Synthetic Phytoceramides Induce Apoptosis with Higher Potency than Ceramides

  1. Onyou Hwang1,
  2. Guncheol Kim2,
  3. Yeon Joo Jang1,
  4. Seong Who Kim1,
  5. Guiyong Choi3,
  6. Hyun Jin Choi1,
  7. Seon Young Jeon2,
  8. Don Gyu Lee2 and
  9. Jae Dam Lee1
  1. 1Department of Biochemistry, University of Ulsan College of Medicine, Seoul, Korea (O.H., Y.J.J., S.W.K., H.J.C., J.D.L.);2Department of Chemistry, College of Natural Sciences, Chungnam National University, Taejon, Korea (G.K., S.Y.J., D.G.L.); and 3The SphingoBiolipid Corp, Yongin, Korea (G.C.)

    Abstract

    Ceramides are naturally occurring compounds recognized to mediate apoptosis. N-acylsphingosines, containing a double bond at carbons 4 and 5 of their sphingoid backbone, are thought to be the active form, because N-acylsphinganines with completely saturated sphingoid are inactive. In the present study, we synthesized a series of N-acyl-4d-ribo-phytosphingosines (phytoceramides) that contain a hydroxyl group at carbon 4 and investigated structure-cytotoxicity relationship of the presumed functional groups in ceramides. N-Acetylphytoceramide (PCer2) and N-hexanoylphytoceramide (PCer6) were found to be more cytotoxic than ceramides as determined by released lactate dehydrogenase activity and morphological criteria. This was not caused by intracellular conversion of phytoceramides to ceramides, because noN-hexanoylsphingosine was formed after incubation of cell lysate with PCer6. Among phytoceramides having acyl chains two to eight carbons long, the cytotoxicity was highest with five or six carbons. The carbonyl group of the amide bond did not seem to be critical, because substitution of the oxygen with sulfur did not influence the cytotoxicity. The phytoceramide-induced cell death was observed to be apoptotic in nature with the use of terminal deoxynucleotidyl transferase dUTP nick-end labeling and propidium iodide staining. Because phytoceramides can be readily synthesized from yeast sources, they may present a potential and economical alternative to ceramide in future studies and therapies.

    Footnotes

    • Send reprint requests to: Jae Dam Lee M.D., Ph.D., Department of Biochemistry, University of Ulsan, College of Medicine 388–1 Poongnap-dong, Songpa-ku Seoul 138–736, Korea. E-mail:jdlee{at}www.amc.seoul.kr

    • This work was supported by the 98 Good Health R&D Program (HMP-98-N-1–0011) of the Korean Ministry of Health and Welfare.

    • Abbreviations:
      HPLC
      high-performance liquid chromatography
      Cer2
      N-acetylsphingosine
      DHCer2
      N-acetylsphinganine
      Cer6
      N-hexanoylsphingosine
      DHCer6
      N-hexanoylsphinganine
      TUNEL
      terminal deoxynucleotidyl transferase dUTP nick-end labeling
      DMSO
      dimethyl sulfoxide
      MS
      mass spectrometry
      ESI
      electrospray ionization
      PCer2
      N-acetyl-4d-ribo-phytosphingosine
      PCer6
      N-hexanoyl-4d-ribo-phytosphingosine
      Pcer6S
      N-thio-hexanoylphytosphingosine
      LDH
      lactate dehydrogenase
      ANOVA
      analysis of variance
      • Received August 29, 2000.
      • Accepted January 31, 2000.
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    1. Molecular Pharmacology May 1, 2001 vol. 59 no. 5 1249-1255
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