Induction of glioma apoptosis by microglia-secreted molecules: The role of nitric oxide and cathepsin B.
Microglia contributes significantly to brain tumor mass, particularly in astrocytic gliomas. Here, we examine the cytotoxic effects of soluble components secreted from microglia culture on glioma cells. Microglia conditioned culture medium (MCM) actively stimulated apoptotic death of glioma cells, and the effects of MCM prepared from LPS- or IFN-gamma-activated microglia were more pronounced. The cytotoxic effects were glioma-specific in that primary cultured rat astrocytes were not affected by MCM. A donor of peroxynitrite induced glioma-specific cell death. In addition, NO synthase inhibitor suppressed glioma cell death induced by activated MCM, indicating that NO is one of the key molecules responsible for glioma cytotoxicity mediated by activated MCM. However, since unstimulated resting microglia produces low or very limited level of NO, MCM may contain other critical molecule(s) that induce glioma apoptosis. To identify the proteins secreted in MCM, proteomic analysis was performed on control or activated medium. Among over 200 protein spots detected by Coomassie blue staining, we identified 26 constitutive and 28 LPS- or IFN-gamma-regulated MCM proteins. Several cathepsin proteases were markedly expressed, which were reduced upon activation. In particular, suppression of cathepsin B by the chemical inhibitors significantly reversed MCM-induced glioma cell death, implying a critical role of this protease in cytotoxicity. Our findings provide evidence on the functional implications of specific microglial-secreted proteins in glioma cytotoxicity, as well as a basis to develop a proteomic databank of both basal and activation-related proteins in microglia.
Hwang
So-Young
SY
Department of Physiology and Biophysics, and Center for Advanced Medical Education by BK21 Project, Inha University, College of Medicine, Nam-Ku, Incheon 402-751, Korea.
Yoo
Byong-Chul
BC
Jung
Jae-won
JW
Oh
Eok-Soo
ES
Hwang
Ji-Sun
JS
Shin
Jin-A
JA
Kim
Song-Yi
SY
Cha
Seok-Ho
SH
Han
Inn-Oc
IO
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D017209:Apoptosis; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005910:Glioma / Q000378:metabolism*; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D017628:Microglia / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D000998:Antiviral Agents; D017077:Culture Media, Conditioned; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D009569:Nitric Oxide; D007371:Interferon-gamma; D002401:Cathepsin B; C533915:Ctsb protein, mouse; C533916:Ctsb protein, rat