Brodňanová M, Cibulka M, Kolísek M
Biomedical Centre Martin, Jessenius Faculty of Medicine in Martin, Comenius University Bratislava, Slovakia
Background: Impaired magnesium (Mg) homeostasis (IMH) is considered one of the important factors involved in neurodegeneration. One of the most common neurodegenerative diseases – Parkinson's disease – is associated with PARK16 locus. This locus includes SLC41A1 gene, encoding an important Na+/Mg2+ exchanger system, which is ubiquitously expressed in all human cells.
Neuroinflammation, another critical component of the neurodegenerative process, is accompanied by increased levels of interleukin 6 (IL-6). IL-6 exhibits both pro-inflammatory and anti-inflammatory effects depending on other factors. Presence of the α-subunit of the IL-6 membrane receptor is not sufficient to trigger the classical signalling pathway, which is considered anti-inflammatory. Also, the β-subunit of the receptor, which is present mainly on the membranes of hepatocytes, megakaryocytes and leukocytes, is necessary. In the absence of the membrane form of the β-subunit, cells are able to respond to the presence of IL-6 after its interaction with the soluble form of β-subunit. The signalling pathway that is triggered in this scenario is called trans-signalling and is considered pro-inflammatory.
Binding of IL-6 to the receptor triggers the phosphorylation of STAT3 molecules by JAK activity. STAT3 subsequently dimerizes and the dimer is transported to the nucleus, where as an active transcription factor it regulates the expression of STAT3-responsive genes. A putative binding site for STAT3 was also predicted to be in the promoter region of SLC41A1 gene in PARK16 locus.
Aim: Therefore, the aim of our study was to elucidate a possible link between neuroinflammation, modelled by the presence of IL-6, and IMH. Our hypothesis is based on the involvement of both components in the process of neurodegeneration.
Methods: In our experiments, we used four IL-6-responsive cell lines expressing genes encoding both the α- and β-subunit of the IL-6 receptor (HEK-BlueTM IL-6, HepG2, U-266 and PANC-1). Cells were treated with recombinant human IL-6 at a concentration of 1, 50 and 100 ng/mL for different time intervals. The specificity of the induced response was verified using the Western blot analysis with specific antibodies against post-translationally modified forms of STAT3. Potential changes in the expression of nine magnesiotropic genes (SLC41A1, SLC41A2, SLC41A3, TRPM7, MAGT1, NIPA1, N33, CNNM2, TRPM6) were analysed at the level of transcription using the RT-qPCR analysis.
Results: A specific response by activating the IL-6/JAK/STAT3 pathway was observed in all four cell lines. Despite the activation of the STAT3 transcription factor in these cells, IL-6 does not affect transcription of the nine studied magnesiotropic genes, after neither 1 nor 24 hours long treatment with lower (1 ng/mL) or higher (50 ng/mL) concentration. However, IL-6 still may interfere with the Mg homeostasis at some other level or needs to act in cooperation with some other factors to do so.
Acknowledgement: This study was supported by grant VEGA 1/0554/19.
