Lipopolysaccharide-induced sensitization of adenylyl cyclase activity in murine macrophages.
LPS is known to modulate macrophage responses during sepsis, including cytokine release, phagocytosis, and proliferation. Although agents that elevate cAMP reverse LPS-induced macrophage functions, whether LPS itself modulates cAMP and whether LPS-induced decreases in proliferation are modulated via a cAMP-dependent pathway are not known. Murine macrophages (RAW264.7 cells) were treated with LPS in the presence or absence of inhibitors of prostaglandin signaling, protein kinases, CaM, Gi proteins, and NF-kappaB translocation or transcription/translation. LPS effects on CaMKII phosphorylation and the expression of relevant adenylyl cyclase (AC) isoforms were measured. LPS caused a significant dose (5-10,000 ng/ml)- and time (1-8 h)-dependent increase in forskolin-stimulated AC activity that was abrogated by pretreatment with SN50 (an NF-kappaB inhibitor), actinomycin D, or cycloheximide, indicating that the effect is mediated via NF-kappaB-dependent transcription and new protein synthesis. Furthermore, LPS decreased the phosphorylation state of CaMKII, and pretreatment with a CaM antagonist attenuated the LPS-induced sensitization of AC. LPS, cAMP, or PKA activation each independently decreased macrophage proliferation. However, inhibition of NF-kappaB had no effect on LPS-induced decreased proliferation, indicating that LPS-induced decreased macrophage proliferation can proceed via PKA-independent signaling pathways. Taken together, these findings indicate that LPS induces sensitization of AC activity by augmenting the stimulatory effect of CaM and attenuating the inhibitory effect of CaMKII on isoforms of AC that are CaMK sensitive.
Osawa
Y
Y
Dept. of Anesthesiology, College of Physicians and Surgeons, Columbia Univ., 630 W. 168th St., P&S Box 46, New York, NY 10032, USA.
Lee
H T
HT
Hirshman
C A
CA
Xu
D
D
Emala
C W
CW
D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D005576:Colforsin / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D011453:Prostaglandins / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis
D002147:Calmodulin; D018841:HSP90 Heat-Shock Proteins; D007527:Isoenzymes; D008070:Lipopolysaccharides; D016328:NF-kappa B; D011453:Prostaglandins; D012333:RNA, Messenger; D005576:Colforsin; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D000262:Adenylyl Cyclases