scholarly journals Putative roles of Ca2+ -independent phospholipase A2 in respiratory chain-associated ROS production in brain mitochondria: influence of docosahexaenoic acid and bromoenol lactone

2014 ◽  
Vol 131 (2) ◽  
pp. 163-176 ◽  
Author(s):  
Caroline Nordmann ◽  
Mikhail Strokin ◽  
Peter Schönfeld ◽  
Georg Reiser
Keyword(s):  
Docosahexaenoic Acid ◽  
Phospholipase A2 ◽  
Respiratory Chain ◽  
Brain Mitochondria ◽  
Ros Production ◽  
Bromoenol Lactone

scholarly journals Effect of salicylate on oxidative phosphorylation and respiration of mitochondrial fragments

1965 ◽  
Vol 97 (1) ◽  
pp. 194-198 ◽  
Author(s):  
JT Miyahara ◽  
R Karler
Keyword(s):  
Oxidative Phosphorylation ◽  
Respiratory Chain ◽  
Direct Effect ◽  
Brain Mitochondria ◽  
Final Effect

1. The effects of salicylate on oxidative phosphorylation and respiration were investigated in liver and brain mitochondria, and in sonically prepared mitochondrial fragments. 2. Salicylate was shown to uncouple oxidative phosphorylation in mitochondrial fragments, as well as in intact mitochondria. The effects of salicylate on mitochondria and oxidative phosphorylation resemble those produced by dinitrophenol. 3. The quantitative effects of salicylate on respiration in vitro were shown to be complex. The final effect may be the resultant of an interaction of a multiplicity of factors, such as the tissue, the substrate, mitochondrial and extramitochondrial influences, and the direct effect on the respiratory chain itself.

NAD(P)H oxidase-mediated reactive oxygen species production alters astrocyte membrane molecular order via phospholipase A2

2009 ◽  
Vol 421 (2) ◽  
pp. 201-210 ◽  
Author(s):  
Donghui Zhu ◽  
Chunhua Hu ◽  
Wenwen Sheng ◽  
Kevin S. Tan ◽  
Mark A. Haidekker ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Phospholipase A2 ◽  
Actin Polymerization ◽  
Plasma Membranes ◽  
Reactive Oxygen ◽  
Membrane Dynamics ◽  
Mitogen Activated Protein Kinase ◽  
Ros Production ◽  
Oxygen Species ◽  
Mitogen Activated Protein

ROS (reactive oxygen species) overproduction is an important underlying factor for the activation of astrocytes in various neuropathological conditions. In the present study, we examined ROS production in astrocytes and downstream effects leading to changes in the signalling cascade, morphology and membrane dynamics using menadione, a redox-active compound capable of inducing intracellular ROS. NAD(P)H oxidase-mediated menadione-induced ROS production, which then stimulated phosphorylation of p38 MAPK (mitogen-activated protein kinase) and ERK1/2 (extracellular-signal-regulated kinase 1/2), and increased actin polymerization and cytoskeletal protrusions. We also showed that astrocyte plasma membranes became more molecularly ordered under oxidative stress, which was abrogated by down-regulating cPLA2 (cytosolic phospholipase A2) either with a pharmacological inhibitor or by RNA interference. In addition, mild disruption of F-actin with cytochalasin D suppressed menadione-enhanced phosphorylation of cPLA2 and membrane alterations. Taken together, these results suggest an important role for ROS derived from NAD(P)H oxidase in activation of astrocytes to elicit biochemical, morphological and biophysical changes reminiscent of reactive astrocytes in pathological conditions.

scholarly journals Massive calcium–activated endocytosis without involvement of classical endocytic proteins

2010 ◽  
Vol 137 (1) ◽  
pp. 111-132 ◽  
Author(s):  
Vincenzo Lariccia ◽  
Michael Fine ◽  
Simona Magi ◽  
Mei-Jung Lin ◽  
Alp Yaradanakul ◽  
...  
Keyword(s):  
Actin Cytoskeleton ◽  
Phospholipase A2 ◽  
Phospholipase C ◽  
Cardiac Myocytes ◽  
Adenosine Triphosphate ◽  
Hek293 Cells ◽  
Membrane Domains ◽  
Baby Hamster Kidney ◽  
Bromoenol Lactone ◽  
Cytoplasmic Side

We describe rapid massive endocytosis (MEND) of >50% of the plasmalemma in baby hamster kidney (BHK) and HEK293 cells in response to large Ca transients. Constitutively expressed Na/Ca exchangers (NCX1) are used to generate Ca transients, whereas capacitance recording and a membrane tracer dye, FM 4–64, are used to monitor endocytosis. With high cytoplasmic adenosine triphosphate (ATP; >5 mM), Ca influx causes exocytosis followed by MEND. Without ATP, Ca transients cause only exocytosis. MEND can then be initiated by pipette perfusion of ATP, and multiple results indicate that ATP acts via phosphatidylinositol-bis 4,5-phosphate (PIP2) synthesis: PIP2 substitutes for ATP to induce MEND. ATP-activated MEND is blocked by an inositol 5-phosphatase and by guanosine 5′-[γ-thio]triphosphate (GTPγS). Block by GTPγS is overcome by the phospholipase C inhibitor, U73122, and PIP2 induces MEND in the presence of GTPγS. MEND can occur in the absence of ATP and PIP2 when cytoplasmic free Ca is clamped to 10 µM or more by Ca-buffered solutions. ATP-independent MEND occurs within seconds during Ca transients when cytoplasmic solutions contain polyamines (e.g., spermidine) or the membrane is enriched in cholesterol. Although PIP2 and cholesterol can induce MEND minutes after Ca transients have subsided, polyamines must be present during Ca transients. MEND can reverse over minutes in an ATP-dependent fashion. It is blocked by brief β-methylcyclodextrin treatments, and tests for involvement of clathrin, dynamins, calcineurin, and actin cytoskeleton were negative. Therefore, we turned to the roles of lipids. Bacterial sphingomyelinases (SMases) cause similar MEND responses within seconds, suggesting that ceramide may be important. However, Ca-activated MEND is not blocked by reagents that inhibit SMases. MEND is abolished by the alkylating phospholipase A2 inhibitor, bromoenol lactone, whereas exocytosis remains robust, and Ca influx causes MEND in cardiac myocytes without preceding exocytosis. Thus, exocytosis is not prerequisite for MEND. From these results and two companion studies, we suggest that Ca promotes the formation of membrane domains that spontaneously vesiculate to the cytoplasmic side.

scholarly journals Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-derived Factor Receptor (PEDF-R)

2021 ◽  
Author(s):  
Jeanee Bullock ◽  
Federica Polato ◽  
Mones Abu-Asab ◽  
Alexandra Bernardo-Colón ◽  
Elma Aflaki ◽  
...  
Keyword(s):  
Retinal Pigment Epithelium ◽  
Phospholipase A2 ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Photoreceptor Outer Segment ◽  
Western Blots ◽  
Rpe Cells ◽  
Photoreceptor Outer Segments ◽  
Transmission Electron ◽  
Bromoenol Lactone

AbstractPurposeTo examine the contribution of PEDF-R to the phagocytosis process. Previously, we identified PEDF-R, the protein encoded by the PNPLA2 gene, as a phospholipase A2 in the retinal pigment epithelium (RPE). During phagocytosis, RPE cells ingest abundant phospholipids and protein in the form of photoreceptor outer segment (POS) tips, which are then hydrolyzed. The role of PEDF-R in RPE phagocytosis is not known.MethodsMice in which PNPLA2 was conditionally knocked out in the RPE were generated (cKO). Mouse RPE/choroid explants were cultured. Human ARPE-19 cells were transfected with siPNPLA2 silencing duplexes. POS were isolated from bovine retinas. The phospholipase A2 inhibitor bromoenol lactone was used. Transmission electron microscopy, immunofluorescence, lipid labeling, pulse-chase experiments, western blots, and free fatty acid and β-hydroxybutyrate assays were performed.ResultsThe RPE of the cKO mice accumulated lipids as well as more abundant and larger rhodopsin particles compared to littermate controls. Upon POS exposure, RPE explants from cKO mice released less β-hydroxybutyrate compared to controls. After POS ingestion during phagocytosis, rhodopsin degradation was stalled both in cells treated with bromoenol lactone and in PNPLA2-knocked-down cells relative to their corresponding controls. Phospholipase A2 inhibition lowered β-hydroxybutyrate release from phagocytic RPE cells. PNPLA2 knock down also resulted in a decline in fatty acids and β-hydroxybutyrate release from phagocytic RPE cells.ConclusionsPEDF-R downregulation delayed POS digestion during phagocytosis. The findings imply that efficiency of RPE phagocytosis depends on PEDF-R, thus identifying a novel contribution of this protein to POS degradation in the RPE.

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