* Department of Pharmacology, College of Medicine, National Taiwan University; and
Department of Microbiology and Immunology, National Yang-Ming University School of Medicine, Taipei, Taiwan
Correspondence: W. W. Lin, Ph.D., Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan. E-mail: wwl{at}ha.mc.ntu.edu.tw
We have explored the regulatory roles played by
Ca2+-dependent signaling on lipopolysaccharide
(LPS)-induced nitric oxide (NO), prostaglandin E2
(PGE2), tumor necrosis factor 
(TNF-), and
interleukin-6 (IL-6) release in mouse peritoneal macrophages. To
elevate intracellular Ca2+, we used thapsigargin (TG) and
UTP. Although LPS alone cannot stimulate NO synthesis, co-addition with
TG, which sustainably increased [Ca2+]i,
resulted in NO release. UTP, via acting on P2Y6 receptors,
can stimulate phosphoinositide (PI) turnover and transient
[Ca2+]i increase, however, it did not possess
the NO priming effect. LPS alone triggered the release of
PGE2, TNF-, and IL-6; all of which were potentiated by
the presence of TG, but not of UTP. The stimulatory effect of LPS plus
TG on NO release was inhibited by the presence of Ro 31-8220, Go6976,
KN-93, PD 098059, or SB 203580, and abolished by BAPTA/AM and nuclear
factor 
B (NF-B) inhibitor, PDTC. PGE2, TNF-, and
IL-6 release by LPS alone were attenuated by Ro 31-8220, Go6976, PD
098059, SB 203580, and PDTC. Using L-NAME, soluble TNF-
receptor, IL-6 antibody, NS-398, and indomethacin, we performed
experiments to understand the cross-regulation by the four mediators.
The results revealed that TNF- up-regulated NO, PGE2,
and IL-6 synthesis; PGE2 up-regulated NO, but
down-regulated TNF- synthesis; and PGE2 and IL-6
mutually up-regulated reciprocally. Taken together, murine peritoneal
macrophages required a sustained [Ca2+]i
increase, which proceeds after TG, but not UTP, stimulation, to enhance
LPS-mediated release of inflammatory mediators, particularly for NO
induction. Activation of PKC-, ERK-, and p38 MAPK-dependent signaling
also are essential for LPS action. The positive regulatory interactions
among these mediators might amplify the inflammatory response caused by
endotoxin.
Key Words: LPS • UTP • Ca2+ • protein kinase
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