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Intestinal alkaline sphingomyelinase hydrolyses and inactivates platelet-activating factor by a phospholipase C activity

Wu, Jun LU ; Nilsson, Åke LU ; Jönsson, Bo A LU ; Stenstad, Hanna LU ; Agace, William LU ; Cheng, Yajun LU and Duan, Rui-Dong LU (2006) In Biochemical Journal 394. p.299-308
Abstract
Alkaline sphingomyelinase (alk-SMase) is a new member of the NPP (nucleotide pyrophophatase/phosphodiesterase) family that hydrolyses SM (sphingomyelin) to generate ceramide ill the intestinal tract. The enzyme may protect the intestinal mucosa front inflammation and tumorigenesis. PAF (platelet-activating factor) is a pro-inflammatory phospholipid involved in pathogenesis of inflammatory bowel diseases. We examined whether alk-SMase call hydrolyse and inactivate PAF. [H-3]Octadecyl-labelled PAF was incubated With purified rat intestinal alk-SMase or recombinant human alk-SMase expressed in COS-7 cells. The hydrolytic products were assayed with TLC and MS. We found that alk-SMase cleaved the phosphocholine head group from PAF and generated... (More)
Alkaline sphingomyelinase (alk-SMase) is a new member of the NPP (nucleotide pyrophophatase/phosphodiesterase) family that hydrolyses SM (sphingomyelin) to generate ceramide ill the intestinal tract. The enzyme may protect the intestinal mucosa front inflammation and tumorigenesis. PAF (platelet-activating factor) is a pro-inflammatory phospholipid involved in pathogenesis of inflammatory bowel diseases. We examined whether alk-SMase call hydrolyse and inactivate PAF. [H-3]Octadecyl-labelled PAF was incubated With purified rat intestinal alk-SMase or recombinant human alk-SMase expressed in COS-7 cells. The hydrolytic products were assayed with TLC and MS. We found that alk-SMase cleaved the phosphocholine head group from PAF and generated 1-O-alkyl-2-acetyl-sn-glycerol. Differing from the activity against SM, the activity against PAF was optimal at pH 7.5, inhibited by EDTA and stimulated by 0.1-0.25 mM Zn2+. The activity was abolished by site Mutation of the predicted metal-binding sites that are conserved in all NPP members. Similar to the activity against SM, the activity against PAF was dependent on bile salt, particularly taurocholate and taurochenodeoxycholate. The V-max for PAF hydrolysis was 374 mu mol . h(-1) . (mg of protein)(-1). The hydrolysis of PAF and SM could be inhibited by the presence of SM and PAF respectively, the inhibition of PAF hydrolysis by SM being stronger. The PAF-induced MAPK (mitogen-activated protein kinase) activation and IL-8 (interleukin 8) release in HT-29 cells, and chemotaxis in leucocytes were abolished by alk-SMase treatment. In Conclusion, alk-SMase hydrolyses and inactivates PAF by a phospholipase C activity. The finding reveals a novel function, by which alk-SMase may counteract the development of intestinal inflammation and colon cancer. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
platelet-activating factor, mitogen-activated protein kinase (MAPK), inflammatory bowel disease, alkaline sphingomyelinase, colon cancer, phospholipase C
in
Biochemical Journal
volume
394
pages
299 - 308
publisher
Portland Press Limited
external identifiers
  • wos:000235476700033
  • pmid:16255717
  • scopus:32944473705
ISSN
0264-6021
DOI
10.1042/BJ20051121
language
English
LU publication?
yes
id
adfe2dbf-3df2-48f4-90d3-129cce7b4e49 (old id 417267)
date added to LUP
2007-10-21 07:21:31
date last changed
2019-05-14 03:17:19
@article{adfe2dbf-3df2-48f4-90d3-129cce7b4e49,
  abstract     = {Alkaline sphingomyelinase (alk-SMase) is a new member of the NPP (nucleotide pyrophophatase/phosphodiesterase) family that hydrolyses SM (sphingomyelin) to generate ceramide ill the intestinal tract. The enzyme may protect the intestinal mucosa front inflammation and tumorigenesis. PAF (platelet-activating factor) is a pro-inflammatory phospholipid involved in pathogenesis of inflammatory bowel diseases. We examined whether alk-SMase call hydrolyse and inactivate PAF. [H-3]Octadecyl-labelled PAF was incubated With purified rat intestinal alk-SMase or recombinant human alk-SMase expressed in COS-7 cells. The hydrolytic products were assayed with TLC and MS. We found that alk-SMase cleaved the phosphocholine head group from PAF and generated 1-O-alkyl-2-acetyl-sn-glycerol. Differing from the activity against SM, the activity against PAF was optimal at pH 7.5, inhibited by EDTA and stimulated by 0.1-0.25 mM Zn2+. The activity was abolished by site Mutation of the predicted metal-binding sites that are conserved in all NPP members. Similar to the activity against SM, the activity against PAF was dependent on bile salt, particularly taurocholate and taurochenodeoxycholate. The V-max for PAF hydrolysis was 374 mu mol . h(-1) . (mg of protein)(-1). The hydrolysis of PAF and SM could be inhibited by the presence of SM and PAF respectively, the inhibition of PAF hydrolysis by SM being stronger. The PAF-induced MAPK (mitogen-activated protein kinase) activation and IL-8 (interleukin 8) release in HT-29 cells, and chemotaxis in leucocytes were abolished by alk-SMase treatment. In Conclusion, alk-SMase hydrolyses and inactivates PAF by a phospholipase C activity. The finding reveals a novel function, by which alk-SMase may counteract the development of intestinal inflammation and colon cancer.},
  author       = {Wu, Jun and Nilsson, Åke and Jönsson, Bo A and Stenstad, Hanna and Agace, William and Cheng, Yajun and Duan, Rui-Dong},
  issn         = {0264-6021},
  keyword      = {platelet-activating factor,mitogen-activated protein kinase (MAPK),inflammatory bowel disease,alkaline sphingomyelinase,colon cancer,phospholipase C},
  language     = {eng},
  pages        = {299--308},
  publisher    = {Portland Press Limited},
  series       = {Biochemical Journal},
  title        = {Intestinal alkaline sphingomyelinase hydrolyses and inactivates platelet-activating factor by a phospholipase C activity},
  url          = {http://dx.doi.org/10.1042/BJ20051121},
  volume       = {394},
  year         = {2006},
}