EMAIL THIS PAGE TO A FRIEND

Journal of neuroinflammation

Cytosolic phospholipase A2 plays a crucial role in ROS/NO signaling during microglial activation through the lipoxygenase pathway.


PMID 26520095

Abstract

Oxidative stress and inflammation are important factors contributing to the pathophysiology of numerous neurological disorders, including Alzheimer's disease, Parkinson's disease, acute stroke, and infections of the brain. There is well-established evidence that proinflammatory cytokines and glutamate, as well as reactive oxygen species (ROS) and nitric oxide (NO), are produced upon microglia activation, and these are important factors contributing to inflammatory responses and cytotoxic damage to surrounding neurons and neighboring cells. Microglial cells express relatively high levels of cytosolic phospholipase A2 (cPLA2), an enzyme known to regulate membrane phospholipid homeostasis and release of arachidonic acid (AA) for synthesis of eicosanoids. The goal for this study is to elucidate the role of cPLA2IV in mediating the oxidative and inflammatory responses in microglial cells. Experiments involved primary microglia cells isolated from transgenic mice deficient in cPLA2α or iPLA2β, as well as murine immortalized BV-2 microglial cells. Inhibitors of cPLA2/iPLA2/cyclooxygenase (COX)/lipoxygenase (LOX) were used in BV-2 microglial cell line. siRNA transfection was employed to knockdown cPLA2 expression in BV-2 cells. Griess reaction protocol was used to determine NO concentration, and CM-H2DCF-DA was used to detect ROS production in primary microglia and BV-2 cells. WST-1 assay was used to assess cell viability. Western blotting was used to assess protein expression levels. Immunocytochemical staining for phalloidin against F-actin was used to demonstrate cell morphology. In both primary and BV-2 microglial cells, stimulation with lipopolysaccharide (LPS) or interferon gamma (IFNγ) resulted in a time-dependent increase in phosphorylation of cPLA2 together with ERK1/2. In BV-2 cells, LPS- and IFNγ-induced ROS and NO production was inhibited by arachidonyl trifluoromethyl ketone (AACOCF3) and pyrrophenone as well as RNA interference, but not BEL, suggesting a link between cPLA2, and not iPLA2, on LPS/IFNγ-induced nitrosative and oxidative stress in microglial cells. Primary microglial cells isolated from cPLA2α-deficient mice generated significantly less NO and ROS as compared with the wild-type mice. Microglia isolated from iPLA2β-deficient mice did not show a decrease in LPS-induced NO and ROS production. LPS/IFNγ induced morphological changes in primary microglia, and these changes were mitigated by AACOCF3. Interestingly, despite that LPS and IFNγ induced an increase in phospho-cPLA2 and prostaglandin E2 (PGE2) release, LPS- and IFNγ-induced NO and ROS production were not altered by the COX-1/2 inhibitor but were suppressed by the LOX-12 and LOX-15 inhibitors instead. In summary, the results in this study demonstrated the role of cPLA2 in microglial activation with metabolic links to oxidative and inflammatory responses, and this was in part regulated by the AA metabolic pathways, namely the LOXs. Further studies with targeted inhibition of cPLA2/LOX in microglia during neuroinflammatory conditions can be valuable to investigate the therapeutic potential in ameliorating neurological disease pathology.

Related Materials

Product #

Image

Description

Molecular Formula

Add to Cart

AV38791
Anti-HTATIP antibody produced in rabbit, affinity isolated antibody
SAB2103488
Anti-HTATIP antibody produced in rabbit, affinity isolated antibody
SAB2104133
Anti-HTATIP, (N-terminal) antibody produced in rabbit, affinity isolated antibody
SAB1407003
Anti-KAT5 antibody produced in mouse, purified immunoglobulin, buffered aqueous solution
HPA016953
Anti-KAT5 antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution
SAB2106496
Anti-KAT5 antibody produced in rabbit, affinity isolated antibody
SAB4504108
Anti-phospho-TIP60 (pSer86) antibody produced in rabbit, affinity isolated antibody
SAB4504732
Anti-phospho-Tip60 (pSer90) antibody produced in rabbit, affinity isolated antibody
SAB2501599
Anti-PLA2G4A antibody produced in goat, affinity isolated antibody, buffered aqueous solution
SAB4500117
Anti-TIP60 antibody produced in rabbit, affinity isolated antibody
SAB4500118
Anti-TIP60 antibody produced in rabbit, affinity isolated antibody
SAB4500119
Anti-TIP60 antibody produced in rabbit, affinity isolated antibody
A231
Arachidonyl trifluoromethyl ketone, oil, ≥97% (NMR)
C21H31F3O
B1552
Bromoenol lactone, ≥98% (TLC)
C16H13BrO2
EHU009531 MISSION® esiRNA, esiRNA targeting human PLA2G4A (esiRNA1)
EHU139921 MISSION® esiRNA, esiRNA human KAT5 (esiRNA1)
EMU060371 MISSION® esiRNA, esiRNA targeting mouse Pla2g4a (esiRNA1)
SAB5300083
Monoclonal Anti-KAT5 antibody produced in mouse, clone 3F9, ascites fluid
WH0005321M1
Monoclonal Anti-PLA2G4A antibody produced in mouse, clone 10G5, purified immunoglobulin, buffered aqueous solution
N9125
N-(1-Naphthyl)ethylenediamine dihydrochloride, ≥98%
C12H14N2 · 2HCl
222488
N-(1-Naphthyl)ethylenediamine dihydrochloride, ACS reagent, >98%
C12H14N2 · 2HCl
SML0499
NCTT-956, ≥97% (NMR)
C17H15N3O4S
74540
Nordihydroguaiaretic acid, ≥97.0% (HPLC)
C18H22O4
N5023
Nordihydroguaiaretic acid, ≥90% (HPLC), from Larrea divaricata (creosote bush)
C18H22O4
S9251
Sulfanilamide, ≥99%
C6H8N2O2S
33626
Sulfanilamide, puriss. p.a., ≥99% (calc. to the dried substance)
C6H8N2O2S