MAOA/H202 regulation by Dexamethasone & other Steroids i

慢性应激与心血管疾病、代谢综合征、神经退行性变、抑郁症和其他精神疾病以及炎症性疾病的发病机制有关。骨骼肌病变和神经元细胞萎缩是内源性和外源性糖皮质激素过量的常见并发症，但其潜在机制仍不清楚，靶向治疗干预有限。 线粒体在细胞内稳态中起关键作用，容纳导致以ATP形式产生能量的多种代谢途径。它们还产生大多数细胞活性氧（ROS）并调节程序性细胞死亡，作为细胞凋亡的看门人。不令人惊讶的是，原发性或继发性线粒体功能障碍与具有明显公共卫生意义的病症如肥胖症、糖尿病、癌症、神经变性、心肌病、抑郁症和衰老的发病机制相关。患有上述疾病的患者通常使用宽CAM模式。 为了系统地研究线粒体基因组学对人类生理学和病理生理学的贡献，我们建立了一个以线粒体为中心的基因数据库，并将其用作定制cDNA微阵列（hMitChip）的开发参考，其中包含501个线粒体相关的核基因。为了验证这种线粒体微阵列检测基因表达变化的能力，我们探讨了地塞米松（Dex），一种合成糖皮质激素，对人骨骼肌细胞线粒体核转录组的药物基因组学效应。 在501个线粒体相关基因中，23个（4.5%）在Dex处理后骨骼肌细胞中显示出显著的转录变化。参与胺代谢的基因（MAOA和SAT）的上调在测试的所有时间间隔均具有统计学意义，而参与蛋白质加工（AKAP 10）、氧化磷酸化（FDX 1、CYBA）、热休克细胞应激反应（HSPD 1）、脂质生物合成（FACL 2）、糖酵解（GCK）和线粒体蛋白合成（TSFM）的基因上调仅在某些时间点具有统计学意义。处理7天下调与脂质代谢（ACADVL、ACADM）、三羧酸循环（DLST）、蛋白质加工（MPI、PPIB）和基因转录（GABPA）相关的基因，而诱导参与细胞凋亡的基因（BNIP 3）。分别参与细胞粘附和核苷酸代谢的CTNNA 1和AK 1基因在暴露于10-6 M Dex 7天后也显着下调。在多次重复实验中获得了可重复的结果，并且在杂交载玻片的反向标记后 对于选定的基因，用hMitChip检测到的转录变化通过QRT-PCR证实，对来自相应微阵列实验以及来自单独的时间过程实验的RNA进行，其中从两个独立的健康男性供体获得的肌细胞暴露于10-6 M的Dex长达9天。在每个时间点观察到MAOA基因的显著诱导。倍数变化随着时间的推移而增加，诱导峰值为21.2？2.7与未处理的细胞相比。在第3天和第5天，DEX处理的细胞和对照细胞之间FDX 1表达不同，而FACL 2表达在第5、7和9天不同，但随时间推移没有显著增加。在暴露于Dex 7天后观察到ACADVL和ACADM的下调趋势，与微阵列结果一致，但未能达到统计学显著性。所选基因的QRT-PCR结果，其在微阵列实验中没有差异表达（即，NNT 1，TOMM 70），因此用作阴性对照，也与微阵列发现一致。 我们报道了儿茶酚胺代谢酶MAO-A是人骨骼肌细胞中糖皮质激素的主要靶点。MAO催化神经活性、血管活性（5-羟色胺、多巴胺、儿茶酚胺）或膳食（酪胺）胺和异生物质的氧化脱氨基，释放反应性醛和H2 O2。糖皮质激素激活MAO-A，随后产生过量的过氧化氢，这可能是糖皮质激素诱导的氧化性组织损伤的一种新途径，针对该途径不同组分的MAO抑制剂或其他对抗疗法和CAM干预措施可作为治疗剂。 遗传研究已经将MAO-A和MAO-B与人类行为的各个方面联系起来，并且环境相互作用调节某些MAO-A多态性的表达;然而，它们在人类中表达的激素调节仍然未被探索。我们已经开始研究肾上腺，性腺和神经类固醇，除了糖皮质激素，对人胎儿星形胶质细胞的单胺氧化酶表达的影响。我们假设雌激素、孕激素、脱氢表雄酮和/或其他神经甾体激素（别孕烯醇酮，THDOC）对单胺氧化酶表达的影响可能与糖皮质激素相反。 看来，hMitChip是一种可靠和新颖的工具，可能被证明有助于系统地研究线粒体基因组学对人类健康和疾病的贡献。

Chronic stress is implicated in the pathogenesis of cardiovascular disease, the metabolic syndrome, neurodegeneration, depression and other psychiatric disorders, and inflammatory diseases. Skeletal myopathy and neuronal cell atrophy are common complications of endogenous and exogenous glucocorticoid excess; yet the underlying mechanisms remain unclear and targeted therapeutic interventions are limited. Mitochondria play a pivotal role in cell homeostasis, housing multiple metabolic pathways that result in energy production in the form of ATP. They also generate most cellular reactive oxygen species (ROS) and regulate programmed cell death, serving as a gatekeeper for apoptosis. Not surprisingly, primary or secondary mitochondrial dysfunction have been associated with the pathogenesis of conditions such as obesity, diabetes, cancer, neurodegeneration, cardiomyopathy, depression and aging, of obvious public health significance. Patients with the aforementioned diseases commonly use wide CAM modalities. To systematically study the contribution of mitochondrial genomics to human physiology and pathophysiology, we built a mitochondria-focused gene database and used it as reference for the development of a customized cDNA microarray (hMitChip), containing 501 mitochondria-related nuclear genes. To validate the ability of this mitochondrial microarray to detect changes in gene expression, we explored the pharmacogenomic effects of dexamethasone (Dex), a synthetic glucocorticoid, on the mitochondrial nuclear transcriptome in human skeletal muscle cells. Of the 501 mitochondria related genes arrayed in hMitChip, 23 (4.5%) displayed significant transcriptional changes in skeletal myocytes after treatment with Dex. Upregulation of genes involved in amine metabolism (MAOA and SAT) was statistically significant at all time intervals tested, whereas for genes involved in protein processing (AKAP10), oxidative phosphorylation (FDX1, CYBA), heat shock cellular stress response (HSPD1), lipid biosynthesis (FACL2), glycolysis (GCK) and mitochondrial protein synthesis (TSFM) upregulation was stastistically significant only at certain time points. Treatment for 7 days downregulated genes related to lipid metabolism (ACADVL, ACADM), the tricarboxylic acid cycle (DLST), protein processing (MPI, PPIB) and gene transcription (GABPA), whereas it induced genes involved in apoptosis (BNIP3). The CTNNA1 and AK1 genes, involved in cell adhesion and nucleotide metabolism, respectively, were also significantly downregulated after 7 days of exposure to 10-6 M of Dex. Reproducible results were obtained across multiple replicate experiments, and after reverse labeling of the hybridized slides For selected genes, the transcriptional changes detected with hMitChip were confirmed by QRT-PCR, carried out on RNA from the corresponding microarray experiments, as well as from separate time-course experiments, in which myocytes obtained from two independent healthy male donors were exposed to 10-6 M of Dex for up to 9 days. A significant induction of the MAOA gene was observed at every time point. Fold change increased over time, with a peak induction of 21.2?2.7 vs. untreated cells at day 5. FDX1 expression differed between Dex-treated and control cells at days 3 and 5, whereas FACL2 expression was different on days 5, 7, and 9, but with no significant increase over time. A trend towards downregulation was observed for ACADVL and ACADM after 7 days of exposure to Dex, consistent with the microarray findings, but failed to reach statistical significance. QRT-PCR results for selected genes, which were not differentially expressed in microarray experiments (i.e., NNT1, TOMM70) and were therefore used as negative controls, were also consistent with the microarray findings. We reported that the catecholamine-metabolizing enzyme MAO-A is a major target for glucocorticoids in human skeletal muscle cells. MAO catalyzes the oxidative deamination of neuroactive, vasoactive (serotonin, dopamine, catecholamines), or dietary (tyramine) amines and xenobiotics, releasing reactive aldehydes and H202. MAO-A activation by glucocorticoids with subsequent excess hydrogen peroxide production could represent a novel pathway for glucocorticoid-induced oxidative tissue damage, and MAO-inhibitors or other allopathic and CAM interventions aimed at different components of that pathway may serve as therapeutic agents. Genetic studies have linked MAO-A and MAO-B to various aspects of human behavior, and environmental interactions modulate the expression of certain MAO-A polymorphisms; however, the hormonal regulation of their expression in humans remains unexplored. We have begun to examine the effects of adrenal, gonadal and neurosteroids, in addition to glucocorticoids, on MAO expression in human fetal astrocytes. We hypothesized that estrogen, progesterone, DHEA and/or other neurosteroids (allopreganolone, THDOC) may elicit effectson on MAO expression opposite to those of glucocorticoids . It would appear that hMitChip is a reliable and novel tool that may prove useful for systematically studying the contribution of mitochondrial genomics to human health and disease.