Anti-HMOX1 monoclonal antibody, clone IP-2-2

Restless legs syndrome is a common complex disorder with different genetic and environmental risk factors. Here we used human cell lines to conduct an RNA-Seq study and observed how the gene showing the most significant association with RLS, MEIS1, acts as a regulator of the expression of many other genes. Some of the genes affected by its expression level are linked to pathways previously reported to be associated with RLS. We found that in cells where MEIS1 expression was either increased or prevented, mineral absorption is the principal dysregulated pathway. The mineral absorption pathway genes, HMOX1 and VDR are involved in iron metabolism and response to vitamin D, respectively. This shows a strong functional link to the known RLS pathways. We observed the same enrichment of the mineral absorption pathway in postmortem brain tissues of RLS patients showing a reduced expression of MEIS1. The expression of genes encoding metallothioneins (MTs) was observed to be dysregulated across the RNA-Seq datasets generated from both human cells and tissues. MTs are highly relevant to RLS as they bind intracellular metals, protect against oxidative stress and interact with ferritins which manage iron level in the central nervous system. Overall, our study suggests that in a subset of RLS patients, the contribution of MEIS1 appears to be associated to its downstream regulation of genes that are more directly involved in pathways that are relevant to RLS. While MTs have been implicated in the pathogenesis of neurodegenerative diseases such as Parkinson's diseases, this is a first report to propose that they have a role in RLS.

Antidepressants, such as imipramine and fluoxetine, are known to alter gene expression patterns by inducing changes in the epigenetic status of neuronal cells. There is also some evidence for the anti-apoptotic effect of various groups of antidepressants; however, this effect is complicated and cell-type dependent. Antidepressants of the tricyclic group, in particular amitriptyline, have been suggested to be beneficial in the treatment of neurodegenerative disorders. We examined whether amitriptyline exerts an anti-apoptotic effect via epigenetic mechanisms. Using DNA microarray, we analyzed global gene expression in mouse primary cultured neocortical neurons after treatment with amitriptyline and imipramine. The neuroprotection-associated genes, activating transcription factor 3 (Atf3) and heme oxygenase 1 (Hmox1), were up-regulated at both mRNA and protein levels by treatment with amitriptyline. Quantitative chromatin immunoprecipitation assay revealed that amitriptyline increased enrichments of trimethylation of histone H3 lysine 4 in the promoter regions of Atf3 and Hmox1 and acetylation of histone H3 lysine 9 in the promoter regions of Atf3, which indicate an active epigenetic status. Amitriptyline pre-treatment attenuated 1-methyl-4-phenylpyridinium ion (MPP+)- or amyloid beta peptide 1-42 (A beta(1-42))-induced neuronal cell death and inhibited the activation of extracellular signal-regulated kinase 1 and 2 (ERK1/2). We found that Atf3 and Hmox1 were also up-regulated after A beta(1-42) treatment, and were further increased when pre-treated with amitriptyline. Interestingly, the highest up-regulation of Atf3 and Hmox1, at least at mRNA level, was observed after co-treatment with A beta(1-42) and amitriptyline, together with the loss of the neuroprotective effect. These findings suggest preconditioning and neuroprotective effects of amitriptyline; however, further investigations are needed for clarifying the contribution of epigenetic up-regulation of Atf3 and Hmox1 genes.