Limits...
Acid sphingomyelinase activity triggers microparticle release from glial cells.

Bianco F, Perrotta C, Novellino L, Francolini M, Riganti L, Menna E, Saglietti L, Schuchman EH, Furlan R, Clementi E, Matteoli M, Verderio C - EMBO J. (2009)

Bottom Line: ATP-induced shedding and IL-1beta release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice.We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL-1beta release.Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL-1beta release, thus, opening new strategies for the treatment of neuroinflammatory diseases.

View Article: PubMed Central - PubMed

Affiliation: CNR Institute of Neuroscience and Department of Medical Pharmacology, University of Milano, Milano, Italy.

ABSTRACT
We have earlier shown that microglia, the immune cells of the CNS, release microparticles from cell plasma membrane after ATP stimulation. These vesicles contain and release IL-1beta, a crucial cytokine in CNS inflammatory events. In this study, we show that microparticles are also released by astrocytes and we get insights into the mechanism of their shedding. We show that, on activation of the ATP receptor P2X7, microparticle shedding is associated with rapid activation of acid sphingomyelinase, which moves to plasma membrane outer leaflet. ATP-induced shedding and IL-1beta release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice. We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL-1beta release. Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL-1beta release, thus, opening new strategies for the treatment of neuroinflammatory diseases.

Show MeSH

Related in: MedlinePlus

A-SMase activity does not control pore opening. (A) Time course analysis of Yo-PRO-1 uptake in astrocytes exposed to 100 μM BzATP in the presence/absence of A-SMase inhibitors. (B) Time course analysis of Yo-PRO-1 uptake in astrocytes from A-SMase wild type (+/+), heterozygous (+/−) or KO (−/−) animals on 100 μM BzATP exposure. (n=2; **P<0.01 versus controls, ANOVA analysis, Dunnett's method).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2664656&req=5

f6: A-SMase activity does not control pore opening. (A) Time course analysis of Yo-PRO-1 uptake in astrocytes exposed to 100 μM BzATP in the presence/absence of A-SMase inhibitors. (B) Time course analysis of Yo-PRO-1 uptake in astrocytes from A-SMase wild type (+/+), heterozygous (+/−) or KO (−/−) animals on 100 μM BzATP exposure. (n=2; **P<0.01 versus controls, ANOVA analysis, Dunnett's method).

Mentions: We then investigated whether p38-dependent activation of A-SMase could be involved in P2X7-induced pore formation. An efficient YO-PRO-1 uptake was observed in microglial pre-treated with imipramine (Figure 6A; Supplementary data), indicating that A-SMase inhibition does not alter the capability of P2X7R to open large membrane pores. Accordingly, no significant differences in YO-PRO-1 uptake kinetics were detected in astrocytes from A-SMase KO and heterozygous mice as compared with astrocytes from wild-type animals (Figure 6B). These data suggest that MP shedding and pore opening are two P2X7-dependent processes that are regulated by two parallel pathways, downstream of p38 phosphorylation (Figure 7).


Acid sphingomyelinase activity triggers microparticle release from glial cells.

Bianco F, Perrotta C, Novellino L, Francolini M, Riganti L, Menna E, Saglietti L, Schuchman EH, Furlan R, Clementi E, Matteoli M, Verderio C - EMBO J. (2009)

A-SMase activity does not control pore opening. (A) Time course analysis of Yo-PRO-1 uptake in astrocytes exposed to 100 μM BzATP in the presence/absence of A-SMase inhibitors. (B) Time course analysis of Yo-PRO-1 uptake in astrocytes from A-SMase wild type (+/+), heterozygous (+/−) or KO (−/−) animals on 100 μM BzATP exposure. (n=2; **P<0.01 versus controls, ANOVA analysis, Dunnett's method).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2664656&req=5

f6: A-SMase activity does not control pore opening. (A) Time course analysis of Yo-PRO-1 uptake in astrocytes exposed to 100 μM BzATP in the presence/absence of A-SMase inhibitors. (B) Time course analysis of Yo-PRO-1 uptake in astrocytes from A-SMase wild type (+/+), heterozygous (+/−) or KO (−/−) animals on 100 μM BzATP exposure. (n=2; **P<0.01 versus controls, ANOVA analysis, Dunnett's method).
Mentions: We then investigated whether p38-dependent activation of A-SMase could be involved in P2X7-induced pore formation. An efficient YO-PRO-1 uptake was observed in microglial pre-treated with imipramine (Figure 6A; Supplementary data), indicating that A-SMase inhibition does not alter the capability of P2X7R to open large membrane pores. Accordingly, no significant differences in YO-PRO-1 uptake kinetics were detected in astrocytes from A-SMase KO and heterozygous mice as compared with astrocytes from wild-type animals (Figure 6B). These data suggest that MP shedding and pore opening are two P2X7-dependent processes that are regulated by two parallel pathways, downstream of p38 phosphorylation (Figure 7).

Bottom Line: ATP-induced shedding and IL-1beta release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice.We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL-1beta release.Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL-1beta release, thus, opening new strategies for the treatment of neuroinflammatory diseases.

View Article: PubMed Central - PubMed

Affiliation: CNR Institute of Neuroscience and Department of Medical Pharmacology, University of Milano, Milano, Italy.

ABSTRACT
We have earlier shown that microglia, the immune cells of the CNS, release microparticles from cell plasma membrane after ATP stimulation. These vesicles contain and release IL-1beta, a crucial cytokine in CNS inflammatory events. In this study, we show that microparticles are also released by astrocytes and we get insights into the mechanism of their shedding. We show that, on activation of the ATP receptor P2X7, microparticle shedding is associated with rapid activation of acid sphingomyelinase, which moves to plasma membrane outer leaflet. ATP-induced shedding and IL-1beta release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice. We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL-1beta release. Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL-1beta release, thus, opening new strategies for the treatment of neuroinflammatory diseases.

Show MeSH
Related in: MedlinePlus