Alkaline sphingomyelinase (alk-SMase) is a new member of the NPP (nucleotide pyrophosphatase/phosphodiesterase) family that hydrolyses SM (sphingomyelin) to generate ceramide in the intestinal tract. The enzyme may protect the intestinal mucosa from inflammation and tumorigenesis. PAF (platelet-activating factor) is a pro-inflammatory phospholipid involved in pathogenesis of inflammatory bowel diseases. We examined whether alk-SMase can hydrolyse and inactivate PAF. [3H]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 alkSMase 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 Vmax for PAF hydrolysis was 374 μmol·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.
Skip Nav Destination
Article navigation
February 2006
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
January 27 2006
Intestinal alkaline sphingomyelinase hydrolyses and inactivates platelet-activating factor by a phospholipase C activity
Jun Wu;
Jun Wu
*Gastroenterology Laboratory, Biomedical Centre, B11, Lund University, S-221 84 Lund, Sweden
Search for other works by this author on:
Åke Nilsson;
Åke Nilsson
*Gastroenterology Laboratory, Biomedical Centre, B11, Lund University, S-221 84 Lund, Sweden
Search for other works by this author on:
Bo A. G. Jönsson;
Bo A. G. Jönsson
†Department of Occupational and Environment Medicine, Institute of Laboratory Medicine, University Hospital, S-221 85 Lund, Sweden
Search for other works by this author on:
Hanna Stenstad;
Hanna Stenstad
‡Immunology Unit, Lund University, S-221 84 Lund, Sweden
Search for other works by this author on:
William Agace;
William Agace
‡Immunology Unit, Lund University, S-221 84 Lund, Sweden
Search for other works by this author on:
Yajun Cheng;
Yajun Cheng
*Gastroenterology Laboratory, Biomedical Centre, B11, Lund University, S-221 84 Lund, Sweden
Search for other works by this author on:
Rui-Dong Duan
Rui-Dong Duan
1
*Gastroenterology Laboratory, Biomedical Centre, B11, Lund University, S-221 84 Lund, Sweden
1To whom correspondence should be addressed (email rui-dong.duan@med.lu.se).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
July 12 2005
Revision Received:
October 12 2005
Accepted:
October 31 2005
Accepted Manuscript online:
October 31 2005
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2006
Biochem J (2006) 394 (1): 299–308.
Article history
Received:
July 12 2005
Revision Received:
October 12 2005
Accepted:
October 31 2005
Accepted Manuscript online:
October 31 2005
Citation
Jun Wu, Åke Nilsson, Bo A. G. Jönsson, Hanna Stenstad, William Agace, Yajun Cheng, Rui-Dong Duan; Intestinal alkaline sphingomyelinase hydrolyses and inactivates platelet-activating factor by a phospholipase C activity. Biochem J 15 February 2006; 394 (1): 299–308. doi: https://doi.org/10.1042/BJ20051121
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.