- J Biol Chem. 2008 Sep 26;283(39):26509-17. Epub 2008 Aug 1.
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Ceramide Generated by Sphingomyelin Hydrolysis and the Salvage Pathway Is Involved in Hypoxia/Reoxygenation-induced Bax Redistribution to Mitochondria in NT-2 Cells.
Departments of Biochemistry and Molecular Biology, Medicine, and Pediatrics, Division of Hematology/Oncology, Medical University of South Carolina, Charleston, South Carolina 29425.
Ceramide functions as an important second messenger in apoptosis signaling pathways. In this report, we show that treatment of NT-2 neuronal precursor cells with hypoxia/reoxygenation (H/R) resulted in ceramide up-regulation. This elevation in ceramide was primarily due to the actions of acid sphingomyelinase and ceramide synthase LASS 5, demonstrating the action of the salvage pathway. Hypoxia/reoxygenation treatment led to Bax translocation from the cytoplasm to mitochondria and cytochrome c release from mitochondria. Down-regulation of either acid sphingomyelinase or LASS 5-attenuated ceramide accumulation and H/R-induced Bax translocation to mitochondria. Overall, we have demonstrated that ceramide up-regulation following H/R is pertinent to Bax activation to promote cell death.
PMID: 18676372 [PubMed - in process]
NT-2 cells, 12-hour hypoxia, 0, 12, 24h reoxygenation
HPLC-MS, qPCR (quantitative PCR = realtime PCR??)
H/R results in ceramide accumulation, sphingomyelin decrease
C14, C16 ceramides most changed
C18:1, C20 also altered
C24 no change
suggests involvement of LASS5 or LASS6
LASS5 (by aPCR and siRNA) siRNA attenuates Bax activation, cyt c release, caspase3 activation
aSMase but not nSMase activity up, and mRNA up, siRNA attenuates Bax activation, therefore ceramide signal from salvage pathway
fumonisin B1 inhibits both de novo and salvage pathway generation of ceramide
how does aSMase get activated? coordinated with LASS5 activation? probably
how does increased short(er) chain ceramides activate Bax? not through Bid
