Glucosylceramide synthesis and synthase expression protect against ceramide-induced stress.
Ceramides (Cers), critical for epidermal barrier function, also can inhibit keratinocyte proliferation, while glucosylceramides (GlcCers) exert pro-mitogenic effects. Since alterations in Cer-to-GlcCer ratios appear to modulate cellular growth versus apoptosis, we assessed whether keratinocytes up-regulate GlcCer synthesis as a protective mechanism against Cer-induced stress. Exogenous sphingomyelinase (SMase) treatment of cultured human keratinocytes (CHK) initially decreased proliferation and cellular sphingomyelin (50-60% decrease; P < 0.001), and increased Cer levels (6.1- to 6.8-fold; P < 0.001). Proliferation recovered to normal rates by 24 h, in parallel with increased cellular GlcCer. Both GlcCer synthesis and GlcCer synthase activity increased significantly by 8 h following SMase (8.2- and 2.4-fold, respectively; P < 0.01 each vs. control), attributed to antecedent increases in GlcCer synthase mRNA and protein expression. Further evidence that GlcCer production is responsible for normalized CHK proliferation includes: a) attenuation of SMase-induced inhibition of proliferation by exogenous GlcCer; and b) enhancement of the SMase effect in cells cotreated with the GlcCer synthase inhibitor, PDMP (D-threo-1-phenyl-2(decanoylamino)-3-morpholino-1-propanol). Finally, although proliferation in immortalized, nontransformed keratinocytes (HaCaT) also was inhibited by SMase, HaCaT cells that overexpress GlcCer synthase were resistant to this effect. Thus, SMase-induced stress initiates a response in keratinocytes that includes upregulation of GlcCer synthesis which may protect against the deleterious effects of excess Cer.
Uchida
Yoshikazu
Y
Dermatology Service, Department of Veterans Affairs Medical Center, San Francisco, CA, USA.
Murata
Satoru
S
Schmuth
Matthias
M
Behne
Martin J
MJ
Lee
Jeong Deuk
JD
Ichikawa
Shinichi
S
Elias
Peter M
PM
Hirabayashi
Yoshio
Y
Holleran
Walter M
WM
D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002478:Cells, Cultured; D002518:Ceramides / Q000096:biosynthesis* / Q000494:pharmacology*; D017931:DNA Primers; D004261:DNA Replication; D005964:Glucosyltransferases / Q000096:biosynthesis*; D006801:Humans; D018384:Oxidative Stress; D020133:Reverse Transcriptase Polymerase Chain Reaction
D002518:Ceramides; D017931:DNA Primers; D005964:Glucosyltransferases; C023246:ceramide glucosyltransferase