Determination of phospholipase activity of PAF acetylhydrolase

Stafforini, Diana M.; McIntyre, Thomas M.

doi:10.1016/j.freeradbiomed.2012.05.031

Cited by 10 publications

(8 citation statements)

References 57 publications

(74 reference statements)

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“…We assessed the lipoprotein associated phospholipase A2 (Lp-PLA 2 ), a proatherogenic enzyme, as a second marker for HDL-associated anti-inflammatory activity [ 34 ]. HDL associated Lp-PLA 2 activity was significantly lower in exudative AMD patients when compared to control patients.…”

Section: Discussionmentioning

confidence: 99%

High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration

Pertl¹,

Kern²,

Weger³

et al. 2016

PLoS ONE

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PurposeHigh-density lipoproteins (HDL) have long been implicated in the pathogenesis of age-related macular degeneration (AMD). However, conflicting results have been reported with regard to the associations of AMD with HDL-cholesterol levels. The present study is the first to assess HDL composition and metrics of HDL function in patients with exudative AMD and control patients.MethodsBlood samples were collected from 29 patients with exudative AMD and 26 age-matched control patients. Major HDL associated apolipoproteins were determined in apoB-depleted serum by immunoturbidimetry or ELISA, HDL-associated lipids were quantified enzymatically. To get an integrated measure of HDL quantity and quality, we assessed several metrics of HDL function, including cholesterol efflux capacity, anti-oxidative and anti-inflammatory activities using apoB-depleted serum from study participants.ResultsIn our study, we observed that the HDL associated acute phase protein serum amyloid A (SAA) was significantly increased in AMD patients (p<0.01), whereas all other assessed apolipoproteins including ApoA-I, apoA-II, apoC-II, apoC-III and apoE as well as major HDL associated lipids were not altered. HDL efflux capacity, anti-oxidative capacity and arylesterase activity were not different in AMD patients when compared with the control group. The ability of apoB-depleted serum to inhibit monocyte NF-κB expression was significantly improved in AMD patients (mean difference (MD) -5.6, p<0.01). Moreover, lipoprotein-associated phospholipase A2 activity, a marker of vascular inflammation, was decreased in AMD subjects (MD -24.1, p<0.01).ConclusionsThe investigated metrics of HDL composition and HDL function were not associated with exudative AMD in this study, despite an increased content of HDL associated SAA in AMD patients. Unexpectedly, anti-inflammatory activity of apoB-depleted serum was even increased in our study. Our data suggest that the investigated parameters of serum HDL function showed no significant association with exudative AMD. However, we cannot exclude that alterations in locally produced HDL may be part of the AMD pathogenesis.

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Section: Discussionmentioning

confidence: 99%

High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration

Pertl¹,

Kern²,

Weger³

et al. 2016

PLoS ONE

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“…SsE-catalyzed hydrolysis of PAF was monitored by thin-layer chromatography (TLC), a colorimetric assay using 2-thio-PAF (8), and the standard assay using [acetyl-3 H]PAF (36). In the TLC assay, PAF (0.29 mM) was mixed with 7 M SsE M28 in 20 l of 20 mM Tris-HCl (pH 8.0) or the buffer (negative control reaction), and the reactions were stopped by adding 20 l of acetonitrile containing 1% formic acid after 20 min incubation at room temperature.…”

Section: Methodsmentioning

confidence: 99%

“…The standard assay was performed as described previously (36). Briefly, 0.1 mM PAF solution was prepared by mixing 42 l of unlabeled PAF and 50 l of [acetyl-…”

Section: Methodsmentioning

confidence: 99%

Characterization of Streptococcal Platelet-Activating Factor Acetylhydrolase Variants That Are Involved in Innate Immune Evasion

Liu

Xie

et al. 2013

Infect Immun

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Human pathogen group A streptococcus (GAS) has developed mechanisms to subvert innate immunity. We recently reported that the secreted esterase produced by serotype M1 GAS (SsE M1 ) reduces neutrophil recruitment by targeting platelet-activating factor (PAF M28 in a covS deletion mutant of GAS increased neutrophil recruitment and reduced skin infection, whereas in trans expression of SsE M28 in GAS reduced neutrophil infiltration and increased skin invasion in subcutaneous infection of mice. These results suggest that the SsE proteins evolved to target PAF for enhancing innate immune evasion and skin invasion.

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“…It is believed to function by cleaving the oxidatively fragmented portion of a damaged phospholipid, thereby starting the reparative process [2–4, 6, 8, 11, 16–18]. Its substrate specificity for shortened chains at the sn -2 position prevents PAFAH-II from hydrolyzing intact phospholipids, making this enzyme an essential component of homeostasis [4, 11].…”

Section: Introductionmentioning

confidence: 99%

Oligomeric state regulated trafficking of human platelet-activating factor acetylhydrolase type-II

Monillas

Caplan

Thévenin

et al. 2015

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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The intracellular enzyme platelet-activating factor acetylhydrolase type-II (PAFAH-II) hydrolyzes platelet-activating factor and oxidatively fragmented phospholipids. PAFAH-II in its resting state is mainly cytoplasmic, and it responds to oxidative stress by becoming increasingly bound to endoplasmic reticulum and Golgi membranes. Numerous studies have indicated that this enzyme is essential for protecting cells from oxidative stress induced apoptosis. However, the regulatory mechanism of the oxidative stress response by PAFAH-II has not been fully resolved. Here, changes to the oligomeric state of human PAFAH-II were investigated as a potential regulatory mechanism toward enzyme trafficking. Native PAGE analysis in vitro and photon counting histogram within live cells showed that PAFAH-II is both monomeric and dimeric. A Gly-2-Ala site-directed mutation of PAFAH-II demonstrated that the N-terminal myristoyl group is required for homodimerization. Additionally, the distribution of oligomeric PAFAH-II is distinct within the cell; homodimers of PAFAH-II were localized to the cytoplasm while monomers were associated to the membranes of the endoplasmic reticulum and Golgi. We propose that the oligomeric state of PAFAH-II drives functional protein trafficking. PAFAH-II localization to the membrane is critical for substrate acquisition and effective oxidative stress protection. It is hypothesized that the balance between monomer and dimer serves as a regulatory mechanism of a PAFAH-II oxidative stress response.

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Determination of phospholipase activity of PAF acetylhydrolase

Cited by 10 publications

References 57 publications

High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration

High-Density Lipoprotein Function in Exudative Age-Related Macular Degeneration

Characterization of Streptococcal Platelet-Activating Factor Acetylhydrolase Variants That Are Involved in Innate Immune Evasion

Oligomeric state regulated trafficking of human platelet-activating factor acetylhydrolase type-II

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