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Lithium protects against paraquat neurotoxicity by NRF2 activation and miR-34a inhibition in SH-SY5Y cells.

Alural B, Ozerdem A, Allmer J, Genc K, Genc S - Front Cell Neurosci (2015)

Bottom Line: Furthermore, lithium significantly decreased both basal and PQ-induced expression of miR-34a.Transfection of miR-34a specific mimic reversed neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium against PQ-toxicity.Our results revealed two novel mechanisms of lithium neuroprotection, namely NRF2 activation and miR-34a suppression.

View Article: PubMed Central - PubMed

Affiliation: Izmir Biomedicine and Genome Center, Dokuz Eylul University Izmir, Turkey ; Department of Neuroscience, Health Science Institute, Dokuz Eylul University Izmir, Turkey.

ABSTRACT
Lithium is a mood stabilizing agent commonly used for the treatment of bipolar disorder. Here, we investigated the potential neuroprotective effect of lithium against paraquat toxicity and its underlying mechanisms in vitro. SH-SY5Y human neuroblastoma cells were treated with paraquat (PQ) 0.5 mM concentration after lithium pretreatment to test lithium's capability in preventing cell toxicity. Cell death was evaluated by LDH, WST-8, and tryphan blue assays. Apoptosis was analyzed using DNA fragmentation, Annexin V immunostaining, Sub G1 cell cycle analysis, and caspase-3 activity assays. BCL2, BAX, and NRF2 protein expression were evaluated by Western-blotting and the BDNF protein level was determined with ELISA. mRNA levels of BCL2, BAX, BDNF, and NRF2 target genes (HO-1, GCS, NQO1), as well as miR-34a expression were analyzed by qPCR assay. Functional experiments were done via transfection with NRF2 siRNA and miR-34a mimic. Lithium treatment prevented paraquat induced cell death and apoptosis. Lithium treated cells showed increased anti-apoptotic protein BCL2 and decreased pro-apoptotic protein BAX expression. Lithium exerted a neurotrophic effect by increasing BDNF protein expression. It also diminished reactive oxygen species production and activated the redox sensitive transcription factor NRF2 and increased its target genes expression. Knockdown of NRF2 abolished neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium. Furthermore, lithium significantly decreased both basal and PQ-induced expression of miR-34a. Transfection of miR-34a specific mimic reversed neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium against PQ-toxicity. Our results revealed two novel mechanisms of lithium neuroprotection, namely NRF2 activation and miR-34a suppression.

No MeSH data available.


Related in: MedlinePlus

Lithium increases BDNF expression and secretion in SH-SY5Y cells. Cells were pretreated with lithium (2 mM) for 24 h prior to PQ exposure. (A) Intracellular BDNF mRNA levels were quantified by real time qPCR. PQ exposure markedly reduced the BDNF mRNA level while lithium pretreatment reversed the PQ effect on the BDNF mRNA level. (B) Secreted BDNF protein levels in culture medium were measured by BDNF ELISA kit. Lithium increased PQ induced BDNF decrease in SH-SY5Y cells. The data are presented as mean ± S.E, n = 5. (*p < 0.05 compared to control and #p < 0.05 compare to PQ treated cells).
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Figure 4: Lithium increases BDNF expression and secretion in SH-SY5Y cells. Cells were pretreated with lithium (2 mM) for 24 h prior to PQ exposure. (A) Intracellular BDNF mRNA levels were quantified by real time qPCR. PQ exposure markedly reduced the BDNF mRNA level while lithium pretreatment reversed the PQ effect on the BDNF mRNA level. (B) Secreted BDNF protein levels in culture medium were measured by BDNF ELISA kit. Lithium increased PQ induced BDNF decrease in SH-SY5Y cells. The data are presented as mean ± S.E, n = 5. (*p < 0.05 compared to control and #p < 0.05 compare to PQ treated cells).

Mentions: Our results showed that lithium alone increased BDNF mRNA expression by 2.7 fold whereas PQ treatment caused a 0.62-fold reduction in BDNF mRNA expression compared to control cells. Lithium pretreatment prevented the decrease in BDNF mRNA levels after treatment with PQ (Figure 4A).


Lithium protects against paraquat neurotoxicity by NRF2 activation and miR-34a inhibition in SH-SY5Y cells.

Alural B, Ozerdem A, Allmer J, Genc K, Genc S - Front Cell Neurosci (2015)

Lithium increases BDNF expression and secretion in SH-SY5Y cells. Cells were pretreated with lithium (2 mM) for 24 h prior to PQ exposure. (A) Intracellular BDNF mRNA levels were quantified by real time qPCR. PQ exposure markedly reduced the BDNF mRNA level while lithium pretreatment reversed the PQ effect on the BDNF mRNA level. (B) Secreted BDNF protein levels in culture medium were measured by BDNF ELISA kit. Lithium increased PQ induced BDNF decrease in SH-SY5Y cells. The data are presented as mean ± S.E, n = 5. (*p < 0.05 compared to control and #p < 0.05 compare to PQ treated cells).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4446540&req=5

Figure 4: Lithium increases BDNF expression and secretion in SH-SY5Y cells. Cells were pretreated with lithium (2 mM) for 24 h prior to PQ exposure. (A) Intracellular BDNF mRNA levels were quantified by real time qPCR. PQ exposure markedly reduced the BDNF mRNA level while lithium pretreatment reversed the PQ effect on the BDNF mRNA level. (B) Secreted BDNF protein levels in culture medium were measured by BDNF ELISA kit. Lithium increased PQ induced BDNF decrease in SH-SY5Y cells. The data are presented as mean ± S.E, n = 5. (*p < 0.05 compared to control and #p < 0.05 compare to PQ treated cells).
Mentions: Our results showed that lithium alone increased BDNF mRNA expression by 2.7 fold whereas PQ treatment caused a 0.62-fold reduction in BDNF mRNA expression compared to control cells. Lithium pretreatment prevented the decrease in BDNF mRNA levels after treatment with PQ (Figure 4A).

Bottom Line: Furthermore, lithium significantly decreased both basal and PQ-induced expression of miR-34a.Transfection of miR-34a specific mimic reversed neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium against PQ-toxicity.Our results revealed two novel mechanisms of lithium neuroprotection, namely NRF2 activation and miR-34a suppression.

View Article: PubMed Central - PubMed

Affiliation: Izmir Biomedicine and Genome Center, Dokuz Eylul University Izmir, Turkey ; Department of Neuroscience, Health Science Institute, Dokuz Eylul University Izmir, Turkey.

ABSTRACT
Lithium is a mood stabilizing agent commonly used for the treatment of bipolar disorder. Here, we investigated the potential neuroprotective effect of lithium against paraquat toxicity and its underlying mechanisms in vitro. SH-SY5Y human neuroblastoma cells were treated with paraquat (PQ) 0.5 mM concentration after lithium pretreatment to test lithium's capability in preventing cell toxicity. Cell death was evaluated by LDH, WST-8, and tryphan blue assays. Apoptosis was analyzed using DNA fragmentation, Annexin V immunostaining, Sub G1 cell cycle analysis, and caspase-3 activity assays. BCL2, BAX, and NRF2 protein expression were evaluated by Western-blotting and the BDNF protein level was determined with ELISA. mRNA levels of BCL2, BAX, BDNF, and NRF2 target genes (HO-1, GCS, NQO1), as well as miR-34a expression were analyzed by qPCR assay. Functional experiments were done via transfection with NRF2 siRNA and miR-34a mimic. Lithium treatment prevented paraquat induced cell death and apoptosis. Lithium treated cells showed increased anti-apoptotic protein BCL2 and decreased pro-apoptotic protein BAX expression. Lithium exerted a neurotrophic effect by increasing BDNF protein expression. It also diminished reactive oxygen species production and activated the redox sensitive transcription factor NRF2 and increased its target genes expression. Knockdown of NRF2 abolished neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium. Furthermore, lithium significantly decreased both basal and PQ-induced expression of miR-34a. Transfection of miR-34a specific mimic reversed neuroprotective, anti-apoptotic, and anti-oxidant effects of lithium against PQ-toxicity. Our results revealed two novel mechanisms of lithium neuroprotection, namely NRF2 activation and miR-34a suppression.

No MeSH data available.


Related in: MedlinePlus