Translational event mediates differential production of tumor necrosis factor-alpha in hyaluronan-stimulated microglia and macrophages.
Recent evidence has demonstrated that hyaluronan synthase 2 mRNA is up-regulated after brain ischemia. After a cerebral ischemic event, microglia and macrophages are the major inflammatory cells and are activated by hyaluronan (HA). However, it is unclear how these cells compare with regard to HA responsiveness. We show here that peritoneal macrophages and RAW 264.7 macrophages produced more than five- and 10-fold more tumor necrosis factor-alpha (TNF-alpha) than primary microglia and BV-2 microglia, respectively. Antibody blockade study showed that CD44, Toll-like receptor-4 receptor and the receptor for HA-mediated motility were responsible for HA-induced TNF-alpha release. Furthermore, HA induced higher levels of phosphorylated MAPK in RAW 264.7 cells when compared with BV-2 cells. HA-mediated TNF-alpha production required p38 MAPK, extracellular-regulated kinase and c-Jun N-terminal kinase phosphorylation in both cell types. The levels of HA-induced TNF-alpha mRNA expression in BV-2 cells were only twofold lower compared with RAW 264.7 cells, suggesting that a translational event is involved in the differential production of TNF-alpha. Western blot analysis revealed that HA treatment resulted in more rapid phosphorylation of eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) and more effective dissociation of 4E-BP1 from eukaryotic initiation factor 4E in RAW 264.7 cells than in BV-2 cells. Additionally, HA-induced phosphorylation of 4E-BP1 was dependent on MAPK signaling, indicating that RAW 264.7 cells exhibited higher levels of hyperphosphorylated 4E-BP1 possibly due to the overactivation of MAPK. The results suggest that resident microglia and blood-derived monocytes/macrophages exhibit differential sensitivities in response to extracellular mediators after brain ischemia.
Neuro-Medical Scientific Center, Buddhist Tzu-Chi General Hospital, Tzu-Chi College of Technology, Hualien, Taiwan.
Mei-Jen
MJ
Wang
Jon-Son
JS
Kuo
Wen-Wen
WW
Lee
Hsin-Yi
HY
Huang
Wu-Fu
WF
Chen
Shinn-Zong
SZ
Lin
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000276:Adjuvants, Immunologic / Q000494:pharmacology*; D000818:Animals; D000831:Animals, Newborn; D000906:Antibodies / Q000494:pharmacology; D015153:Blotting, Western / Q000379:methods; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016326:Extracellular Matrix Proteins / Q000276:immunology; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D018960:Hyaluronan Receptors / Q000276:immunology; D006820:Hyaluronic Acid / Q000494:pharmacology*; D047468:Immunoprecipitation / Q000379:methods; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects*; D017628:Microglia / Q000187:drug effects*; D010750:Phosphoproteins / Q000378:metabolism; D014176:Protein Biosynthesis / Q000187:drug effects*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013997:Time Factors; D051197:Toll-Like Receptor 4 / Q000276:immunology; D014157:Transcription Factors / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D000276:Adjuvants, Immunologic; D000906:Antibodies; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C087000:EIF4EBP1 protein, human; C111931:ELF4 protein, human; D004791:Enzyme Inhibitors; D016326:Extracellular Matrix Proteins; D018960:Hyaluronan Receptors; D008070:Lipopolysaccharides; D010750:Phosphoproteins; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D014157:Transcription Factors; D014409:Tumor Necrosis Factor-alpha; C098946:hyaluronan-mediated motility receptor; D006820:Hyaluronic Acid