Abstract
Alkaline sphingomyelinase (Alk-SMase) is a key enzyme in the intestinal tract for digestion of dietary sphingomyelin (SM), which generates lipid messengers with cell-cycle regulating effects. The enzyme is significantly decreased in ulcerative colitis and colon cancer. Based on this information, we wanted to investigate whether the enzyme had preventive effects against murine colitis. We report herein a method to express a biologically active Alk-SMase from Pichia pastoris yeast cells. By using the expressed enzyme to treat a rat colitis model induced by dextran sulfate sodium, we found that intrarectal instillation of Alk-SMase once daily for 1 week significantly reduced the inflammation score and protected the colonic epithelium from inflammatory destruction. We found a tendency for decreased tumor necrosis factor (TNF)-α expression in the Alk-SMase-treated group. This study, for the first time, provides a method to produce the enzyme and shows the potential applicability of the enzyme in the treatment of inflammatory bowel diseases.
Similar content being viewed by others
References
Xavier RJ, Podolsky DK. Unravelling the pathogenesis of inflammatory bowel disease. Nature. 2007;448:427–434. doi:10.1038/nature06005.
Duan RD, Hertervig E, Nyberg L, et al. Distribution of alkaline sphingomyelinase activity in human beings and animals. Tissue and species differences. Dig Dis Sci. 1996;41:1801–1806. doi:10.1007/BF02088748.
Lundgren P, Nilsson Å, Duan RD. Distribution and properties of neutral ceramidase activity in rat intestinal tract. Dig Dis Sci. 2001;46:765–772. doi:10.1023/A:1010792031910.
Hannun YA, Obeid LM. Principles of bioactive lipid signalling: lessons from sphingolipids. Nat Rev Mol Cell Biol. 2008;9:139–150. doi:10.1038/nrm2329.
Nilsson Å. The presence of sphingomyelin- and ceramide-cleaving enzymes in the small intestinal tract. Biochim Biophys Acta. 1969;176:339–347.
Duan RD, Cheng Y, Hansen G, et al. Purification, localization, and expression of human intestinal alkaline sphingomyelinase. J Lipid Res. 2003;44:1241–1250. doi:10.1194/jlr.M300037-JLR200.
Duan RD, Bergman T, Xu N, et al. Identification of human intestinal alkaline sphingomyelinase as a novel ecto-enzyme related to the nucleotide phosphodiesterase family. J Biol Chem. 2003;278:38528–38536. doi:10.1074/jbc.M305437200.
Hertervig E, Nilsson A, Nyberg L, Duan RD. Alkaline sphingomyelinase activity is decreased in human colorectal carcinoma. Cancer. 1997;79:448–453.
Wu J, Cheng Y, Nilsson A, Duan RD. Identification of one exon deletion of intestinal alkaline sphingomyelinase in colon cancer HT-29 cells and a differentiation-related expression of the wild-type enzyme in Caco-2 cells. Carcinogenesis. 2004;25:1327–1333. doi:10.1093/carcin/bgh140.
Cheng Y, Wu J, Hertervig E, et al. Identification of aberrant forms of alkaline sphingomyelinase (NPP7) associated with human liver tumorigenesis. Br J Cancer. 2007;97:1441–1448. doi:10.1038/sj.bjc.6604013.
Sjöqvist U, Hertervig E, Nilsson A, et al. Chronic colitis is associated with a reduction of mucosal alkaline sphingomyelinase activity. Inflamm Bowel Dis. 2002;8:258–263. doi:10.1097/00054725-200207000-00004.
Wu J, Nilsson A, Jonsson BA, et al. Intestinal alkaline sphingomyelinase hydrolyses and inactivates platelet-activating factor by a phospholipase C activity. Biochem J. 2006;394:299–308. doi:10.1042/BJ20051121.
Stoffel W. Chemical synthesis of choline-labeled lecithins and sphingomyelin. Methods Enzymol. 1975;36:533–541. doi:10.1016/0076-6879(75)35181-1.
Blank M, Cress EA, Smith ZL, Snyder F. Meats and fish consumed in the American diet contain substantial amounts of ether-linked phospholipids. J Nutr. 1992;122:1656–1661.
Wallace JL, MacNaughton WK, Morris GP, Beck PL. Inhibition of leukotriene synthesis markedly accelerates healing in a rat model of inflammatory bowel disease. Gastroenterology. 1989;96:29–36.
Frodin M, Sekine N, Roche E, et al. Glucose, other secretagogues, and nerve growth factor stimulate mitogen- activated protein kinase in the insulin-secreting beta-cell line, INS-1. J Biol Chem. 1995;270:7882–7889. doi:10.1074/jbc.270.14.7882.
Kotarsky K, Boketoft A, Bristulf J, et al. Lysophosphatidic acid binds to and activates GPR92, a G protein-coupled receptor highly expressed in gastrointestinal lymphocytes. J Pharmacol Exp Ther. 2006;318:619–628. doi:10.1124/jpet.105.098848.
Peixoto A, Monteiro M, Rocha B, Veiga-Fernandes H. Quantification of multiple gene expression in individual cells. Genome Res. 2004;14:1938–1947. doi:10.1101/gr.2890204.
Wu J, Cheng Y, Palmberg C, Bergman T, Nilsson A, Duan RD. Cloning of alkaline sphingomyelinase from rat intestinal mucosa and adjusting of the hypothetical protein XP_221184 in GenBank. Biochim Biophys Acta. 2005;1687:94–102.
Cheng Y, Nilsson Å, Tömquist E, Duan RD. Purification, characterization and expression of rat intestinal alkaline sphingomyelinase. J Lipid Res. 2002;43:316–324.
Cereghino GP, Cereghino JL, Ilgen C, Cregg JM. Production of recombinant proteins in fermenter cultures of the yeast Pichia pastoris. Curr Opin Biotechnol. 2002;13:329–332. doi:10.1016/S0958-1669(02)00330-0.
Wu J, Hansen GH, Nilsson A, Duan RD. Functional studies of human intestinal alkaline sphingomyelinase by deglycosylation and mutagenesis. Biochem J. 2005;386:153–160. doi:10.1042/BJ20041455.
Klotz U, Schwab M. Topical delivery of therapeutic agents in the treatment of inflammatory bowel disease. Adv Drug Deliv Rev. 2005;57:267–279. doi:10.1016/j.addr.2004.08.007.
Gaudio E, Taddei G, Vetuschi A, et al. Dextran sulfate sodium (DSS) colitis in rats: clinical, structural, and ultrastructural aspects. Dig Dis Sci. 1999;44:1458–1475. doi:10.1023/A:1026620322859.
Weber CR, Turner JR. Inflammatory bowel disease: is it really just another break in the wall? Gut. 2007;56:6–8. doi:10.1136/gut.2006.104182.
Jarnerot G, Hertervig E, Friis-Liby I, et al. Infliximab as rescue therapy in severe to moderately severe ulcerative colitis: a randomized, placebo-controlled study. Gastroenterology. 2005;128:1805–1811. doi:10.1053/j.gastro.2005.03.003.
Maines LW, Fitzpatrick LR, French KJ, et al. Suppression of ulcerative colitis in mice by orally available inhibitors of sphingosine kinase. Dig Dis Sci. 2008;53:997–1012. doi:10.1007/s10620-007-0133-6.
Nassif A, Longo WE, Mazuski JE, Vernava AM, Kaminski DL. Role of cytokines and platelet-activating factor in inflammatory bowel disease. Implications for therapy. Dis Colon Rectum. 1996;39:217–223. doi:10.1007/BF02068079.
Ewer AK. Role of platelet-activating factor in the pathophysiology of necrotizing enterocolitis. Acta Paediatr Suppl. 2002;91:2–5. doi:10.1080/08035250260095717.
Saslowsky DE, Lencer WI. Conversion of apical plasma membrane sphingomyelin to ceramide attenuates the intoxication of host cells by cholera toxin. Cell Microbiol. 2008;10:67–80.
Bock J, Liebisch G, Schweimer J, Schmitz G, Rogler G. Exogenous sphingomyelinase causes impaired intestinal epithelial barrier function. World J Gastroenterol. 2007;13:5217–5225.
Possemiers S, Van Camp J, Bolca S, Verstraete W. Characterization of the bactericidal effect of dietary sphingosine and its activity under intestinal conditions. Int J Food Microbiol. 2005;105:59–70. doi:10.1016/j.ijfoodmicro.2005.05.007.
Acknowledgments
The work was supported by the grants from the Swedish Cancer Society, the Albert Påhlsson Foundation, the Swedish Research Council, and the Research Foundation of Lund University Hospital. Dr. Yajun Cheng is thanked for technical assistance and Dr. Åke Nilsson for stimulating discussions. The current address for Jun Wu is Beijing Institute of Biotechnology, Beijing, 100071, China.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Andersson, D., Kotarsky, K., Wu, J. et al. Expression of Alkaline Sphingomyelinase in Yeast Cells and Anti-inflammatory Effects of the Expressed Enzyme in a Rat Colitis Model. Dig Dis Sci 54, 1440–1448 (2009). https://doi.org/10.1007/s10620-008-0509-2
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10620-008-0509-2