaining the cellular cholesterol homeostasis both in glial cells and macrophage. The disturbed cholesterol as well as VLCFA metabolisms in glial cells might be related to the neurodegeneration in X-ALD. Less