Lipid-orchestrated paracrine circuit coordinates mast cell maturation and anaphylaxis through functional interaction with fibroblasts.

Interaction of mast cells (MCs) with fibroblasts is essential for MC maturation within tissue microenvironments, although the underlying mechanism is incompletely understood. Through a phenotypic screening of >30 mouse lines deficient in lipid-related genes, we found that deletion of the lysophosphatidic acid (LPA) receptor LPA1, like that of the phospholipase PLA2G3, the prostaglandin D2 (PGD2) synthase L-PGDS, or the PGD2 receptor DP1, impairs MC maturation and thereby anaphylaxis. Mechanistically, MC-secreted PLA2G3 acts on extracellular vesicles (EVs) to supply lysophospholipids, which are converted by fibroblast-derived autotaxin (ATX) to LPA. Fibroblast LPA1 then integrates multiple pathways required for MC maturation by facilitating integrin-mediated MC-fibroblast adhesion, IL-33-ST2 signaling, L-PGDS-driven PGD2 generation, and feedforward ATX-LPA1 amplification. Defective MC maturation resulting from PLA2G3 deficiency is restored by supplementation with LPA1 agonists or PLA2G3-modified EVs. Thus, the lipid-orchestrated paracrine circuit involving PLA2G3-driven lysophospholipid, eicosanoid, integrin, and cytokine signaling fine-tunes MC-fibroblast communication, ensuring MC maturation.

Lyso-PAF, a biologically inactive phospholipid, contributes to RAF1 activation.

Phospholipase A2, group VII (PLA2G7) is widely recognized as a secreted, lipoprotein-associated PLA2 in plasma that converts phospholipid platelet-activating factor (PAF) to a biologically inactive product Lyso-PAF during inflammatory response. We report that intracellular PLA2G7 is selectively important for cell proliferation and tumor growth potential of melanoma cells expressing mutant NRAS, but not cells expressing BRAF V600E. Mechanistically, PLA2G7 signals through its product Lyso-PAF to contribute to RAF1 activation by mutant NRAS, which is bypassed by BRAF V600E. Intracellular Lyso-PAF promotes p21-activated kinase 2 (PAK2) activation by binding to its catalytic domain and altering ATP kinetics, while PAK2 significantly contributes to S338-phosphorylation of RAF1 in addition to PAK1. Furthermore, the PLA2G7-PAK2 axis is also required for full activation of RAF1 in cells stimulated by epidermal growth factor (EGF) or cancer cells expressing mutant KRAS. Thus, PLA2G7 and Lyso-PAF exhibit intracellular signaling functions as key elements of RAS-RAF1 signaling.

The DDHD2-STXBP1 interaction mediates long-term memory via generation of saturated free fatty acids.

The phospholipid and free fatty acid (FFA) composition of neuronal membranes plays a crucial role in learning and memory, but the mechanisms through which neuronal activity affects the brain's lipid landscape remain largely unexplored. The levels of saturated FFAs, particularly of myristic acid (C14:0), strongly increase during neuronal stimulation and memory acquisition, suggesting the involvement of phospholipase A1 (PLA1) activity in synaptic plasticity. Here, we show that genetic ablation of the PLA1 isoform DDHD2 in mice dramatically reduces saturated FFA responses to memory acquisition across the brain. Furthermore, DDHD2 loss also decreases memory performance in reward-based learning and spatial memory models prior to the development of neuromuscular deficits that mirror human spastic paraplegia. Via pulldown-mass spectrometry analyses, we find that DDHD2 binds to the key synaptic protein STXBP1. Using STXBP1/2 knockout neurosecretory cells and a haploinsufficient STXBP1+/- mouse model of human early infantile encephalopathy associated with intellectual disability and motor dysfunction, we show that STXBP1 controls targeting of DDHD2 to the plasma membrane and generation of saturated FFAs in the brain. These findings suggest key roles for DDHD2 and STXBP1 in lipid metabolism and in the processes of synaptic plasticity, learning, and memory.

Modulation of immunity by the secreted phospholipase A2 family.

Among the phospholipase A2 (PLA2 ) superfamily, which typically catalyzes the sn-2 hydrolysis of phospholipids to yield fatty acids and lysophospholipids, the secreted PLA2 (sPLA2 ) family contains 11 isoforms in mammals. Individual sPLA2 s have unique enzymatic specificity toward fatty acids and polar heads of phospholipid substrates and display distinct tissue/cellular distributions, suggesting their distinct physiological functions. Recent studies using knockout and/or transgenic mice for a full set of sPLA2 s have revealed their roles in modulation of immunity and related disorders. Application of mass spectrometric lipidomics to these mice has enabled to identify target substrates and products of individual sPLA2 s in given tissue microenvironments. sPLA2 s hydrolyze not only phospholipids in the plasma membrane of activated, damaged or dying mammalian cells, but also extracellular phospholipids such as those in extracellular vesicles, microbe membranes, lipoproteins, surfactants, and dietary phospholipids, thereby exacerbating or ameliorating various diseases. The actions of sPLA2 s are dependent on, or independent of, the generation of fatty acid- or lysophospholipid-derived lipid mediators according to the pathophysiological contexts. In this review, we make an overview of our current understanding of the roles of individual sPLA2 s in various immune responses and associated diseases.

A novel phospholipase A2 is a core component of the typhoid toxin genetic islet.

S. Typhi, the cause of typhoid fever, is a bacterial pathogen of substantial global importance. Typhoid toxin is a secreted AB-type toxin that is a key S. Typhi virulence factor encoded within a 5-gene genetic islet. Four genes in this islet have well-defined roles in typhoid toxin biology, however the function of the fifth gene is unknown. Here, we investigate the function of this gene, which we name ttaP. We show that ttaP is co-transcribed with the typhoid toxin subunit cdtB, and we perform genomic analyses that indicate that TtaP is very highly conserved in typhoid toxin islets found in diverse salmonellae. We show that TtaP is a distant homolog of group XIV secreted phospholipase A2 (PLA2) enzymes, and experimentally demonstrate that TtaP is a bona fide PLA2. Sequence and structural analyses indicate that TtaP differs substantially from characterized PLA2s, and thus represents a novel class of PLA2. Secretion assays revealed that TtaP is neither co-secreted with typhoid toxin, nor is it required for toxin secretion. Although TtaP is a phospholipase that remains associated with the S. Typhi cell, assays that probed for altered cell envelope integrity failed to identify any differences between wild-type S. Typhi and a ttaP deletion strain. Collectively, this study identifies a biochemical activity for the lone uncharacterized typhoid toxin islet gene and lays the groundwork for exploring how this gene factors into S. Typhi pathogenesis. This study further identifies a novel class of PLA2, enzymes that have a wide range of industrial applications.

Whole transcriptome sequencing reveals the activity of the PLA2 family members in Androctonus crassicauda (Scorpionida: Buthidae) venom gland.

Phospholipase A2 is the most abundant venom gland enzyme, whose activity leads to the activation of the inflammatory response by accumulating lipid mediators. This study aimed to identify, classify, and investigate the properties of venom PLA2 isoforms. Then, the present findings were confirmed by chemically measuring the activity of PLA2. The sequences representing PLA2 annotation were extracted from the Androctonus crassicauda transcriptome dataset using BLAS searches against the local PLA2 database. We found several cDNA sequences of PLA2 classified and named by conducting multiple searches as platelet-activating factor acetylhydrolases, calcium-dependent PLA2s, calcium-independent PLA2s, and secreted PLA2s. The largest and smallest isoforms of these proteins range between approximately 70.34 kDa (iPLA2) and 17.75 kDa (cPLA2). Among sPLA2 isoforms, sPLA2GXIIA and sPLA2G3 with ORF encoding 169 and 299 amino acids are the smallest and largest secreted PLA2, respectively. These results collectively suggested that A. crassicauda venom has PLA2 activity, and the members of this protein family may have important biological roles in lipid metabolism. This study also revealed the interaction between members of PLA2s in the PPI network. The results of this study would greatly help with the classification, evolutionary relationships, and interactions between PLA2 family proteins in the gene network.

Group III secreted phospholipase A2 -driven lysophospholipid pathway protects against allergic asthma.

Asthma is a chronic inflammatory disease of the airways characterized by recurrent episodes of airway obstruction, hyperresponsiveness, remodeling, and eosinophilia. Phospholipase A2 s (PLA2 s), which release fatty acids and lysophospholipids from membrane phospholipids, have been implicated in exacerbating asthma by generating pro-asthmatic lipid mediators, but an understanding of the association between individual PLA2 subtypes and asthma is still incomplete. Here, we show that group III-secreted PLA2 (sPLA2 -III) plays an ameliorating, rather than aggravating, role in asthma pathology. In both mouse and human lungs, sPLA2 -III was expressed in bronchial epithelial cells and decreased during the asthmatic response. In an ovalbumin (OVA)-induced asthma model, Pla2g3-/- mice exhibited enhanced airway hyperresponsiveness, eosinophilia, OVA-specific IgE production, and type 2 cytokine expression as compared to Pla2g3+/+ mice. Lipidomics analysis showed that the pulmonary levels of several lysophospholipids, including lysophosphatidylcholine, lysophosphatidylethanolamine, and lysophosphatidic acid (LPA), were decreased in OVA-challenged Pla2g3-/- mice relative to Pla2g3+/+ mice. LPA receptor 2 (LPA2 ) agonists suppressed thymic stromal lymphopoietin (TSLP) expression in bronchial epithelial cells and reversed airway hyperresponsiveness and eosinophilia in Pla2g3-/- mice, suggesting that sPLA2 -III negatively regulates allergen-induced asthma at least by producing LPA. Thus, the activation of the sPLA2 -III-LPA pathway may be a new therapeutic target for allergic asthma.

Melatonin suppresses atherosclerosis by ferroptosis inhibition via activating NRF2 pathway.

Melatonin (MLT), a conserved small indole compound, exhibits anti-inflammatory and antioxidant properties, contributing to its cardioprotective effects. Lipoprotein-associated phospholipase A2 (Lp-PLA2) is associated with atherosclerosis disease risk, and is known as an atherosclerosis risk biomarker. This study aimed to investigate the impact of MLT on Lp-PLA2 expression in the atherosclerotic process and explore the underlying mechanisms involved. In vivo, ApoE-/- mice were fed a high-fat diet, with or without MLT administration, after which the plaque area and collagen content were assessed. Macrophages were pretreated with MLT combined with ox-LDL, and the levels of ferroptosis-related proteins, NRF2 activation, mitochondrial function, and oxidative stress were measured. MLT administration significantly attenuated atherosclerotic plaque progression, as evidenced by decreased plaque area and increased collagen. Compared with those in the high-fat diet (HD) group, the levels of glutathione peroxidase 4 (GPX4) and SLC7A11 (xCT, a cystine/glutamate transporter) in atherosclerotic root macrophages were significantly increased in the MLT group. In vitro, MLT activated the nuclear factor-E2-related Factor 2 (NRF2)/SLC7A11/GPX4 signaling pathway, enhancing antioxidant capacity while reducing lipid peroxidation and suppressing Lp-PLA2 expression in macrophages. Moreover, MLT reversed ox-LDL-induced ferroptosis, through the use of ferrostatin-1 (a ferroptosis inhibitor) and/or erastin (a ferroptosis activator). Furthermore, the protective effects of MLT on Lp-PLA2 expression, antioxidant capacity, lipid peroxidation, and ferroptosis were decreased in ML385 (a specific NRF2 inhibitor)-treated macrophages and in AAV-sh-NRF2 treated ApoE-/- mice. MLT suppresses Lp-PLA2 expression and atherosclerosis processes by inhibiting macrophage ferroptosis and partially activating the NRF2 pathway.

A protein-lipid complex that detoxifies free fatty acids.

Fatty acids (FAs) are well known to serve as substrates for reactions that provide cells with membranes and energy. In contrast to these metabolic reactions, the physiological importance of FAs themselves known as free FAs (FFAs) in cells remains obscure. Since accumulation of FFAs in cells is toxic, cells must develop mechanisms to detoxify FFAs. One such mechanism is to sequester free polyunsaturated FAs (PUFAs) into a droplet-like structure assembled by Fas-Associated Factor 1 (FAF1), a cytosolic protein. This sequestration limits access of PUFAs to Fe2+ , thereby preventing Fe2+ -catalyzed PUFA peroxidation. Consequently, assembly of the FAF1-FFA complex is critical to protect cells from ferroptosis, a cell death pathway triggered by PUFA peroxidation. The observations that free PUFAs in cytosol are not randomly diffused but rather sequestered into a membraneless complex should open new directions to explore signaling pathways by which FFAs regulate cellular physiology.

VpdC is a ubiquitin-activated phospholipase effector that regulates Legionella vacuole expansion during infection.

Intravacuolar pathogens need to gradually expand their surrounding vacuole to accommodate the growing number of bacterial offspring during intracellular replication. Here we found that Legionella pneumophila controls vacuole expansion by fine-tuning the generation of lysophospholipids within the vacuolar membrane. Upon allosteric activation by binding to host ubiquitin, the type IVB (Dot/Icm) effector VpdC converts phospholipids into lysophospholipids which, at moderate concentrations, are known to promote membrane fusion but block it at elevated levels by generating excessive positive membrane curvature. Consequently, L. pneumophila overproducing VpdC were prevented from adequately expanding their surrounding membrane, trapping the replicating bacteria within spatially confined vacuoles and reducing their capability to proliferate intracellularly. Quantitative lipidomics confirmed a VpdC-dependent increase in several types of lysophospholipids during infection, and VpdC production in transiently transfected cells caused tubulation of organelle membranes as well as mitochondria fragmentation, processes that can be phenocopied by supplying cells with exogenous lysophospholipids. Together, these results demonstrate an important role for bacterial phospholipases in vacuolar expansion.

Effectiveness of a novel rat model of off-target PLA2R1 knockout to renal impairment.

Phospholipase A2 receptor 1 (PLA2R1) plays a crucial role in various diseases, including membranous nephropathy. However, the precise implications of PLA2R1 deficiency remain poorly understood. In this study, we created PLA2R1 knockout rats to explore potential consequences resulting from the loss of the PLA2R1 gene. Unexpectedly, our PLA2R1 knockout rats exhibited symptoms resembling those of chronic kidney disease after an 8-week observation period. Notably, several rats developed persistent proteinuria, a hallmark of renal dysfunction. Immunohistochemical and immunofluorescence analyses revealed insignificant glomerular fibrosis, reduced podocyte count, and augmented glomerular expression of complement C3 (C3) compared to immunoglobin A (IgA) and immunoglobin G(IgG) in the rat model. These findings suggest that the loss of PLA2R1 may contribute to the pathogenesis of membranous nephropathy and related conditions. Our knockout rat model provides a valuable tool for investigating the underlying pathology of PLA2R1-associated diseases, and may facilitate the development of targeted therapies for membranous nephropathy and other related disorders.

The role of lysosomal phospholipase A2 in the catabolism of bis(monoacylglycerol)phosphate and association with phospholipidosis.

Bis(monoacylglycerol)phosphate (BMP) is an acidic glycerophospholipid localized to late endosomes and lysosomes. However, the metabolism of BMP is poorly understood. Because many drugs that cause phospholipidosis inhibit lysosomal phospholipase A2 (LPLA2, PLA2G15, LYPLA3) activity, we investigated whether this enzyme has a role in BMPcatabolism. The incubation of recombinant human LPLA2 (hLPLA2) and liposomes containing the naturally occurring BMP (sn-(2-oleoyl-3-hydroxy)-glycerol-1-phospho-sn-1'-(2'-oleoyl-3'-hydroxy)-glycerol (S,S-(2,2',C18:1)-BMP) resulted in the deacylation of this BMP isomer. The deacylation rate was 70 times lower than that of dioleoyl phosphatidylglycerol (DOPG), an isomer and precursor of BMP. The release rates of oleic acid from DOPG and four BMP stereoisomers by LPLA2 differed. The rank order of the rates of hydrolysis were DOPG>S,S-(3,3',C18:1)-BMP>R,S-(3,1',C18:1)-BMP>R,R-(1,1',C18:1)>S,S-(2,2')-BMP. The cationic amphiphilic drug amiodarone (AMD) inhibited the deacylation of DOPG and BMP isomers by hLPLA2 in a concentration-dependent manner. Under these experimental conditions, the IC50s of amiodarone-induced inhibition of the four BMP isomers and DOPG were less than 20 µM and approximately 30 µM, respectively. BMP accumulation was observed in AMD-treated RAW 264.7 cells. The accumulated BMP was significantly reduced by exogenous treatment of cells with active recombinant hLPLA2 but not with diisopropylfluorophosphate-inactivated recombinant hLPLA2. Finally, a series of cationic amphiphilic drugs known to cause phospholipidosis were screened for inhibition of LPLA2 activity as measured by either the transacylation or fatty acid hydrolysis of BMP or phosphatidylcholine as substrates. Fifteen compounds demonstrated significant inhibition with IC50s ranging from 6.8 to 63.3 µM. These results indicate that LPLA2 degrades BMP isomers with different substrate specificities under acidic conditions and may be the key enzyme associated with BMP accumulation in drug-induced phospholipidosis.

Contribution of transient receptor potential vanilloid 1 (TRPV1) channel to cholinergic contraction of rat bladder smooth muscle.

Transient receptor potential vanilloid 1 (TRPV1) is a calcium-permeable ion channel that is gated by the pungent constituent of red chili pepper, capsaicin, and by related chemicals from the group of vanilloids, in addition to noxious heat. It is expressed mostly in sensory neurons to act as a detector of painful stimuli produced by pungent chemicals and high temperatures. Although TRPV1 is also found outside the sensory nervous system, its expression and function in the bladder detrusor smooth muscle (DSM) remain controversial. Here, by using Ca2+ imaging and patch clamp on isolated rat DSM cells, in addition to tensiometry on multicellular DSM strips, we show that TRPV1 is expressed functionally in only a fraction of DSM cells, in which it acts as an endoplasmic reticulum Ca2+-release channel responsible for the capsaicin-activated [Ca2+]i rise. Carbachol-stimulated contractions of multicellular DSM strips contain a TRPV1-dependent component, which is negligible in the circular DSM but reaches ≤50% in the longitudinal DSM. Activation of TRPV1 in rat DSM during muscarinic cholinergic stimulation is ensured by phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists. Immunofluorescence detection of TRPV1 protein in bladder sections and isolated DSM cells confirmed both its preferential expression in the longitudinal DSM sublayer and its targeting to the endoplasmic reticulum. We conclude that TRPV1 is an essential contributor to the cholinergic contraction of bladder longitudinal DSM, which might be important for producing spatial and/or temporal anisotropy of bladder wall deformation in different regions during parasympathetic stimulation. KEY POINTS: The transient receptor potential vanilloid 1 (TRPV1) heat/capsaicin receptor/channel is localized in the endoplasmic reticulum membrane of detrusor smooth muscle (DSM) cells of the rat bladder, operating as a calcium-release channel. Isolated DSM cells are separated into two nearly equal groups, within which the cells either show or do not show TRPV1-dependent [Ca2+]i rise. Carbachol-stimulated, muscarinic ACh receptor-mediated contractions of multicellular DSM strips contain a TRPV1-dependent component. This component is negligible in the circular DSM but reaches ≤50% in longitudinal DSM. Activation of TRPV1 in rat DSM during cholinergic stimulation involves phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists.

Identification of the glycosylphosphatidylinositol-specific phospholipase A2 (GPI-PLA2) that mediates GPI fatty acid remodeling in Trypanosoma brucei.

The biosynthesis of glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) in the parasitic protozoan Trypanosoma brucei involves fatty acid remodeling of the GPI precursor molecules before they are transferred to protein in the endoplasmic reticulum. The genes encoding the requisite phospholipase A2 and A1 activities for this remodeling have thus far been elusive. Here, we identify a gene, Tb927.7.6110, that encodes a protein that is both necessary and sufficient for GPI-phospholipase A2 (GPI-PLA2) activity in the procyclic form of the parasite. The predicted protein product belongs to the alkaline ceramidase, PAQR receptor, Per1, SID-1, and TMEM8 (CREST) superfamily of transmembrane hydrolase proteins and shows sequence similarity to Post-GPI-Attachment to Protein 6 (PGAP6), a GPI-PLA2 that acts after transfer of GPI precursors to protein in mammalian cells. We show the trypanosome Tb927.7.6110 GPI-PLA2 gene resides in a locus with two closely related genes Tb927.7.6150 and Tb927.7.6170, one of which (Tb927.7.6150) most likely encodes a catalytically inactive protein. The absence of GPI-PLA2 in the null mutant procyclic cells not only affected fatty acid remodeling but also reduced GPI anchor sidechain size on mature GPI-anchored procyclin glycoproteins. This reduction in GPI anchor sidechain size was reversed upon the re-addition of Tb927.7.6110 and of Tb927.7.6170, despite the latter not encoding GPI precursor GPI-PLA2 activity. Taken together, we conclude that Tb927.7.6110 encodes the GPI-PLA2 of GPI precursor fatty acid remodeling and that more work is required to assess the roles and essentiality of Tb927.7.6170 and the presumably enzymatically inactive Tb927.7.6150.

Identification and characterization of GLYCEROLIPASE A1 for wound-triggered JA biosynthesis in Nicotiana benthamiana leaves.

Although many important discoveries have been made regarding the jasmonate signaling pathway, how jasmonate biosynthesis is initiated is still a major unanswered question in the field. Previous evidences suggest that jasmonate biosynthesis is limited by the availability of fatty acid precursor, such as ⍺-linolenic acid (⍺-LA). This indicates that the lipase responsible for releasing α-LA in the chloroplast, where early steps of jasmonate biosynthesis take place, is the key initial step in the jasmonate biosynthetic pathway. Nicotiana benthamiana glycerol lipase A1 (NbGLA1) is homologous to N. attenuata GLA1 (NaGLA1) which has been reported to be a major lipase in leaves for jasmonate biosynthesis. NbGLA1 was studied for its potential usefulness in a species that is more common in laboratories. Virus-induced gene silencing of both NbGLA1 and NbGLA2, another homolog, resulted in more than 80% reduction in jasmonic acid (JA) biosynthesis in wounded leaves. Overexpression of NbGLA1 utilizing an inducible vector system failed to increase JA, indicating that transcriptional induction of NbGLA1 is insufficient to trigger JA biosynthesis. However, co-treatment with wounding in addition to NbGLA1 induction increased JA accumulation several fold higher than the gene expression or wounding alone, indicating an enhancement of the enzyme activity by wounding. Domain-deletion of a 126-bp C-terminal region hypothesized to have regulatory roles increased NbGLA1-induced JA level. Together, the data show NbGLA1 to be a major lipase for wound-induced JA biosynthesis in N. benthamiana leaves and demonstrate the use of inducible promoter-driven construct of NbGLA1 in conjunction with its transient expression in N. benthamiana as a useful system to study its protein function.

Modulation of lipid profile by secretory phospholipase A2 group IIA: Verification with a transgenic mouse model.

We previously demonstrated a positive relation of secretory phospholipase A2 group IIA (sPLA2-IIA) with circulating high-density lipoprotein cholesterol (HDL-C) in patients with coronary artery disease, and sPLA2-IIA increased cholesterol efflux in THP-1 cells through peroxisome proliferator-activated receptor-γ (PPAR-γ)/liver X receptor α/ATP-binding cassette transporter A1 (ABCA1) signaling pathway. The aim of the present study was to examine the role of sPLA2-IIA over-expression on lipid profile in a transgenic mouse model. Fifteen apoE-/- and C57BL/7 female mice received bone marrow transplantation from transgenic SPLA2-IIA mice, and treated with specific PPAR-γ inhibitor GW9662. High fat diet was given after one week of bone marrow transplantation, and animals were sacrificed after twelve weeks. Immunohistochemical staining showed over-expression of sPLA2-IIA protein in the lung and spleen. The circulating level of HDL-C, but not that of low-density lipoprotein cholesterol (LDL-C), total cholesterol, or total triglyceride, was increased by sPLA2-IIA over-expression, and was subsequently reversed by GW9662 treatment. Over-expression of sPLA2-IIA resulted in augmented expression of cholesterol transporter ABCA1 at mRNA level in the aortas, and at protein level in macrophages, co-localized with macrophage specific antigen CD68. GW9662 exerted potent inhibitory effects on sPLA2-IIA-induced ABCA1 expression. Conclusively, we demonstrated the effects of sPLA2-IIA on circulating HDL-C level and the expression of ABCA1, possibly through regulation of PPAR-γ signaling in transgenic mouse model, that is in concert with the conditions in patients with coronary artery disease.

Mobile health technologies in an interventional hybrid study on actinic keratosis: Results from an early phase randomized controlled trial investigating the safety and efficacy of a cytosolic phospholipase A2 inhibitor gel in photodamaged skin.

Hybrid trials are a new trend in dermatological research that leverage mobile health technologies to decentralize a subset of clinical trial elements and thereby reduce the number of in-clinic visits. In a Phase I/IIa randomized controlled hybrid trial, the safety and efficacy of an anti-proliferative and anti-inflammatory drug inhibiting cytosolic phospholipase A2 (AVX001) was tested using 1%, 3% or vehicle gel in 60 patients with actinic keratosis (AK) and assessed in-clinic as well as remotely. Over the course of 12 weeks, patients were assessed in-clinic at baseline, end of treatment (EOT) and end of study (EOS), as well as 9 times remotely on a weekly to biweekly basis. Safety outcomes comprising local skin reactions (LSR; 0-5), adverse events (AE) and cosmesis, were graded in-clinic and remotely using patient-obtained smartphone photographs (PSPs) and questionnaires; efficacy was assessed in-clinic based on clinically visible clearance of AK target area of >50%. A total of 55 participants (91.7%) completed the treatment course. The average submission rate of PSPs was high (≥85%), of which 93% were of sufficient quality. No serious AE were reported and only two experienced temporary LSR >2 (scale 0-4) and cosmesis remained stable throughout the study. Based on the mild AE and LSR profile, daily application of AVX001 gel for 1 month appears safe, tolerable, and cosmetically acceptable for use in patients with AK. At EOT, AVX001 achieved a subtle treatment response with clearance of AK target area of >50% in 18% of patients. Remote and in-clinic assessments of LSRs were in high agreement, suggesting that the use of mobile health technologies in early-phase hybrid studies of AK does not compromise patient safety.

Targeted metabolomic analysis of serum free fatty acids: Lipidomics disturbance in patients with superior limbic keratoconjunctivitis.

This study aimed to identify and quantify free fatty acids (FFAs), secretory phospholipase A2 group IIa (sPLA2-IIa) and cytosolic phospholipase A2 (cPLA2) in serum of superior limbic keratoconjunctivitis (SLK) patients and explored the association between FFAs, sPLA2-IIa and cPLA2 variations and SLK. Targeted metabolomic analysis of FFAs in serum was performed by gas chromatography tandem mass spectrometry (GC-MS/MS) analysis on 16 SLK patients (43.88 ± 7.88 years; female: 62.50%) and 25 healthy controls (43.12 ± 7.88 years; female: 64.00%). Qualitative and absolute quantitative results of FFAs were obtained and classified according to gender and thyroid tests. Differential lipid metabolites, metabolomic pathways and biomarkers were further evaluated. The serum sPLA2-IIa and cPLA2 were determined by enzyme linked immunosorbent assay (ELISA). Among 40 FFAs identified, 6 FFAs showed significant changes (P < 0.05) in SLK patients, including 4 decreased and 2 increased. They were mainly related to unsaturated fatty acid biosynthesis, α-linolenic acid and linoleic acid metabolism, and fatty acid biosynthesis. When dividing the data by gender or abnormal thyroid tests, some comparable FFAs alterations displayed in SLK patients. The ROC analysis revealed that the AUC values of linoleic acid, γ-linolenic acid, cis-8,11,14-eicosatrienoic acid, stearic acid, and palmitic acid, were all greater than 0.8. The serum concentrations of sPLA2-IIa and cPLA2 in patients with SLK were significantly higher than that in healthy controls. Lipidomics disturbance might be the potential mechanism of SLK. Serum FFA biomarkers associated with SLK have potential for the diagnosis and treatment of the disease.

PLAAT1 deficiency alleviates high-fat diet-induced hepatic lipid accumulation in mice.

The phospholipase A and acyltransferase (PLAAT) family is composed of three isoforms in mice (PLAAT1, 3, and 5), all of which function as phospholipid-metabolizing enzymes exhibiting phospholipase A1 /A2 and acyltransferase activities. Plaat3-deficient (Plaat3-/- ) mice were previously reported to show lean phenotype and remarkable hepatic fat accumulation under high-fat diet (HFD) feeding, while Plaat1-/- mice have not been analyzed. In the present study, we generated Plaat1-/- mice and investigated the effects of PLAAT1 deficiency on HFD-induced obesity, hepatic lipid accumulation, and insulin resistance. After HFD treatment, PLAAT1 deficiency caused a lower body weight gain compared to wild-type mice. Plaat1-/- mice also showed reduced liver weight with negligible hepatic lipid accumulation. In accordance with these findings, PLAAT1 deficiency improved HFD-induced hepatic dysfunction and lipid metabolism disorders. Lipidomics analysis in the liver revealed that in Plaat1-/- mice, the levels of various glycerophospholipids tended to increase, while all classes of lysophospholipids examined tended to decrease, suggesting that PLAAT1 functions as phospholipase A1 /A2 in the liver. Interestingly, the HFD treatment of wild-type mice significantly increased the mRNA level of PLAAT1 in the liver. Furthermore, the deficiency did not appear to elevate the risk of insulin resistance in contrast to PLAAT3 deficiency. These results suggested that the suppression of PLAAT1 improves HFD-induced overweight and concomitant hepatic lipid accumulation.

Causal Effect of Lipoprotein-Associated Phospholipase A2 Activity on Ischemic Stroke: A Mendelian Randomization Study.

BACKGROUND: The relationship between ischemic stroke (IS) and lipoprotein-associated phospholipase A2 (Lp-PLA2) activity is still unclear, and there is a dearth of stratified research on the relationship between Lp-PLA2 activity and different IS subtypes. Therefore, Mendelian randomization (MR) was used in this study to examine the relationship between genetically proxied Lp-PLA2 activity and the risks of IS and its subtypes. METHODS: Based on information from a meta-analysis of genome-wide association studies, which included 13,664 European people, five single nucleotide polymorphisms related to Lp-PLA2 activity were chosen as instrumental variables. Summary statistics information about the MEGESTROKE consortium with the European group (40,585 cases and 406,111 controls) include any IS (AIS; n = 34,217), large-artery stroke (LAS; n = 4,373), cardioembolic stroke (CES; n = 7,193), and small-vessel stroke (SVS; n = 5,386). In order to determine the causal relationships between Lp-PLA2 activity and IS as well as its subtypes, the inverse-variance-weighted (IVW) approach was chosen as the primary analysis. Significant estimates were then tested by sensitivity analysis to rule out heterogeneity and pleiotropy. RESULTS: IVW showed that Lp-PLA2 activity was causally associated with LAS (odds ratio = 3.25, 95% confidence interval = 1.65-6.41, p = 0.0007) but not with other subtypes of stroke. Sensitivity analysis for causal estimates between Lp-PLA2 activity and LAS showed no significant heterogeneity or pleiotropy. CONCLUSIONS: These MR analyses support a causal effect of Lp-PLA2 activity on LAS but not on AIS, CES, or SVS, which suggests that serum Lp-PLA2 activity might be a biomarker for prediction of LAS.