Toll-like receptor expression in human keratinocytes: nuclear factor kappaB controlled gene activation by Staphylococcus aureus is toll-like receptor 2 but not toll-like receptor 4 or platelet activating factor receptor dependent.
Cultured primary human keratinocytes were screened for their expression of various members of the toll-like receptor (TLR) family. Keratinocytes were found to constitutively express TLR1, TLR2, TLR3, TLR5, and TLR9 but not TLR4, TLR6, TLR7, TLR8, or TLR10 as shown by polymerase chain reaction analysis. The expression of the crucial receptor for signaling of staphylococcal compounds TLR2 was also confirmed by immunohistochemistry, in contrast to TLR4, which showed a negative staining pattern. Next, we analyzed the activation of the proinflammatory nuclear transcription factor kappaB by Staphylococcus aureus strain 8325-4. Using nuclear extract gel shifts, RelA staining, and luciferase reporter transfection plasmids we found a clear induction of nuclear factor kappaB translocation by the bacteria. This translocation induced the transcription of nuclear factor kappaB controlled genes such as inducible nitric oxide synthetase, COX2, and interleukin-8. Transcription of these genes was followed by production of increased amounts of interleukin-8 protein and NO. Inhibition experiments using monoclonal antibodies and the specific platelet activating factor receptor inhibitor CV3988 showed that nuclear factor kappaB activation by S. aureus was TLR2 but not TLR4 or platelet activating factor receptor dependent. In line, the purified staphylococcal cell wall components lipoteichoic acid and peptidoglycan, known to signal through TLR2, also showed nuclear factor kappaB translocation in human keratinocytes, indicating a crucial role of the staphylococcal cell wall in the innate immune stimulation of human keratinocytes. These results help to explain the complex activation of human keratinocytes by S. aureus and its cell wall components in various inflammatory disorders of the skin.
Mempel
Martin
M
Department of Dermatology and Allergy, Biederstein , Munich, Germany. m.mempel@lrz.tum.de
Voelcker
Verena
V
Köllisch
Gabriele
G
Plank
Christian
C
Rad
Roland
R
Gerhard
Markus
M
Schnopp
Christina
C
Fraunberger
Peter
P
Walli
Autar K
AK
Ring
Johannes
J
Abeck
Dietrich
D
Ollert
Markus
M
D002478:Cells, Cultured; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D016209:Interleukin-8 / Q000235:genetics / Q000378:metabolism; D015603:Keratinocytes / Q000166:cytology / Q000382:microbiology / Q000502:physiology*; D008562:Membrane Glycoproteins / Q000235:genetics*; D016328:NF-kappa B / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D010742:Phospholipid Ethers / Q000494:pharmacology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface / Q000235:genetics*; D043562:Receptors, G-Protein-Coupled / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013203:Staphylococcal Infections / Q000503:physiopathology*; D013211:Staphylococcus aureus / Q000472:pathogenicity; D051194:Toll-Like Receptor 1; D051201:Toll-Like Receptor 10; D051195:Toll-Like Receptor 2; D051196:Toll-Like Receptor 3; D051197:Toll-Like Receptor 4; D051198:Toll-Like Receptor 5; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; D015533:Transcriptional Activation; D014774:Virulence
D016209:Interleukin-8; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D010742:Phospholipid Ethers; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C495342:TLR2 protein, human; C495344:TLR3 protein, human; C495345:TLR4 protein, human; C495346:TLR5 protein, human; C495348:TLR7 protein, human; C495349:TLR8 protein, human; C495371:TLR9 protein, human; D051194:Toll-Like Receptor 1; D051201:Toll-Like Receptor 10; D051195:Toll-Like Receptor 2; D051196:Toll-Like Receptor 3; D051197:Toll-Like Receptor 4; D051198:Toll-Like Receptor 5; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; C039351:platelet activating factor receptor; C037913:CV 3988; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II