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Stress Tolerance and Protein O‐GlcNAc Regulation in Neuroblastoma Cells under Hypoglycemic Condition
Author(s) -
Nagy Zsófia,
Kátai Emese,
Miseta Attila,
Nagy Tamas
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.721.2
Subject(s) - phosphorylation , western blot , chemistry , viability assay , carbohydrate metabolism , flow cytometry , cell culture , metabolism , glucose uptake , cell , tau protein , enzyme , endocrinology , medicine , biochemistry , microbiology and biotechnology , biology , alzheimer's disease , insulin , disease , genetics , gene
Alzheimer's disease is often referred to as type III diabetes since many studies demonstrated that glucose metabolism is significantly decreased in these patients. The exact mechanism how this metabolic change leads to Alzheimer's is still unclear. However Protein O‐linked N‐acetylglucosamine modification (O‐GlcNAc) may play a pivotal role in this process. O‐GlcNAc modification is a reversible post‐translational modification influenced by substrate availability (hexosamine pathway and UDP‐GlcNAc) which depends on glucose metabolism. Importantly, O‐GlcNAc and phosphorylation seems to be in balance on Tau‐protein, which may form neurofibrillary tangles if hyperphosphorylated. Despite of O‐GlcNAc's obvious connection to glucose metabolism, the level of O‐GlcNAc modification under hypoglycemic condition was not in the focus of research. Thus, the objective of this study was to investigate the relationship between O‐GlcNAc modification and Tau phosphorylation in a hypoglycemic cell culture model. We cultured the neuroblastoma cell line SH‐SY5Y in various amount of glucose‐containing media (5.5, 3, 1.5, 1, 0.5, 0.25 mM). In the first set of experiments, the glucose concentrations were kept constant for 96 hours. In a second set of experiments, we let the cells run out of glucose. Cell viability was analyzed by flow cytometry and direct microscopic observation. O‐GlcNAc, Tau‐Ps199, total Tau levels were detected by Western‐blot. We have found that SH‐SY5Y cells were resistant to very low level of glucose since the viability, cell morphology did not change significantly when compared to 5.5 mM glucose condition, as long as the media glucose was replenished to maintain the low, but steady level. However, when the cells were left to metabolize all of the glucose, the viability quickly deteriorated. O‐GlcNAc levels and P‐Tau levels were also steady, independent of the glucose concentration. In the second set of experiments, soon after glucose was completely depleted by the cells, O‐GlcNAc initially decreased then significantly elevated when detected with O‐GlcNAc specific antibody CTD110.6. In conclusion, our data suggest that hypoglycemic glucose concentrations could be still sufficient for neuronal cells, however glucose fasting quickly leads to cellular stress and eventually, cell death. In the future, we plan to study whether nutrient supplements, such as beta‐hydroxy‐butirate or galactose might alleviate or delay the effect of the unavailable glucose. Support or Funding Information This work was funded by the Research Fund of the University of Pécs, Faculty of Medicine (AOK‐KA 2018) and the EU founded Hungarian project GINOP 2.3.2.‐15 (grant No.: 2.3.2.‐15‐2016‐00050). This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .