Excretory-secretory products produced by paragonimus westermani differentially regulate the nitric oxide production and viability of microglial cells.
BACKGROUND
Tissue-invading helminth parasites secrete a large amount of cysteine proteases that may play critical roles in tissue invasion and immune modulation. However, roles of excretory-secretory products (ESP) secreted by Paragonimus westermani in the activation and death of microglial cells in brain are poorly understood.


OBJECTIVE
In the present study, we investigated whether ESP could regulate microglial nitric oxide (NO) production and viability.


METHODS
The NO production and cell viability were assessed by respectively measuring the formation of nitrite and the release of lactate dehyrogenase.


RESULTS
At a low (0.2 microg/ml) concentration, ESP significantly stimulated NO production with no apparent cell injury or death in cultured microglial cells. However, at high (> or =2 microg/ml) concentrations, ESP induced severe cell death. Inhibition of inducible NO synthase significantly reduced the NO productivity, but not cytotoxicity, of ESP. Similarly, inhibitors of the extracellular signal-regulated kinase, p38 and nuclear factor kappa B also blocked only the NO productivity of ESP. Interestingly, heat inactivation did not hamper the ability of ESP to stimulate microglial NO production. Similarly, pretreatment with thiol-crosslinking reagents dramatically reduced both proteolytic activity and cytotoxicity of ESP, but did not alter NO production in microglial cells. Interestingly, although cysteine protease competitive inhibitors and thiol-alkylating reagents markedly reduced the proteolytic activity of ESP, they did not influence the NO productivity and cytotoxicity of ESP.


CONCLUSIONS
The present results indicate that the NO production and cytotoxicity by ESP may be differentially regulated via unknown mechanisms, not related with cysteine protease activity.
Jin
Youngnam
Y
Department of Pharmacology, Ewha Medical School and Ewha Institute of Neuroscience, Ewha Woman's University, Seoul, South Korea.
Lee
Jae-Chul
JC
Choi
In Young
IY
Kim
Eun A
EA
Shin
Myeong Heon
MH
Kim
Won-Ki
WK
D000818:Animals; D001685:Biological Factors / Q000378:metabolism* / Q000494:pharmacology*; D002478:Cells, Cultured; D006358:Hot Temperature; D007770:L-Lactate Dehydrogenase / Q000032:analysis / Q000378:metabolism; D017628:Microglia / Q000378:metabolism* / Q000473:pathology*; D009336:Necrosis / Q000473:pathology*; D009566:Nitrates / Q000032:analysis / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D048492:Paragonimus westermani / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D001685:Biological Factors; D009566:Nitrates; D009569:Nitric Oxide; D007770:L-Lactate Dehydrogenase