Lysophosphatidic Acid Receptor Research Articles

Targeting autotaxin impacts disease advance in the SOD1-G93A mouse model of amyotrophic lateral sclerosis.

A preclinical strategy to broaden the search of potentially effective treatments in amyotrophic lateral sclerosis (ALS) relies on identifying factors controlling motor neuron (MN) excitability. These partners might be part of still unknown pathogenic pathways and/or useful for the design of new interventions to affect disease progression. In this framework, the bioactive membrane‐derived phospholipid lysophosphatidic acid (LPA) affects MN excitability through LPA receptor 1 (LPA1). Furthermore, LPA1 knockdown is neuroprotective in transgenic ALS SOD1‐G93A mice. On this basis, we raised the hypothesis that the major LPA‐synthesizing ectoenzyme, autotaxin (ATX), regulates MN excitability and is a potential target to modulate disease development in ALS mice. We show here that PF‐8380, a specific ATX inhibitor, reduced intrinsic membrane excitability (IME) of hypoglossal MNs in brainstem slices, supporting that baseline ATX activity regulates MN IME. PF‐8380‐induced alterations were prevented by a small‐interfering RNA directed against mRNA for lpa1 . These outcomes support that impact of ATX‐originated lysophospholipids on MN IME engages, at least, the G‐protein‐coupled receptor LPA1. Interestingly, mRNA atx levels increased in the spinal cord of pre‐symptomatic (1–2 months old) SOD1‐G93A mice, thus preceding MN loss. The rise in transcripts levels also occurred in cultured spinal cord MNs from SOD1‐G93A embryos, suggesting that mRNA atx upregulation in MNs is an etiopathogenic event in the ALS cell model. Remarkably, chronic administration in the drinking water of the orally bioavailable ATX inhibitor PF‐8380 delayed MN loss, motor deterioration and prolonged life span in ALS mice. Treatment also led to a reduction in LPA1‐immunoreactive patches in transgenic animals mostly in MNs. These outcomes support that neuroprotective effects of interfering with ATX in SOD1‐G93A mice rely, at least in part, on LPA1 knockdown in MNs. Therefore, we propose ATX as a potential target and/or a biomarker in ALS and highlight ATX inhibitors as reasonable tools with therapeutic usefulness for this lethal pathology.

Role of Aberrantly Activated Lysophosphatidic Acid Receptor 1 Signaling Mediated Inflammation in Renal Aging.

The increasing load of senescent cells is a source of aging, and chronic inflammation plays a pivotal role in cellular senescence. In addition, senescent renal tubular epithelial cells are closely associated with renal aging. Lysophosphatidic acid (LPA) is a bioactive lipid mainly produced by the catalytic action of autotaxin (ATX), and its ligation to LPA receptor-1 (LPAR1) is associated with chronic inflammation and renal fibrosis; however, its role in renal aging is unclear. Male 2-, 12-, and 24-month-old C57BL/6 mice and Human renal proximal tubular epithelial cells (HRPTEpiC) were used in the present study. DNA damage and oxidative stress-induced senescence were simulated using doxorubicin (DOXO) and H2O2, respectively. The aged kidney showed decreased renal function, increased fractional mesangial area, and tubulointerstitial fibrosis. Both aged kidney and senescent cells showed increased levels of LPAR1, Nuclear factor κB (NF-κB), and inflammatory cytokines. In addition, LPAR1-knockdown reduced NF-κB and subsequent inflammatory cytokine induction, and NF-κB-knockdown resulted in decreased LPAR1 expression. Our study revealed a positive feedback loop between LPAR1 and NF-κB, which reinforces the role of inflammatory response, suggesting that blocking of aberrantly activated LPAR1 may reduce excessive inflammation, thereby providing a new possible therapeutic strategy to attenuate renal aging.

The ameliorative effect of bromelain on STZ-induced type 1 diabetes in rats through Oxi-LDL/LPA/LPAR1 pathway

AimsDiabetes, a serious worldwide problem, is modulated via inflammation and oxidative stress. Bromelain, a natural compound, recently attracts interest due to its anti-inflammatory effects, while its mode of action remains not properly understood. Thus, investigating the antidiabetic effect of bromelain is promising. Materials and methodsRats were randomized into normal group, STZ group (were administrated single intraperitoneal (i.p) injection of 55 mg/kg streptozotocin (STZ)) and STZ + Bro group (were administrated single i.p injection of STZ, 72 h later were i.p administrated 10 mg/kg/day bromelain for 15 days). Wound healing ability was investigated for different groups. Spectrophotometry, ELISA, histopathological and immunohistochemical techniques were applied. Key findingsBromelain significantly decreased fasting blood glucose, serum triglycerides and cholesterol and hepatic malondialdehyde levels compared with STZ group. Moreover, Bromelain significantly increased serum albumin and total protein levels and percentage of wound healing compared with STZ group. These results were confirmed through the histopathological examination of liver, pancreas, and skin tissues. Investigating the molecular mechanism underlying these effects, STZ injection caused significant increase in hepatic oxidized-LDL (Oxi-LDL) and lysophosphatidic acid (LPA) levels and hepatic lysophosphatidic acid receptor 1 (LPAR1), and beta secretase (BACE1) protein tissue expressions, while bromelain significantly aborted these effects. Thus, STZ caused upregulation of Oxi-LDL/LPA/LPAR1/BACE1 pathway, while bromelain significantly ameliorated these effects. SignificanceTo our best knowledge, this study represents the 1st study investigating Oxi-LDL/LPA/LPAR1/BACE1 pathway in STZ-induced diabetes in rats, in addition to the promising ameliorative effect of bromelain in STZ-induced diabetes in rats.

Downregulation of acylglycerol kinase suppresses high-glucose-induced endothelial-mesenchymal transition in human retinal microvascular endothelial cells through regulating the LPAR1/TGF-β/Notch signaling pathway.

The endothelial-mesenchymal transition (EndMT) participates in the progression of diabetic retinopathy (DR), but cell-intrinsic factors modulating this process remain elusive. In this study, we explored the role of lysophosphatidic acid (LPA) - producing enzyme, acylglycerol kinase (AGK), in the EndMT of human retinal microvascular endothelial cells (HRECs) under high-glucose (HG) conditions. We found that AGK was significantly elevated in HG-treated cells. In addition, AGK knockdown reversed the HG-induced EndMT in HRECs, which was evidenced by the increased endothelial markers (CD31 and VE-cadherin) and decreased mesenchymal markers (FSP1 and α-SMA). Furthermore, downregulation of AGK inhibited the HG-induced activation of transforming growth factor β (TGF-β)/Notch pathways, whereas exogenous TGF-β1 (10 ng/mL) impeded the inhibitory effects of AGK knockdown on HG-induced EndMT in HRECs. Additionally, the silencing of AGK abolished the HG-induced upregulation of LPA and its receptor, LPA receptor 1 (LPAR1), and overexpression of LPAR1 further rescued the AGK knockdown-mediated inhibition of the EndMT process. In conclusion, we demonstrate that downregulation of AGK suppresses HG-induced EndMT in HRECs through regulating the LPAR1/TGF-β/Notch signaling pathway, indicating that AGK might be a potential therapeutic target for the treatment of DR.

Wound Healing Effect of Gintonin Involves Lysophosphatidic Acid Receptor/Vascular Endothelial Growth Factor Signaling Pathway in Keratinocytes.

Gintonin, a novel compound of ginseng, is a ligand of the lysophosphatidic acid (LPA) receptor. The in vitro and in vivo skin wound healing effects of gintonin remain unknown. Therefore, the objective of this study was to investigate the effects of gintonin on wound healing-linked responses, especially migration and proliferation, in skin keratinocytes HaCaT. In this study, 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide assay, Boyden chamber migration assay, scratch wound healing assay, and Western blot assay were performed. A tail wound mouse model was used for the in vivo test. Gintonin increased proliferation, migration, and scratch closure in HaCaT cells. It also increased the release of vascular endothelial growth factor (VEGF) in HaCaT cells. However, these increases, induced by gintonin, were markedly blocked by treatment with Ki16425, an LPA inhibitor, PD98059, an ERK inhibitor, 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis (acetoxymethyl ester), a calcium chelator, and U73122, a PLC inhibitor. The VEGF receptor inhibitor axitinib also attenuated gintonin-enhanced HaCaT cell proliferation. Gintonin increased the phosphorylation of AKT and ERK1/2 in HaCaT cells. In addition, gintonin improved tail wound healing in mice. These results indicate that gintonin may promote wound healing through LPA receptor activation and/or VEGF release-mediated downstream signaling pathways. Thus, gintonin could be a beneficial substance to facilitate skin wound healing.

Lysophosphatidic acid receptor 6 regulated by miR-27a-3p attenuates tumor proliferation in breast cancer

PurposeLysophosphatidic acid (LPA) is a bioactive molecule which participates in many physical and pathological processes. Although LPA receptor 6 (LPAR6), the last identified LPA receptor, has been reported to have diverse effects in multiple cancers, including breast cancer, its effects and functioning mechanisms are not fully known.MethodsMultiple public databases were used to investigate the mRNA expression of LPAR6, its prognostic value, and potential mechanisms in breast cancer. Western blotting was performed to validate the differential expression of LPAR6 in breast cancer tissues and their adjacent tissues. Furthermore, in vitro experiments were used to explore the effects of LPAR6 on breast cancer. Additionally, TargetScan and miRWalk were used to identify potential upstream regulating miRNAs and validated the relationship between miR-27a-3p and LPAR6 via real-time polymerase chain reaction and an in vitro rescue assay.ResultsLPAR6 was significantly downregulated in breast cancer at transcriptional and translational levels. Decreased LPAR6 expression in breast cancer is significantly correlated with poor overall survival, disease-free survival, and distal metastasis-free survival, particularly for hormone receptor-positive patients, regardless of lymph node metastatic status. In vitro gain and loss-of-function assays indicated that LPAR6 attenuated breast cancer cell proliferation. The analyses of TCGA and METABRIC datasets revealed that LPAR6 may regulate the cell cycle signal pathway. Furthermore, the expression of LPAR6 could be positively regulated by miR-27a-3p. The knockdown of miR-27a-3p increased cell proliferation, and ectopic expression of LPAR6 could partly rescue this phenotype.ConclusionLPAR6 acts as a tumor suppressor in breast cancer and is positively regulated by miR-27a-3p.

Autotaxin-LPA-LPP3 Axis in Energy Metabolism and Metabolic Disease.

Besides serving as a structural membrane component and intermediate of the glycerolipid metabolism, lysophosphatidic acid (LPA) has a prominent role as a signaling molecule through its binding to LPA receptors at the cell surface. Extracellular LPA is primarily produced from lysophosphatidylcholine (LPC) through the activity of secreted lysophospholipase D, autotaxin (ATX). The degradation of extracellular LPA to monoacylglycerol is mediated by lipid phosphate phosphatases (LPPs) at the cell membrane. This review summarizes and interprets current literature on the role of the ATX-LPA-LPP3 axis in the regulation of energy homeostasis, insulin function, and adiposity at baseline and under conditions of obesity. We also discuss how the ATX-LPA-LPP3 axis influences obesity-related metabolic complications, including insulin resistance, fatty liver disease, and cardiomyopathy.

Lysophospholipid (LPA) receptors in GtoPdb v.2021.3

Lysophosphatidic acid (LPA) receptors (nomenclature as agreed by the NC-IUPHAR Subcommittee on Lysophospholipid Receptors [55, 19, 82, 129]) are activated by the endogenous phospholipid LPA. The first receptor, LPA1, was identified as ventricular zone gene-1 (vzg-1) [40], This discovery represented the beginning of the de-orphanisation of members of the endothelial differentiation gene (edg) family, as other LPA and sphingosine 1-phosphate (S1P) receptors were found. Five additional LPA receptors (LPA2,3,4,5,6) have since been identified [82] and their gene nomenclature codified for human LPAR1, LPAR2, etc. (HUGO Gene Nomenclature Committee, HGNC) and Lpar1, Lpar2, etc. for mice (Mouse Genome Informatics Database, MGI) to reflect species and receptor function of their corresponding proteins. The crystal structure of LPA1 is solved and indicates that LPA accesses the extracellular binding pocket, consistent with its proposed delivery via autotaxin [13]. These studies have also implicated cross-talk with endocannabinoids via phosphorylated intermediates that can also activate these receptors. The binding affinities to LPA1 of unlabeled, natural LPA and anandamide phosphate (AEAp) were measured using backscattering interferometry (pKd = 9) [83, 104]. Utilization of this method indicated affinities that were 77-fold lower than when measured using radioactivity-based protocols [128]. Targeted deletion of LPA receptors has clarified signalling pathways and identified physiological and pathophysiological roles. Multiple groups have independently published validation of all six LPA receptors described in these tables, and further validation was achieved using a distinct read-out via a novel TGFα "shedding* assay [48]. LPA has been proposed to be a ligand for GPR35 [94], supported by a study revealing that LPA modulates macrophage function through GPR35 [54]. However chemokine (C-X-C motif) ligand 17 (CXCL17) is reported to be a ligand for GPR35/CXCR8 [76]. Moreover, LPA has also been described as an agonist for the transient receptor potential (Trp) ion channels TRPV1 [87] and TRPA1 [58]. All of these proposed non-GPCR receptor identities require confirmation and are not currently recognized as bona fide LPA receptors.

Generation of an Lpar1-EGFP Fusion Knock-in Transgenic Mouse Line

Lysophosphatidic acid (LPA) is a lysophospholipid that acts as an extracellular signal through the activation of cognate G protein-coupled receptors (GPCRs). There are six known LPA receptors (LPA1–6). The first such receptor, LPA1, was identified in the embryonic brain and has been studied extensively for gene expression throughout the body, including through studies of receptor-null mice. However, identifying receptor protein expression in situ and in vivo within living cells and tissues has been difficult because of biologically low receptor expression and variable antibody specificity. To visualize native LPA1 receptor expression in situ, we generated a knock-in mouse produced by homologous recombination in murine embryonic stem (ES) cells to replace a wildtype Lpar1 allele with a mutant allele created by in-frame fusion of EGFP to the 4th exon of Lpar1 (Lpar1-EGFP knock-in allele). Homozygous knock-in mice appeared normal and the expected mendelian ratios of knock-in allele transmission were present in females and males. Histological assessments of the fetal and adult central nervous system (CNS) demonstrated expression patterns that were consistent with prior in situ hybridization studies. This new mouse line will be useful for studies of LPA1 in the developing and adult CNS, as well as other tissues, and for receptor assessments in living tissues and disease models.

Asthmatic allergen inhalation sensitises carotid bodies to lysophosphatidic acid

The carotid bodies are multimodal sensors that regulate various autonomic reflexes. Recent evidence demonstrates their role in immune reflex regulation. Our previous studies using the allergen (ovalbumin) sensitised and exposed Brown Norway rat model of asthma suggest that carotid bodies mediate asthmatic bronchoconstriction through a lysophosphatidic acid (LPA) receptor (LPAr)-protein kinase C epsilon (PKCε)-transient receptor potential vanilloid one channel (TRPV1) pathway. Whilst naïve carotid bodies respond to LPA, whether their response to LPA is enhanced in asthma is unknown. Here, we show that asthmatic sensitisation of Brown Norway rats involving repeated aerosolised allergen challenges over 6 days, results in an augmentation of the carotid bodies’ acute sensitivity to LPA. Increased expression of LPAr in the carotid bodies and petrosal ganglia likely contributed to this sensitivity. Importantly, allergen sensitisation of the carotid bodies to LPA did not alter their hypoxic response, nor did hypoxia augment LPA sensitivity acutely. Our data demonstrate the ability of allergens to sensitise the carotid bodies, highlighting the likely role of the carotid bodies and blood-borne inflammatory mediators in asthma.

LPA1-mediated PKD2 activation promotes LPA-induced tissue factor expression via the p38α and JNK2 MAPK pathways in smooth muscle cells

Tissue factor (TF) is the principal initiator of blood coagulation and is necessary for thrombosis. We previously reported that lysophosphatidic acid (LPA), a potent bioactive lipid, highly induces TF expression at the transcriptional level in vascular smooth muscle cells. To date, however, the specific role of the LPA receptor is unknown, and the intracellular signaling pathways that lead to LPA induction of TF have been largely undetermined. In the current study, we found that LPA markedly induced protein kinase D (PKD) activation in mouse aortic smooth muscle cells (MASMCs). Small-interfering RNA-mediated knockdown of PKD2 blocked LPA-induced TF expression and activity, indicating that PKD2 is the key intracellular mediator of LPA signaling leading to the expression and cell surface activity of TF. Furthermore, our data reveal a novel finding that PKD2 mediates LPA-induced TF expression via the p38α and JNK2 MAPK signaling pathways, which are accompanied by the PKD-independent MEK1/2-ERK-JNK pathway. To identify the LPA receptor(s) responsible for LPA-induced TF expression, we isolated MASMCs from LPA receptor-knockout mice. Our results demonstrated that SMCs isolated from LPA receptor 1 (LPA1)-deficient mice completely lost responsiveness to LPA stimulation, which mediates induction of TF expression and activation of PKD and p38/JNK MAPK, indicating that LPA1 is responsible for PKD2-mediated activation of JNK2 and p38α. Taken together, our data reveal a new signaling mechanism in which the LPA1-PKD2 axis mediates LPA-induced TF expression via the p38α and JNK2 pathways. This finding provides new insights into LPA signaling, the PKD2 pathway, and the mechanisms of coagulation/atherothrombosis.

2-Carba-lysophosphatidic acid is a novel \u03b2-lysophosphatidic acid analogue with high potential for lysophosphatidic acid receptor activation and autotaxin inhibition

Cyclic phosphatidic acid (cPA) is a naturally occurring phospholipid mediator that, along with its chemically stabilized analogue 2-carba-cyclic phosphatidic acid (2ccPA), induces various biological activities in vitro and in vivo. Although cPA is similar to lysophosphatidic acid (LPA) in structure and synthetic pathway, some of cPA biological functions apparently differ from those reported for LPA. We previously investigated the pharmacokinetic profile of 2ccPA, which was found to be rapidly degraded, especially in acidic conditions, yielding an unidentified compound. Thus, not only cPA but also its degradation compound may contribute to the biological activity of cPA, at least for 2ccPA. In this study, we determined the structure and examined the biological activities of 2-carba-lysophosphatidic acid (2carbaLPA) as a 2ccPA degradation compound, which is a type of β-LPA analogue. Similar to LPA and cPA, 2carbaLPA induced the phosphorylation of the extracellular signal-regulated kinase and showed potent agonism for all known LPA receptors (LPA1–6) in the transforming growth factor-α (TGFα) shedding assay, in particular for LPA3 and LPA4. 2carbaLPA inhibited the lysophospholipase D activity of autotaxin (ATX) in vitro similar to other cPA analogues, such as 2ccPA, 3-carba-cPA, and 3-carba-LPA (α-LPA analogue). Our study shows that 2carbaLPA is a novel β-LPA analogue with high potential for the activation of some LPA receptors and ATX inhibition.

CircLPAR1/miR-212-3p/ZNF217 feedback loop promotes amyloid β-induced neuronal injury in Alzheimer’s Disease

BackgroundAlzheimer’s Disease (AD) is a neurodegenerative disorder characterized by progressive memory loss and cognition and language impairment. CircRNA lysophosphatidic acid receptor 1 (circLPAR1) was found to be increased in AD patients, however, the potential role of circLPAR1 in AD process remains unclear. MethodsBeta-amyloid (Aβ) 25–35-stimulated CHP-212 and IMR-32 cells were used to perform expression and function analyses. The expression of genes and proteins was determined by qRT-PCR and Western blot. Cell proliferation and apoptosis were analyzed using cell counting kit-8 (CCK-8) assay, flow cytometry, and Western blot, respectively. ELISA analysis was used to detect the levels of interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). The levels of reactive oxygen species (ROS), lactate dehydrogenase (LDH) and superoxide dismutase (SOD) were detected using commercial kits. The direct interactions between miR-212-3p and ZNF217 (Zinc finger protein 217) or circLPAR1 was verified using dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. ResultsCircLPAR1 was highly expressed in AD patients and Aβ25-35-stimulated CHP-212 and IMR-32 cells. Knockdown of circLPAR1 suppressed Aβ25-35-induced neuronal apoptosis, inflammation, and oxidative stress. Mechanistically, circLPAR1 competitively bound to miR-212-3p to elevate its target ZNF217. Rescue experiments suggested that miR-212-3p inhibition reversed circLPAR1 silencing-evoked inhibition on neuronal injury under Aβ25-35 stimulation. Moreover, miR-212-3p re-expression reduced Aβ25-35-induced neuronal apoptosis, inflammation, and oxidative stress, which were abolished by ZNF217 up-regulation. ConclusionCircLPAR1 promotes Aβ25-35-induced apoptosis, inflammation, and oxidative stress via miR-212-3p/ZNF217 axis, suggesting a new insight into the pathogenesis of AD.

Drivers and suppressors of triple-negative breast cancer

To identify regulators of triple-negative breast cancer (TNBC), gene expression profiles of malignant parts of TNBC (mTNBC) and normal adjacent (nadj) parts of the same breasts have been compared. We are interested in the roles of estrogen receptor β (ERβ) and the cytochrome P450 family (CYPs) as drivers of TNBC. We examined by RNA sequencing the mTNBC and nadj parts of five women. We found more than a fivefold elevation in mTNBC of genes already known to be expressed in TNBC: BIRC5/survivin, Wnt-10A and -7B, matrix metalloproteinases (MMPs), chemokines, anterior gradient proteins, and lysophosphatidic acid receptor and the known basal characteristics of TNBC, sox10, ROPN1B, and Col9a3. There were two unexpected findings: 1) a strong induction of CYPs involved in activation of fatty acids (CYP4), and in inactivation of calcitriol (CYP24A1) and retinoic acid (CYP26A1); and 2) a marked down-regulation of FOS, FRA1, and JUN, known tethering partners of ERβ. ERβ is expressed in 20 to 30% of TNBCs and is being evaluated as a target for treating TNBC. We used ERβ+ TNBC patient-derived xenografts in mice and found that the ERβ agonist LY500703 had no effect on growth or proliferation. Expression of CYPs was confirmed by immunohistochemistry in formalin-fixed and paraffin-embedded (FFPE) TNBC. In TNBC cell lines, the CYP4Z1-catalyzed fatty acid metabolite 20-hydroxyeicosatetraenoic acid (20-HETE) increased proliferation, while calcitriol decreased proliferation but only after inhibition of CYP24A1. We conclude that CYP-mediated pathways can be drivers of TNBC but that ERβ is unlikely to be a tumor suppressor because the absence of its main tethering partners renders ERβ functionless on genes involved in proliferation and inflammation.

Lysophosphatidic Acid Signaling in Cancer Cells: What Makes LPA So Special?

Lysophosphatidic acid (LPA) refers to a family of simple phospholipids that act as ligands for G protein-coupled receptors. While LPA exerts effects throughout the body in normal physiological circumstances, its pathological role in cancer is of great interest from a therapeutic viewpoint. The numerous LPA receptors (LPARs) are coupled to a variety of G proteins, and more than one LPAR is typically expressed on any given cell. While the individual receptors signal through conventional GPCR pathways, LPA is particularly efficacious in stimulating cancer cell proliferation and migration. This review addresses the mechanistic aspects underlying these pro-tumorigenic effects. We provide examples of LPA signaling responses in various types of cancers, with an emphasis on those where roles have been identified for specific LPARs. While providing an overview of LPAR signaling, these examples also reveal gaps in our knowledge regarding the mechanisms of LPA action at the receptor level. The current understanding of the LPAR structure and the roles of LPAR interactions with other receptors are discussed. Overall, LPARs provide insight into the potential molecular mechanisms that underlie the ability of individual GPCRs (or combinations of GPCRs) to elicit a unique spectrum of responses from their agonist ligands. Further knowledge of these mechanisms will inform drug discovery, since GPCRs are promising therapeutic targets for cancer.

Identification of GNA12-driven gene signatures and key signaling networks in ovarian cancer.

With the focus on defining the oncogenic network stimulated by lysophosphatidic acid (LPA) in ovarian cancer, the present study sought to interrogate the oncotranscriptome regulated by the LPA-mediated signaling pathway. LPA, LPA-receptor (LPAR) and LPAR-activated G protein 12 α-subunit, encoded by G protein subunit α 12 (GNA12), all serve an important role in ovarian cancer progression. While the general signaling mechanism regulated by LPA/LPAR/GNA12 has previously been characterized, the global transcriptomic network regulated by GNA12 in ovarian cancer pathophysiology remains largely unknown. To define the LPA/LPAR/GNA12-orchestrated oncogenic networks in ovarian cancer, transcriptomic and bioinformatical analyses were conducted using SKOV3 cells, in which the expression of GNA12 was silenced. Array analysis was performed in Agilent SurePrint G3 Human Comparative Genomic Hybridization 8×60 microarray platform. The array results were validated using Kuramochi cells. Gene and functional enrichment analyses were performed using Database for Annotation, Visualization and Integrated Discovery, Search Tool for Retrieval of Interacting Genes and Cytoscape algorithms. The results indicated a paradigm in which GNA12 drove ovarian cancer progression by upregulating a pro-tumorigenic network with AKT1, VEGFA, TGFB1, BCL2L1, STAT3, insulin-like growth factor 1 and growth hormone releasing hormone as critical hub and/or bottleneck nodes. Moreover, GNA12 downregulated a growth-suppressive network involving proteasome 20S subunit (PSM) β6, PSM α6, PSM ATPase 5, ubiquitin conjugating enzyme E2 E1, PSM non-ATPase 10, NDUFA4 mitochondrial complex-associated, NADH:ubiquinone oxidoreductase subunit B8 and anaphase promoting complex subunit 1 as hub or bottleneck nodes. In addition to providing novel insights into the LPA/LPAR/GNA12-regulated oncogenic networks in ovarian cancer, the present study identified several potential nodes in this network that could be assessed for targeted therapy.

Inhibition of autotaxin by bile salts and bile salt-like molecules increases its expression by feedback regulation

BackgroundAutotaxin is an enzyme that converts lysophospholipid into lysophosphatidic acid (LPA), a highly potent signaling molecule through a range of LPA receptors. It is therefore important to investigate which factors play a role in regulating ATX expression. Since we have reported that ATX levels increase dramatically in patients with various forms of cholestasis, we embarked on a study to reveal factors that influence the enzyme activity ATX as well as its expression level in vitro and in vivo. MethodsBile from cholestatic patients was fractionated by HPLC and analyzed for modulation of ATX activity. ATX expression was measured in fibroblasts upon stimulation or inhibition of LPA signaling. ResultsSurprisingly, ATX activity was stimulated by most forms of its product LPA, but it was inhibited by bile salts and bile salt-like molecules, particularly by 3-OH sulfated bile salts and sulfated progesterone metabolites that are known to accumulate during chronic cholestasis and cholestasis of pregnancy, respectively. Activation of fibroblasts by LPA decreased ATX expression by 72%. Conversely, inhibition of LPA signaling increased ATX expression 3-fold, indicating strong feedback regulation by LPA signaling. In fibroblasts, we could verify that inhibition of ATX activity by bile salts induces its expression. Furthermore, induction of cholestasis in mice causes increased plasma ATX activity. ConclusionsMultiple biliary compounds that accumulate in the systemic circulation during cholestasis inhibit ATX activity and thereby increase ATX expression through feedback regulation. This mechanism may contribute to increased serum ATX activity in patients with cholestasis.

TNRC6C-AS1 Promotes Thyroid Cancer Progression by Upregulating LPAR5 via miR-513c-5p.

BackgroundConsidering the combined role of long non-coding RNA (lncRNAs)-microRNA (miRNA)-mRNA in tumorigenesis, the purpose of this study was to investigate how TNRC6C-AS1 regulates the expression of lysophosphatidic acid receptor 5 (LPAR5) by modulating miR-513c-5p, thus influencing the progression of thyroid cancer (THCA).MethodsqRT-PCR and Western blotting were performed to detect the expression levels of TNRC6C-AS1, miR-513c-5p, and LPAR5 in THCA tissues and cell lines. The viability, proliferation, migration, and invasion were assessed using CCK-8, BrdU, wound healing, and transwell migration assays, respectively. Dual-luciferase reporter assay, RIP assay, and RNA pull-down assay were used to evaluate the relationship between TNRC6C-AS1, miR-513c-5p, and LPAR5.ResultsTNRC6C-AS1 was highly expressed in THCA tissues, and knockout of TNRC6C-AS1 reduced the viability, proliferation, migration, and invasion of THCA cells. TNRC6C-AS1 competitively adsorbed miR-513c-5p. In addition, the biological function of TNRC6C-AS1 was blocked by knocking down the thyroid cell line TNRC6C-AS1 with miR-513c-5p inhibitor transfection. LPAR5 is the target gene for miR-513c-5p, which has the ability to eliminate the influence of miR-513c-5p on THCA cells.ConclusionThe TNRC6C-AS1/miR-513c-5p/LPAR5 axis is a novel signaling pathway that modulates THCA progression and may be a potential target for cancer therapy.

Establishment of a novel, cell-based autotaxin assay

Autotaxin (ATX) plays an important role in (patho-)physiological lysophosphatidic acid (LPA) signaling. Here we describe the establishment of novel cell-based ATX assay formats. ATX-mediated LPA generation is detected by using a stable LPA receptor reporter cell line. In a first assay variant, ATX-mediated LPA generation is started in the absence of cells and the reaction mix is transferred to the reporter cells after stopping the reaction (two-tube assay). In a second assay variant, ATX is added to the reporter cells expressing the known autotaxin binding partners integrin β1, integrin β3 and the LPA receptor 1. LPA generation is started in the presence of cells and is detected in real-time (one-tube assay). Structurally diverse ATX inhibitors with different binding modes were characterized in both cell-based assay variants and were also tested in the well-established biochemical choline release assay. ATX inhibitors displayed similar potencies, regardless if the assay was performed in the absence or presence of cells, and comparable results were obtained in all three assay formats. In summary, our novel cell-based ATX assay formats are well-suited for sensitive detection of enzyme activity as well as for the characterization of ATX inhibitors in the presence and absence of cells.

LPA and Autotaxin: Potential Drug Targets in Asthma?

Lysophosphatidic acid (LPA) is a versatile lysolipid, and activates a variety of signaling cascades in many cell types. Extracellular LPA is produced from lysophosphatidylcholine (LPC) by the enzyme autotaxin (ATX), and binds to a family of G-protein coupled receptors on its target cells. Research by many groups continues to support the idea that LPA, and the ATX-LPA axis, have important roles in asthma and allergic airway inflammation. In vitro studies have shown that LPA activates many cell types implicated in airway inflammation, including eosinophils, mast cells, dendritic cells, lymphocytes, airway epithelial cells, and airway smooth muscle cells. In animal models ATX and LPA receptor antagonists have been shown to attenuate allergic airway inflammation and hyperreactivity, cardinal features of asthma in humans. ATX and LPA antagonists are currently under active development to treat lung fibrosis, cancer, and other conditions. If compounds with acceptable safety profiles can be identified, then it seems likely that they will be useful in inflammatory lung diseases like asthma.