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Ethanol-mediated activation of the NLRP3 inflammasome in iPS cells and iPS cells-derived neural progenitor cells.

De Filippis L, Halikere A, McGowan H, Moore JC, Tischfield JA, Hart RP, Pang ZP - Mol Brain (2016)

Bottom Line: Ethanol exposure for 24 hours or 7 days does not affect the proliferation of iPS cells and NPCs, but primes an innate immune-like response by activating the NLR family pyrin domain containing 3 (NLRP3) inflammasome pathway.This leads to an increase of microtubule-associated protein 1A/1B-light chain 3(+) (LC3B(+)) autophagic puncta and impairment of the mitochondrial and lysosomal distribution.In addition, a decrease of mature neurons derived from differentiating NPCs is evident in ethanol pre-exposed compared to control NPCs.

View Article: PubMed Central - PubMed

Affiliation: Child Health Institute of New Jersey, Rutgers University-Robert Wood Johnson Medical School, room 3233D, 89 French Street, New Brunswick, NJ, 08901, USA. ld473@rwjms.rutgers.edu.

ABSTRACT

Background: Alcohol abuse produces an enormous impact on health, society, and the economy. Currently, there are very limited therapies available, largely due to the poor understanding of mechanisms underlying alcohol use disorders (AUDs) in humans. Oxidative damage of mitochondria and cellular proteins aggravates the progression of neuroinflammation and neurological disorders initiated by alcohol abuse.

Results: Here we show that ethanol exposure causes neuroinflammation in both human induced pluripotent stem (iPS) cells and human neural progenitor cells (NPCs). Ethanol exposure for 24 hours or 7 days does not affect the proliferation of iPS cells and NPCs, but primes an innate immune-like response by activating the NLR family pyrin domain containing 3 (NLRP3) inflammasome pathway. This leads to an increase of microtubule-associated protein 1A/1B-light chain 3(+) (LC3B(+)) autophagic puncta and impairment of the mitochondrial and lysosomal distribution. In addition, a decrease of mature neurons derived from differentiating NPCs is evident in ethanol pre-exposed compared to control NPCs. Moreover, a second insult of a pro-inflammatory factor in addition to ethanol preexposure enhances innate cellular inflammation in human iPS cells.

Conclusions: This study provides strong evidence that neuronal inflammation contributes to the pathophysiology of AUDs through the activation of the inflammasome pathway in human cellular models.

No MeSH data available.


Related in: MedlinePlus

Cooperative effects of ethanol and peroxide challenges on inflammasome markers NLRP3 and Casp1 in iPS cells. On day 7 following 24hr or 7d ethanol treatment, iPS cells were exposed for 14hr to 5 or 10μM H2O2 and immunostained with antibodies against NLRP3 and Casp1 (a), and LC3B (b). At 10μM H2O2, iPS cells pretreated with ethanol for 7d displayed a dramatic increase in death. Scale bars: 50 μm (a), 10μm (b)
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Fig7: Cooperative effects of ethanol and peroxide challenges on inflammasome markers NLRP3 and Casp1 in iPS cells. On day 7 following 24hr or 7d ethanol treatment, iPS cells were exposed for 14hr to 5 or 10μM H2O2 and immunostained with antibodies against NLRP3 and Casp1 (a), and LC3B (b). At 10μM H2O2, iPS cells pretreated with ethanol for 7d displayed a dramatic increase in death. Scale bars: 50 μm (a), 10μm (b)

Mentions: Accordingly, we observed a cumulative increase of the inflammasome-related markers Casp1 and NLRP3 (Fig. 7), of Casp3+ cells (Fig. 6a and 8a), and of LC3B puncta (Fig. 7b and 8b) in iPS cells that had undergone the double challenge compared to the single challenge (Fig. 6 and 7), showing that ethanol treatment induces long-term and long-lasting metabolic changes in the cell that can drive an enhanced response to any additional damage. On the contrary, while an increase in the number of Casp3+ cells was evident with peroxide or ethanol treatment alone in NPCs, no significant difference was detectable between cells that had undergone the double challenge compared to a single challenge (Fig. 9). This suggests that NPCs are more resilient than iPS cells to cumulative damages and/or that in our cell-based system the range of sensitivity is too narrow to reach statistical significance. Consistently, LC3B puncta appeared increased by each single challenge, but a quantitative evaluation in double challenged cells was impaired by the altered cell morphology. These data suggest that ethanol exposure in iPS cells and NPCs results in greater sensitivity to oxidative stress, which may contribute to the pathophysiology of neurogenesis in humans.Fig. 7


Ethanol-mediated activation of the NLRP3 inflammasome in iPS cells and iPS cells-derived neural progenitor cells.

De Filippis L, Halikere A, McGowan H, Moore JC, Tischfield JA, Hart RP, Pang ZP - Mol Brain (2016)

Cooperative effects of ethanol and peroxide challenges on inflammasome markers NLRP3 and Casp1 in iPS cells. On day 7 following 24hr or 7d ethanol treatment, iPS cells were exposed for 14hr to 5 or 10μM H2O2 and immunostained with antibodies against NLRP3 and Casp1 (a), and LC3B (b). At 10μM H2O2, iPS cells pretreated with ethanol for 7d displayed a dramatic increase in death. Scale bars: 50 μm (a), 10μm (b)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC4862119&req=5

Fig7: Cooperative effects of ethanol and peroxide challenges on inflammasome markers NLRP3 and Casp1 in iPS cells. On day 7 following 24hr or 7d ethanol treatment, iPS cells were exposed for 14hr to 5 or 10μM H2O2 and immunostained with antibodies against NLRP3 and Casp1 (a), and LC3B (b). At 10μM H2O2, iPS cells pretreated with ethanol for 7d displayed a dramatic increase in death. Scale bars: 50 μm (a), 10μm (b)
Mentions: Accordingly, we observed a cumulative increase of the inflammasome-related markers Casp1 and NLRP3 (Fig. 7), of Casp3+ cells (Fig. 6a and 8a), and of LC3B puncta (Fig. 7b and 8b) in iPS cells that had undergone the double challenge compared to the single challenge (Fig. 6 and 7), showing that ethanol treatment induces long-term and long-lasting metabolic changes in the cell that can drive an enhanced response to any additional damage. On the contrary, while an increase in the number of Casp3+ cells was evident with peroxide or ethanol treatment alone in NPCs, no significant difference was detectable between cells that had undergone the double challenge compared to a single challenge (Fig. 9). This suggests that NPCs are more resilient than iPS cells to cumulative damages and/or that in our cell-based system the range of sensitivity is too narrow to reach statistical significance. Consistently, LC3B puncta appeared increased by each single challenge, but a quantitative evaluation in double challenged cells was impaired by the altered cell morphology. These data suggest that ethanol exposure in iPS cells and NPCs results in greater sensitivity to oxidative stress, which may contribute to the pathophysiology of neurogenesis in humans.Fig. 7

Bottom Line: Ethanol exposure for 24 hours or 7 days does not affect the proliferation of iPS cells and NPCs, but primes an innate immune-like response by activating the NLR family pyrin domain containing 3 (NLRP3) inflammasome pathway.This leads to an increase of microtubule-associated protein 1A/1B-light chain 3(+) (LC3B(+)) autophagic puncta and impairment of the mitochondrial and lysosomal distribution.In addition, a decrease of mature neurons derived from differentiating NPCs is evident in ethanol pre-exposed compared to control NPCs.

View Article: PubMed Central - PubMed

Affiliation: Child Health Institute of New Jersey, Rutgers University-Robert Wood Johnson Medical School, room 3233D, 89 French Street, New Brunswick, NJ, 08901, USA. ld473@rwjms.rutgers.edu.

ABSTRACT

Background: Alcohol abuse produces an enormous impact on health, society, and the economy. Currently, there are very limited therapies available, largely due to the poor understanding of mechanisms underlying alcohol use disorders (AUDs) in humans. Oxidative damage of mitochondria and cellular proteins aggravates the progression of neuroinflammation and neurological disorders initiated by alcohol abuse.

Results: Here we show that ethanol exposure causes neuroinflammation in both human induced pluripotent stem (iPS) cells and human neural progenitor cells (NPCs). Ethanol exposure for 24 hours or 7 days does not affect the proliferation of iPS cells and NPCs, but primes an innate immune-like response by activating the NLR family pyrin domain containing 3 (NLRP3) inflammasome pathway. This leads to an increase of microtubule-associated protein 1A/1B-light chain 3(+) (LC3B(+)) autophagic puncta and impairment of the mitochondrial and lysosomal distribution. In addition, a decrease of mature neurons derived from differentiating NPCs is evident in ethanol pre-exposed compared to control NPCs. Moreover, a second insult of a pro-inflammatory factor in addition to ethanol preexposure enhances innate cellular inflammation in human iPS cells.

Conclusions: This study provides strong evidence that neuronal inflammation contributes to the pathophysiology of AUDs through the activation of the inflammasome pathway in human cellular models.

No MeSH data available.


Related in: MedlinePlus