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RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells

BACKGROUND: Intracerebral hemorrhage (ICH) is a devastating neurological injury associated with significant mortality. Necroptosis is a newly identified type of programmed necrosis initiated by the activation of tumor necrosis factor alpha. Evidences had demonstrated the importance of necroptosis in...

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Autores principales: Su, Xingfen, Wang, Handong, Lin, Yuanxiang, Chen, Fuxiang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Dove Medical Press 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6247969/
https://www.ncbi.nlm.nih.gov/pubmed/30532542
http://dx.doi.org/10.2147/NDT.S181074
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author Su, Xingfen
Wang, Handong
Lin, Yuanxiang
Chen, Fuxiang
author_facet Su, Xingfen
Wang, Handong
Lin, Yuanxiang
Chen, Fuxiang
author_sort Su, Xingfen
collection PubMed
description BACKGROUND: Intracerebral hemorrhage (ICH) is a devastating neurological injury associated with significant mortality. Necroptosis is a newly identified type of programmed necrosis initiated by the activation of tumor necrosis factor alpha. Evidences had demonstrated the importance of necroptosis in neuronal cell death. Necrostatin-1 is a specific inhibitor of necroptosis. The present study was carried out to explore whether RIP1/RIP3 pathways participate in hemin induced cell death in HT-22 hippocampal neuronal cells and investigate the potential neuroprotection of necrostatin-1 in hemin induced cell death in HT-22. METHODS: First, different concentrations of hemin (0, 25, 50, 100 μmol/L) were added to HT-22 cells. Propidium iodide (PI) positive cells and cell viability were measured at 24 hours after hemin treatment. Then, necrostatin-1, pan-caspase inhibitor Benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (z-VAD-fmk) and reactive oxygen species (ROS) scavenger butylated hydroxyanisole (BHA) were applied to hemin-treated HT-22 cells. PI positive cells and cell viability were measured at 24 hours after hemin treatment. MitoSox Red was used to indicate ROS level. Last, the effect of RIP3 in hemin induced HT-22 cell death was explored through RIP3 knockdown using siRNA. PI positive cells, cell viability and ROS lever were measured at 24 h after hemin treatment. RESULTS: Hemin could induce a dose dependent cell death in HT22 neural cells. RIP1 specific inhibitor necrostatin-1 significantly inhibited cell death induced by hemin in HT-22 cells, greatly reducing PI positive cells, dramatically improving cell viability and decreasing ROS accumulation. BHA could significantly inhibit PI positive cells induced by hemin in HT-22 cells. Furthermore, silencing of RIP3 using siRNA attenuated hemin induced cell death in HT-22 cells, greatly reducing PI positive cells, dramatically improving cell viability and decreasing ROS accumulation. CONCLUSION: These data revealed that RIP1/RIP3 might mediate hemin induced cell death in HT-22 cells, and necrostatin-1 played a neuroprotection role in hemin induced cell death in HT-22. RIP1 and RIP3 might represent novel therapeutic targets for ICH.
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spelling pubmed-62479692018-12-07 RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells Su, Xingfen Wang, Handong Lin, Yuanxiang Chen, Fuxiang Neuropsychiatr Dis Treat Original Research BACKGROUND: Intracerebral hemorrhage (ICH) is a devastating neurological injury associated with significant mortality. Necroptosis is a newly identified type of programmed necrosis initiated by the activation of tumor necrosis factor alpha. Evidences had demonstrated the importance of necroptosis in neuronal cell death. Necrostatin-1 is a specific inhibitor of necroptosis. The present study was carried out to explore whether RIP1/RIP3 pathways participate in hemin induced cell death in HT-22 hippocampal neuronal cells and investigate the potential neuroprotection of necrostatin-1 in hemin induced cell death in HT-22. METHODS: First, different concentrations of hemin (0, 25, 50, 100 μmol/L) were added to HT-22 cells. Propidium iodide (PI) positive cells and cell viability were measured at 24 hours after hemin treatment. Then, necrostatin-1, pan-caspase inhibitor Benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone (z-VAD-fmk) and reactive oxygen species (ROS) scavenger butylated hydroxyanisole (BHA) were applied to hemin-treated HT-22 cells. PI positive cells and cell viability were measured at 24 hours after hemin treatment. MitoSox Red was used to indicate ROS level. Last, the effect of RIP3 in hemin induced HT-22 cell death was explored through RIP3 knockdown using siRNA. PI positive cells, cell viability and ROS lever were measured at 24 h after hemin treatment. RESULTS: Hemin could induce a dose dependent cell death in HT22 neural cells. RIP1 specific inhibitor necrostatin-1 significantly inhibited cell death induced by hemin in HT-22 cells, greatly reducing PI positive cells, dramatically improving cell viability and decreasing ROS accumulation. BHA could significantly inhibit PI positive cells induced by hemin in HT-22 cells. Furthermore, silencing of RIP3 using siRNA attenuated hemin induced cell death in HT-22 cells, greatly reducing PI positive cells, dramatically improving cell viability and decreasing ROS accumulation. CONCLUSION: These data revealed that RIP1/RIP3 might mediate hemin induced cell death in HT-22 cells, and necrostatin-1 played a neuroprotection role in hemin induced cell death in HT-22. RIP1 and RIP3 might represent novel therapeutic targets for ICH. Dove Medical Press 2018-11-15 /pmc/articles/PMC6247969/ /pubmed/30532542 http://dx.doi.org/10.2147/NDT.S181074 Text en © 2018 Su et al. This work is published and licensed by Dove Medical Press Limited The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution – Non Commercial (unported, v3.0) License (http://creativecommons.org/licenses/by-nc/3.0/). By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed.
spellingShingle Original Research
Su, Xingfen
Wang, Handong
Lin, Yuanxiang
Chen, Fuxiang
RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title_full RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title_fullStr RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title_full_unstemmed RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title_short RIP1 and RIP3 mediate hemin-induced cell death in HT22 hippocampal neuronal cells
title_sort rip1 and rip3 mediate hemin-induced cell death in ht22 hippocampal neuronal cells
topic Original Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6247969/
https://www.ncbi.nlm.nih.gov/pubmed/30532542
http://dx.doi.org/10.2147/NDT.S181074
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