Abstract
The expression of 5-lipoxygenase (5-LOX) is affected by aging and regulated by epigenetic mechanisms including DNA methylation. We used methylation-sensitive restriction endonucleases (AciI, BstUI, HpaII, and HinP1I) to assess 5-LOX DNA methylation in brain and heart tissue samples from young (2 months) and old (22 months) mice. We also measured mRNA content for 5-LOX and the DNA methyltransferases DNMT1 and DNMT3a. In young mice, the 5-LOX mRNA content was significantly greater in the heart compared to the brain; 5-LOX DNA methylation was lower, except in the AciI assay in which it was higher in the heart. Aging decreased 5-LOX mRNA content in the heart and increased it in the brain. Aging also increased 5-LOX DNA methylation and this effect was site- (i.e., enzyme) and tissue-specific. Generally, DNMT1 and DNMT3a mRNA content was lower in the brain regions compared to the heart; the only effect of aging was observed in the mRNA content of DNMT3a, which was decreased in the heart of old mice. These results indicate a complex tissue-specific and aging-dependent interplay between the DNA methylation system and 5-LOX mRNA content. Interpretation of this data must take into account that the tissue samples contained a mixture of various cell types.
Highlights
5-lipoxygenase (5-LOX; EC 1.13.11.34; encoded by the ALOX5 gene) is the key enzyme in the biosynthesis of lipid signaling molecules, that is, the inflammatory leukotrienes and the anti-inflammatory lipoxins [1, 2]
Using the 5-LOX DNA methylation assay, which is based on the ability of methylation-sensitive endonucleases to digest only unmethylated recognition sites, we found that the four endonucleases—AciI, BstUI, HinP1I, and HpaII—revealed a differential tissue- and brain-regionspecific methylation status of 5-LOX DNA (Figure 4)
In line with previous reports [14], we confirmed that 5LOX mRNA is expressed in the mouse brain, albeit at levels significantly lower than 5-LOX mRNA levels measured in heart tissue
Summary
5-lipoxygenase (5-LOX; EC 1.13.11.34; encoded by the ALOX5 gene) is the key enzyme in the biosynthesis of lipid signaling molecules, that is, the inflammatory leukotrienes and the anti-inflammatory lipoxins [1, 2]. The 5-LOX pathway has been considered in mechanisms of neural plasticity ranging from neurogenesis and neural differentiation [9] to regulation of synaptic plasticity [10, 11]. Regulation of the brain expression of 5-LOX participates in mechanisms of neuroprotection [8]. In the CNS of rats and mice, 5-LOX expression increases during aging [14, 17] and in brain ischemia [18, 19]. 5-LOX gene expression is regulated by epigenetic mechanisms including modifications of DNA methylation [1, 24– 26]. Epigenetics encompasses the modification of chromatin structure that leads to regulation of the gene expression and Neural Plasticity mRNA 5-LOX DNMT1. DNA methylation at the sites of CpG dinucleotides has been implicated in gene regulation in virtually all tissues, including the brain [28]. It was observed that the genomic DNA methylation in patients with coronary artery disease is significantly higher than in controls [29]
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