Putrescine as indicator of manganese neurotoxicity: Dose-response study in human SH-SY5Y cells

Fernandes, Jolyn; Chandler, Joshua D.; Liu, Ken H.; Uppal, Karan; Go, Young‐Mi; Jones, Dean P.

doi:10.1016/j.fct.2018.04.042

Cited by 11 publications

(7 citation statements)

References 77 publications

(105 reference statements)

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“…Mn is directly important in this pathway because conversion of arginine to urea and ornithine is catalyzed by arginase, a Mn-dependent enzyme. We previously found that Mn altered putrescine and other polyamines derived from ornithine (Fernandes et al, 2018). In addition, Mn decreased arginine and increased N-acetyl-L-glutamate 5-semialdehyde, the second to last step in ornithine biosynthesis.…”

Section: Discussionmentioning

confidence: 97%

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Metabolomic Responses to Manganese Dose in SH-SY5Y Human Neuroblastoma Cells

Fernandes

Chandler

Liu

et al. 2019

Toxicological Sciences

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Manganese (Mn)-associated neurotoxicity has been well recognized. However, Mn is also an essential nutrient to maintain physiological function. Our previous study of human neuroblastoma SH-SY5Y cells showed that Mn treatment comparable to physiological and toxicological concentrations in human brain resulted in different mitochondrial responses, yet cellular metabolic responses associated with such different outcomes remain uncharacterized. Herein, SH-SY5Y cells were examined for metabolic responses discriminated by physiological and toxicological levels of Mn using high-resolution metabolomics (HRM). Before performing HRM, we examined Mn dose (from 0 to100 lM) and time effects on cell death. Although we did not observe any immediate cell death after 5 h exposure to any of the Mn concentrations assessed (0-100 lM), cell loss was present after a 24-h recovery period in cultures treated with Mn ! 50 lM. Exposure to Mn for 5 h resulted in a wide range of changes in cellular metabolism including amino acids (AA), neurotransmitters, energy, and fatty acids metabolism. Adaptive responses at 10 lM showed increases in neuroprotective AA metabolites (creatine, phosphocreatine, phosphoserine). A 5-h exposure to 100 mM Mn, a time before any cell death occurred, resulted in decreases in energy and fatty acid metabolites (hexose-1,6 bisphosphate, acyl carnitines). The results show that adjustments in AA metabolism occur in response to Mn that does not cause cell death while disruption in energy and fatty acid metabolism occur in response to Mn that results in subsequent cell death. The present study establishes utility for metabolomics analyses to discriminate adaptive and toxic molecular responses in a human in vitro cellular model that could be exploited in evaluation of Mn toxicity.

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Section: Discussionmentioning

confidence: 97%

“…Cells extracts were analyzed as described previously (Fernandes et al, 2018;Go et al, 2014b;Liu et al, 2016). Briefly, thawed extracts were incubated at 4 C for 30 min, centrifuged at 16 100 g for 10 min to remove protein and transferred to a refrigerated (4 C) autosampler for analysis.…”

Section: High-resolution Metabolomicsmentioning

confidence: 99%

Metabolomic Responses to Manganese Dose in SH-SY5Y Human Neuroblastoma Cells

Fernandes

Chandler

Liu

et al. 2019

Toxicological Sciences

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“…SH-SY5Y, human neuroblastoma cell line, obtained from the American Type Culture Collection (ATCC-Manassas, VA) was cultured and exposed to Mn dose as previously described ( Supplementary Datasheet 1 ) (Fernandes et al, 2017; Fernandes et al, 2018; Fernandes et al, 2019). Briefly, cells were cultured in Dulbecco’s modified Eagle medium/Ham’s F12 medium and, at 80% confluency, were treated with Mn (0, 1, 5, 10, 50, 100 μM as MnCl 2 ; Sigma-Aldrich, St. Louis, MO) for 5 h. The 5-h time point was chosen to correlate transcriptomic changes with cellular Mn concentration comprising physiological (≤10 μM Mn treatment) to pathophysiological Mn levels (≥50 μM Mn treatment) that was optimized previously (Fernandes et al, 2017) based on the recommended dosage of treatment and accumulation of cellular Mn relevant to in vivo levels from human brain and in vitro studies (Csaszma et al, 2003; Bowman and Aschner, 2014; Kumar et al, 2015; Fernandes et al, 2017).…”

Section: Methodsmentioning

confidence: 99%

Transcriptome Analysis Reveals Distinct Responses to Physiologic versus Toxic Manganese Exposure in Human Neuroblastoma Cells

et al. 2019

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Manganese (Mn) is an essential trace element, which also causes neurotoxicity in exposed occupational workers. Mn causes mitochondrial toxicity; however, little is known about transcriptional responses discriminated by physiological and toxicological levels of Mn. Identification of such mechanisms could provide means to evaluate risk of Mn toxicity and also potential avenues to protect against adverse effects. To study the Mn dose-response effects on transcription, analyzed by RNA-Seq, we used human SH-SY5Y neuroblastoma cells exposed for 5 h to Mn (0 to 100 μM), a time point where no immediate cell death occurred at any of the doses. Results showed widespread effects on abundance of protein-coding genes for metabolism of reactive oxygen species, energy sensing, glycolysis, and protein homeostasis including the unfolded protein response and transcriptional regulation. Exposure to a concentration (10 μM Mn for 5 h) that did not result in cell death after 24-h increased abundance of differentially expressed genes (DEGs) in the protein secretion pathway that function in protein trafficking and cellular homeostasis. These include BET1 (Golgi vesicular membrane-trafficking protein), ADAM10 (ADAM metallopeptidase domain 10), and ARFGAP3 (ADP-ribosylation factor GTPase-activating protein 3). In contrast, 5-h exposure to 100 μM Mn, a concentration that caused cell death after 24 h, increased abundance of DEGs for components of the mitochondrial oxidative phosphorylation pathway. Integrated pathway analysis results showed that protein secretion gene set was associated with amino acid metabolites in response to 10 μM Mn, while oxidative phosphorylation gene set was associated with energy, lipid, and neurotransmitter metabolites at 100 μM Mn. These results show that differential effects of Mn occur at a concentration which does not cause subsequent cell death compared to a concentration that causes subsequent cell death. If these responses translate to effects on the secretory pathway and mitochondrial functions in vivo , differential activities of these systems could provide a sensitive basis to discriminate sub-toxic and toxic environmental and occupational Mn exposures.

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“…At first approximation, we focused on data linked to PA metabolism. Literature data show that added Mn 2+ dose-dependently (1–100 µM) increases putrescine content (3.2–4.5 nmol/mg protein) in human SH-SY5Y cells [ 88 ]. Besides, the increase of putrescine content positively correlates with the PA metabolite N-acetylspermidine, but negatively with GABA related metabolites [ 88 ].…”

Section: Discussionmentioning

confidence: 99%

Dual Role for Astroglial Copper-Assisted Polyamine Metabolism during Intense Network Activity

et al. 2021

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Astrocytes serve essential roles in human brain function and diseases. Growing evidence indicates that astrocytes are central players of the feedback modulation of excitatory Glu signalling during epileptiform activity via Glu-GABA exchange. The underlying mechanism results in the increase of tonic inhibition by reverse operation of the astroglial GABA transporter, induced by Glu-Na+ symport. GABA, released from astrocytes, is synthesized from the polyamine (PA) putrescine and this process involves copper amino oxidase. Through this pathway, putrescine can be considered as an important source of inhibitory signaling that counterbalances epileptic discharges. Putrescine, however, is also a precursor for spermine that is known to enhance gap junction channel communication and, consequently, supports long-range Ca2+ signaling and contributes to spreading of excitatory activity through the astrocytic syncytium. Recently, we presented the possibility of neuron-glia redox coupling through copper (Cu+/Cu2+) signaling and oxidative putrescine catabolism. In the current work, we explore whether the Cu+/Cu2+ homeostasis is involved in astrocytic control on neuronal excitability by regulating PA catabolism. We provide supporting experimental data underlying this hypothesis. We show that the blockade of copper transporter (CTR1) by AgNO3 (3.6 µM) prevents GABA transporter-mediated tonic inhibitory currents, indicating causal relationship between copper (Cu+/Cu2+) uptake and the catabolism of putrescine to GABA in astrocytes. In addition, we show that MnCl2 (20 μM), an inhibitor of the divalent metal transporter DMT1, also prevents the astrocytic Glu-GABA exchange. Furthermore, we observed that facilitation of copper uptake by added CuCl2 (2 µM) boosts tonic inhibitory currents. These findings corroborate the hypothesis that modulation of neuron-glia coupling by copper uptake drives putrescine → GABA transformation, which leads to subsequent Glu-GABA exchange and tonic inhibition. Findings may in turn highlight the potential role of copper signaling in fine-tuning the activity of the tripartite synapse.

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Putrescine as indicator of manganese neurotoxicity: Dose-response study in human SH-SY5Y cells

Cited by 11 publications

References 77 publications

Metabolomic Responses to Manganese Dose in SH-SY5Y Human Neuroblastoma Cells

Metabolomic Responses to Manganese Dose in SH-SY5Y Human Neuroblastoma Cells

Transcriptome Analysis Reveals Distinct Responses to Physiologic versus Toxic Manganese Exposure in Human Neuroblastoma Cells

Dual Role for Astroglial Copper-Assisted Polyamine Metabolism during Intense Network Activity

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