Dysfunction of estrogen-related receptor alpha-dependent hepatic VLDL secretion contributes to sex disparity in NAFLD/NASH development.

Rationale: Men and postmenopausal women are more prone to developing non-alcoholic fatty liver disease/steatohepatitis (NAFLD/NASH) than premenopausal women. However, the pathological links and underlying mechanisms of this disparity are still elusive. The sex-difference in hepatic very low-density lipoprotein (VLDL) assembly and secretion may contribute to NAFLD development. Estrogen-related receptor alpha (ERRα) is a key regulator of several metabolic processes. We hypothesized that ERRα plays a role contributing to the sex-difference in hepatic VLDL assembly and secretion. Methods: VLDL secretion and essential genes governing said process were assessed in male and female mice. Liver-specific ERRα-deficient (ERRαLKO) mice were generated to assess the rate of hepatic VLDL secretion and alteration in target gene expression. Overexpression of either microsomal triglyceride transfer protein (Mttp) or phospholipase A2 G12B (Pla2g12b) by adenovirus was performed to test if the fatty liver phenotype in male ERRαLKO mice was due to defects in hepatic VLDL secretion. Female ERRαLKO mice were put on a diet high in saturated fat, fructose and cholesterol (HFHC) to promote NASH development. Wild type female mice were either ovariectomized or treated with tamoxifen to induce a state of estrogen deficiency or disruption in estrogen signaling. Adenovirus was used to overexpress ERRα in these mice to test if ERRα was sufficient to rescue the suppressed VLDL secretion due to estrogen dysfunction. Finally, wild type male mice on a high-fat diet (HFD) were treated with an ERRα inverse agonist to assess if suppressing ERRα activity pharmacologically would lead to fatty liver development. Results: ERRα is an indispensable mediator modulating hepatic triglyceride-rich very low-density lipoprotein (VLDL-TG) assembly and secretion through coordinately controlling target genes apolipoprotein B (Apob), Mttp and Pla2g12b in a sex-different manner. Hepatic VLDL-TG secretion is blunted in ERRαLKO mice, leading to hepatosteatosis which exacerbates endoplasmic reticulum stress and inflammation paving ways for NASH development. Importantly, ERRα acts downstream of estrogen/ERα signaling in contributing to the sex-difference in hepatic VLDL secretion effecting hepatic lipid homeostasis. Conclusions: Our results highlight ERRα as a key mediator which contributes to the sex disparity in NAFLD development, suggesting that selectively restoring ERRα activity in the liver may be a novel strategy for treating NAFLD/NASH.

PLA2G10 facilitates the cell-cycle progression of soft tissue leiomyosarcoma cells at least by elevating cyclin E1/CDK2 expression.

Soft tissue leiomyosarcoma (STLMS) is a major histological subtype of adult sarcoma. Although the molecular mechanisms ofLMS have been gradually revealed, no valid therapeutic targets have been identified. In this study, we performed a systematic screening to explore relapse-associated genes in STLMS, using data from The Cancer Genome Atlas-Sarcoma (TCGA-SARC). Then, we investigated the functional role of the gene with the best relapse-prediction value in STLMS by both in-vitro and in-vivo studies. Results showed that AMH and PLA2G10 were two genes with area under curve (AUC) values higher than 0.80 in ROC analysis when detecting relapse. Patients in the high AMH or PLA2G10 expression group had significantly worse relapse-free survival (RFS) compared to the respective low expression group. PLA2G10 was highly expressed in STLMS, but not in other sarcoma subtypes. PLA2G10 overexpression promoted SK-LMS-1 cell growth and G1/S transition, while PLA2G10 knockdown slowed the growth and resulted in G1 phase arrest. PLA2G10 overexpression markedly increased the expression of CDK2 and cyclin E1, but did not influence CDK4, CDK6, cyclin D1, CDK1 or cyclin A expression. PLA2G10 overexpression enhanced SK-LMS-1 cell-derived xenograft tumor growth in nude mice, while PLA2G10 inhibition slowed the growth. Mutation of two critical catalyzing amino acid residues (p.H88A and p.D89A) abrogated the capability of PLA2G10 to catalyze the production of arachidonic acid (AA), and also canceled the regulatory effects on cyclin E1 and CDK2 expression, as well as G1/S transition. In conclusion, PLA2G10 was a specific relapse-associated gene in STLMS. It facilitated the cell-cycle progression of STLMS cells at least by elevating the expression of cyclin E1 and CDK2. The hydrolytic activity was crucial for its oncogenic properties.

Estrogen-related receptor γ regulates hepatic triglyceride metabolism through phospholipase A2 G12B.

Hypersecretion of hepatic very LDL (VLDL)-associated triglyceride (TG) is the hallmark of hypertriglyceridemia. The estrogen-related receptor γ (ERRγ), an orphan nuclear receptor, plays crucial roles in the regulation of metabolic homeostasis, including TG formation in the liver. It remains unclear whether ERRγ regulates hepatic VLDL-TG secretion. We demonstrated that knockdown of ERRγ impairs hepatic VLDL-TG secretion in mice, whereas overexpression of ERRγ favors the secretion, indicating a novel role of ERRγ in hepatic TG metabolism. We found that ERRγ transcriptionally regulates the expression of PLA2G12B by binding to the promoter region of the Pla2g12b gene. In Pla2g12b-null mice, ERRγ fails to regulate hepatic VLDL-TG secretion. There is an apparent accumulation of large lipid droplets in the liver of Pla2g12b-null mice. These data suggest that ERRγ is a novel regulator of hepatic VLDL-TG secretion, which is mediated through the action on PLA2G12B.-Chen, L., Wu, M., Zhang, S., Tan, W., Guan, M., Feng, L., Chen, C., Tao, J., Chen, L., Qu, L. Estrogen-related receptor γ regulates hepatic triglyceride metabolism through phospholipase A2 G12B.

Secreted PLA2 group X orchestrates innate and adaptive immune responses to inhaled allergen.

Phospholipase A2 (PLA2) enzymes regulate the formation of eicosanoids and lysophospholipids that contribute to allergic airway inflammation. Secreted PLA2 group X (sPLA2-X) was recently found to be increased in the airways of asthmatics and is highly expressed in airway epithelial cells and macrophages. In the current study, we show that allergen exposure increases sPLA2-X in humans and in mice, and that global deletion of Pla2g10 results in a marked reduction in airway hyperresponsiveness (AHR), eosinophil and T cell trafficking to the airways, airway occlusion, generation of type-2 cytokines by antigen-stimulated leukocytes, and antigen-specific immunoglobulins. Further, we found that Pla2g10-/- mice had reduced IL-33 levels in BALF, fewer type-2 innate lymphoid cells (ILC2s) in the lung, less IL-33-induced IL-13 expression in mast cells, and a marked reduction in both the number of newly recruited macrophages and the M2 polarization of these macrophages in the lung. These results indicate that sPLA2-X serves as a central regulator of both innate and adaptive immune response to proteolytic allergen.

Activation of farnesoid X receptor promotes triglycerides lowering by suppressing phospholipase A2 G12B expression.

As a novel mediator of hepatic very low-density lipoproteins (VLDL) secretion, phospholipase A2 G12B (PLA2G12B) is transcriptionally regulated by hepatocyte nuclear factor-4 alpha (HNF-4α). Farnesoid X receptor (FXR) plays a critical role in maintaining bile acids and triglycerides (TG) homeostasis. Here we report that FXR regulates serum TG level in part through PLA2G12B. Activation of FXR by chenodeoxycholic acid (CDCA) or GW4064 significantly decreased PLA2G12B expression in HepG2 cells. PLA2G12B expression was transcriptionally repressed due to an FXR-mediated up-regulation of small heterodimer partner (SHP) which functionally suppresses HNF-4α activity. We found that hepatic PLA2G12B expression was suppressed and serum TG level reduced in high fat diet mice treated with CDCA. Concurrently, CDCA treatment lowered hepatic VLDL-TG secretion. Our data demonstrate that activation of FXR promotes TG lowering, not only by decreasing de novo lipogenesis but also reducing hepatic secretion of TG-rich VLDL particles in part through suppressing PLA2G12B expression.

Progesterone-induced Acrosome Exocytosis Requires Sequential Involvement of Calcium-independent Phospholipase A2ß (iPLA2ß) and Group X Secreted Phospholipase A2 (sPLA2).

Phospholipase A2 (PLA2) activity has been shown to be involved in the sperm acrosome reaction (AR), but the molecular identity of PLA2 isoforms has remained elusive. Here, we have tested the role of two intracellular (iPLA2ß and cytosolic PLA2α) and one secreted (group X) PLA2s in spontaneous and progesterone (P4)-induced AR by using a set of specific inhibitors and knock-out mice. iPLA2ß is critical for spontaneous AR, whereas both iPLA2ß and group X secreted PLA2 are involved in P4-induced AR. Cytosolic PLA2α is dispensable in both types of AR. P4-induced AR spreads over 30 min in the mouse, and kinetic analyses suggest the presence of different sperm subpopulations, using distinct PLA2 pathways to achieve AR. At low P4 concentration (2 µm), sperm undergoing early AR (0-5 min post-P4) rely on iPLA2ß, whereas sperm undergoing late AR (20-30 min post-P4) rely on group X secreted PLA2. Moreover, the role of PLA2s in AR depends on P4 concentration, with the PLA2s being key actors at low physiological P4 concentrations (≤2 µm) but not at higher P4 concentrations (~10 µm).

Phospholipase A2 Group III and Group X Have Opposing Associations with Prognosis in Colorectal Cancer.

BACKGROUND: Although secretory phospholipase A2 (sPLA2) has been shown to be involved in various biological processes, its specific roles in sub-types of cancer development remain to be elucidated. MATERIALS AND METHODS: We examined the expression of sPLA2 group III (GIII) in 142 patients with colorectal cancer using immunohistochemistry, and its correlation with clinicopathological features and outcomes. In addition, we examined the co-expression of sPLA2GIII and sPLA2GX using serial tissue sections to clarify the roles of both proteins in colorectal carcinogenesis. RESULTS: In 66 cases, diffuse staining of sPLA2GIII was seen; this was defined as the group with high expression. High expression was associated with a significantly higher rate of lymph node metastasis (p=0.02) and poorer survival (p=0.03) compared with low expression. Patients with low sPLA2GIII and high sPLA2GX expression had a significantly higher survival rate than those with high sPLA2GIII and low sPLA2GX expression (p=0.038). CONCLUSION: sPLA2GIII expression may be used as a risk factor for lymph node metastasis and a prognostic marker in colorectal cancer. In addition, sPLA2GIII and sPLA2GX may play opposing roles in colorectal carcinogenesis.

Lack of group X secreted phospholipase A2 increases survival following pandemic H1N1 influenza infection.

The role of Group X secreted phospholipase A2 (GX-sPLA2) during influenza infection has not been previously investigated. We examined the role of GX-sPLA2 during H1N1 pandemic influenza infection in a GX-sPLA2 gene targeted mouse (GX(-/-)) model and found that survival after infection was significantly greater in GX(-/-) mice than in GX(+/+) mice. Downstream products of GX-sPLA2 activity, PGD2, PGE2, LTB4, cysteinyl leukotrienes and Lipoxin A4 were significantly lower in GX(-/-) mice BAL fluid. Lung microarray analysis identified an earlier and more robust induction of T and B cell associated genes in GX(-/-) mice. Based on the central role of sPLA2 enzymes as key initiators of inflammatory processes, we propose that activation of GX-sPLA2 during H1N1pdm infection is an early step of pulmonary inflammation and its inhibition increases adaptive immunity and improves survival. Our findings suggest that GX-sPLA2 may be a potential therapeutic target during influenza.

Secreted phospholipases A2are differentially expressed and epigenetically silenced in human breast cancer cells.

Secreted phospholipases A2 (sPLA2s) have recently been associated with several cancers, but their role in breast cancer is unknown. Here we demonstrate that mRNA expression of group IIA, III and X sPLA2s differs both in vivo in tumour biopsies and in breast cancer cells in vitro. Their expression is differentially regulated by DNA methylation and histone acetylation and, significantly, all three genes are silenced in aggressive triple negative cells due to both mechanisms. The transcription start site promoter region and the upstream CpG islands, exclusive to the group X sPLA2 gene, have variable roles in the regulation of sPLA2 expression. Our results suggest that the differential expression of hGIIA, hGIII and hGX sPLA2s in breast cancer cells is a consequence of various degrees of epigenetic silencing due to DNA hypermethylation and histone deacetylation.

Phospholipase A2 in experimental allergic bronchitis: a lesson from mouse and rat models.

BACKGROUND: Phospholipases A2 (PLA2) hydrolyzes phospholipids, initiating the production of inflammatory lipid mediators. We have previously shown that in rats, sPLA2 and cPLA2 play opposing roles in the pathophysiology of ovalbumin (OVA)-induced experimental allergic bronchitis (OVA-EAB), an asthma model: Upon disease induction sPLA2 expression and production of the broncho-constricting CysLTs are elevated, whereas cPLA2 expression and the broncho-dilating PGE2 production are suppressed. These were reversed upon disease amelioration by treatment with an sPLA2 inhibitor. However, studies in mice reported the involvement of both sPLA2 and cPLA2 in EAB induction. OBJECTIVES: To examine the relevance of mouse and rat models to understanding asthma pathophysiology. METHODS: OVA-EAB was induced in mice using the same methodology applied in rats. Disease and biochemical markers in mice were compared with those in rats. RESULTS: As in rats, EAB in mice was associated with increased mRNA of sPLA2, specifically sPLA2gX, in the lungs, and production of the broncho-constricting eicosanoids CysLTs, PGD2 and TBX2 in bronchoalveolar lavage (BAL). In contrast, EAB in mice was associated also with elevated cPLA2 mRNA and PGE2 production. Yet, treatment with an sPLA2 inhibitor ameliorated the EAB concomitantly with reverting the expression of both cPLA2 and sPLA2, and eicosanoid production. CONCLUSIONS: In both mice and rats sPLA2 is pivotal in OVA-induced EAB. Yet, amelioration of asthma markers in mouse models, and human tissues, was observed also upon cPLA2 inhibition. It is plausible that airway conditions, involving multiple cell types and organs, require the combined action of more than one, essential, PLA2s.

Regulation and function of epithelial secreted phospholipase A2 group X in asthma.

RATIONALE: Indirect airway hyperresponsiveness (AHR) is a fundamental feature of asthma that is manifest as exercise-induced bronchoconstriction (EIB). Secreted phospholipase A2 group X (sPLA2-X) plays a key role in regulating eicosanoid formation and the development of inflammation and AHR in murine models. OBJECTIVES: We sought to examine sPLA2-X in the airway epithelium and airway wall of patients with asthma, the relationship to AHR in humans, and the regulation and function of sPLA2-X within the epithelium. METHODS: We precisely phenotyped 34 patients with asthma (19 with and 15 without EIB) and 10 normal control subjects to examine in vivo differences in epithelial gene expression, quantitative morphometry of endobronchial biopsies, and levels of secreted protein. The regulation of sPLA2-X gene (PLA2G10) expression was examined in primary airway epithelial cell cultures. The function of epithelial sPLA2-X in eicosanoid formation was examined using PLA2 inhibitors and murine tracheal epithelial cells with Pla2g10 deletion. MEASUREMENTS AND MAIN RESULTS: We found that sPLA2-X protein is increased in the airways of patients with asthma and that epithelial-derived sPLA2-X may be increased in association with indirect AHR. The expression of sPLA2-X increases during in vitro epithelial differentiation; is regulated by inflammatory signals including tumor necrosis factor, IL-13, and IL-17; and is both secreted from the epithelium and directly participates in the release of arachidonic acid by epithelial cells. CONCLUSIONS: These data reveal a relationship between epithelial-derived sPLA2-X and indirect AHR in asthma and that sPLA2-X serves as an epithelial regulator of inflammatory eicosanoid formation. Therapies targeting epithelial sPLA2-X may be useful in asthma.

Group X secreted phospholipase A2 limits the development of atherosclerosis in LDL receptor-null mice.

OBJECTIVE: Several secreted phospholipases A2 (sPLA2s), including group IIA, III, V, and X, have been linked to the development of atherosclerosis, which led to the clinical testing of A-002 (varespladib), a broad sPLA2 inhibitor for the treatment of coronary artery disease. Group X sPLA2 (PLA2G10) has the most potent hydrolyzing activity toward phosphatidylcholine and is believed to play a proatherogenic role. METHODS AND RESULTS: Here, we show that Ldlr(-/-) mice reconstituted with bone marrow from mouse group X-deficient mice (Pla2g10(-/-)) unexpectedly display a doubling of plaque size compared with Pla2g10(+/+) chimeric mice. Macrophages of Pla2g10(-/-) mice are more susceptible to apoptosis in vitro, which is associated with a 4-fold increase of plaque necrotic core in vivo. In addition, chimeric Pla2g10(-/-) mice show exaggerated T lymphocyte (Th)1 immune response, associated with enhanced T-cell infiltration in atherosclerotic plaques. Interestingly, overexpression of human PLA2G10 in murine bone marrow cells leads to significant reduction of Th1 response and to 50% reduction of lesion size. CONCLUSIONS: PLA2G10 expression in bone marrow cells controls a proatherogenic Th1 response and limits the development of atherosclerosis. The results may provide an explanation for the recently reported inefficacy of A-002 (varespladib) to treat patients with coronary artery disease. Indeed, A-002 is a nonselective sPLA2 inhibitor that inhibits both proatherogenic (groups IIA and V) and antiatherogenic (group X) sPLA2s. Our results suggest that selective targeting of individual sPLA2 enzymes may be a better strategy to treat cardiovascular diseases.

Group X secretory PLA2 in neutrophils plays a pathogenic role in abdominal aortic aneurysms in mice.

Group X secretory PLA(2) (sPLA(2)-X) is expressed in neutrophils and plays a role in the pathogenesis of neutrophil-mediated tissue inflammation and injury. This study tested the hypothesis that sPLA(2)-X in neutrophils may contribute to the pathogenesis of abdominal aortic aneurysms (AAA) using sPLA(2)-X(-/-) mice. AAA was created by application of CaCl(2) to external surface of aorta. As a result, the aortas of sPLA(2)-X(-/-) mice had smaller diameters (percent increase from baseline; 24.8 ± 3.5% vs. 49.9 ± 9.1%, respectively; P < 0.01), a reduced grade of elastin degradation, and lower activities of elastase and gelatinase (26% and 19% lower, respectively) after CaCl(2) treatment compared with sPLA(2)-X(+/+) mice. In sPLA(2)-X(+/+) mice, immunofluorescence microscopic images showed that the immunoreactivity of sPLA(2)-X was detected only in neutrophils within aortic walls 3 days, 1, 2, and 6 wk after CaCl(2) treatment, whereas the immunoreactivity was not detected in macrophages or mast cells in aortic walls. sPLA(2)-X immunoreactivity also was colocalized in cells expressing matrix metalloproteinase (MMP)-9. Neutrophils isolated from sPLA(2)-X(-/-) mice had lower activities of elastase, gelatinase, and MMP-9 in response to stimuli compared with sPLA(2)-X(+/+) mice. The attenuated release of elastase and gelatinase from sPLA(2)-X(-/-) neutrophils was reversed by exogenous addition of mouse sPLA(2)-X protein. The adoptive transfer of sPLA(2)-X(+/+) neutrophils days 0 and 3 after CaCl(2) treatment reversed aortic diameters and elastin degradation grades in the lethally irradiated sPLA(2)-X(+/+) mice reconstituted with sPLA(2)-X(-/-) bone marrow to an extent similar to that seen in sPLA(2)-X(+/+) mice. In conclusion, sPLA(2)-X in neutrophils plays a pathogenic role in AAA in a mice model.

Blockade of human group X secreted phospholipase A2 (GX-sPLA2)-induced airway inflammation and hyperresponsiveness in a mouse asthma model by a selective GX-sPLA2 inhibitor.

Group X (GX) phospholipase A(2), a member of a large group of secreted phospholipases A(2) (sPLA(2)s), has recently been demonstrated to play an important in vivo role in the release of arachidonic acid and subsequent formation of eicosanoids. In a Th2 cytokine-driven mouse asthma model, deficiency of mouse GX (mGX)-sPLA(2) significantly impairs development of the asthma phenotype. In this study, we generated mGX-sPLA(2)(-/-) mice with knock-in of human GX (hGX)-sPLA(2) (i.e. hGX-sPLA(2)(+/+) knock-in mice) to understand more fully the role of GX-sPLA(2) in these allergic pulmonary responses and to assess the effect of pharmacological blockade of the GX-sPLA(2)-mediated responses. Knock-in of hGX-sPLA(2) in mGX-sPLA(2)(-/-) mice restored the allergen-induced airway infiltration by inflammatory cells, including eosinophils, goblet cell metaplasia, and hyperresponsiveness to methacholine in the mGX-sPLA(2)-deficient mice. This knock-in mouse model enabled the use of a highly potent indole-based inhibitor of hGX-sPLA(2), RO061606 (which is ineffective against mGX-sPLA(2)), to assess the potential utility of GX-sPLA(2) blockade as a therapeutic intervention in asthma. Delivery of RO061606 via mini-osmotic pumps enabled the maintenance in vivo in the mouse asthma model of plasma inhibitor concentrations near 10 µm, markedly higher than the IC(50) for inhibition of hGX-sPLA(2) in vitro. RO061606 significantly decreased allergen-induced airway inflammation, mucus hypersecretion, and hyperresponsiveness in the hGX-sPLA(2)(+/+) knock-in mouse. Thus, development of specific hGX-sPLA(2) inhibitors may provide a new pharmacological opportunity for the treatment of patients with asthma.

Group X secretory phospholipase A2 enhances TLR4 signaling in macrophages.

Secretory phospholipase A(2)s (sPLA(2)) hydrolyze glycerophospholipids to liberate lysophospholipids and free fatty acids. Although group X (GX) sPLA(2) is recognized as the most potent mammalian sPLA(2) in vitro, its precise physiological function(s) remains unclear. We recently reported that GX sPLA(2) suppresses activation of the liver X receptor in macrophages, resulting in reduced expression of liver X receptor-responsive genes including ATP-binding cassette transporters A1 (ABCA1) and G1 (ABCG1), and a consequent decrease in cellular cholesterol efflux and increase in cellular cholesterol content (Shridas et al. 2010. Arterioscler. Thromb. Vasc. Biol. 30: 2014-2021). In this study, we provide evidence that GX sPLA(2) modulates macrophage inflammatory responses by altering cellular cholesterol homeostasis. Transgenic expression or exogenous addition of GX sPLA(2) resulted in a significantly higher induction of TNF-α, IL-6, and cyclooxygenase-2 in J774 macrophage-like cells in response to LPS. This effect required GX sPLA(2) catalytic activity, and was abolished in macrophages that lack either TLR4 or MyD88. The hypersensitivity to LPS in cells overexpressing GX sPLA(2) was reversed when cellular free cholesterol was normalized using cyclodextrin. Consistent with results from gain-of-function studies, peritoneal macrophages from GX sPLA(2)-deficient mice exhibited a significantly dampened response to LPS. Plasma concentrations of inflammatory cytokines were significantly lower in GX sPLA(2)-deficient mice compared with wild-type mice after LPS administration. Thus, GX sPLA(2) amplifies signaling through TLR4 by a mechanism that is dependent on its catalytic activity. Our data indicate this effect is mediated through alterations in plasma membrane free cholesterol and lipid raft content.

Group X secretory phospholipase A2 regulates the expression of steroidogenic acute regulatory protein (StAR) in mouse adrenal glands.

We developed C57BL/6 mice with targeted deletion of group X secretory phospholipase A(2) (GX KO). These mice have approximately 80% higher plasma corticosterone concentrations compared with wild-type (WT) mice under both basal and adrenocorticotropic hormone (ACTH)-induced stress conditions. This increased corticosterone level was not associated with increased circulating ACTH or a defect in the hypothalamic-pituitary axis as evidenced by a normal response to dexamethasone challenge. Primary cultures of adrenal cells from GX KO mice exhibited significantly increased corticosteroid secretion compared with WT cells. Conversely, overexpression of GX secretory phospholipase A(2) (sPLA(2)), but not a catalytically inactive mutant form of GX sPLA(2), significantly reduced steroid production 30-40% in Y1 mouse adrenal cell line. This effect was reversed by the sPLA(2) inhibitor, indoxam. Silencing of endogenous M-type receptor expression did not restore steroid production in GX sPLA(2)-overexpressing Y1 cells, ruling out a role for this sPLA(2) receptor in this regulatory process. Expression of steroidogenic acute regulatory protein (StAR), the rate-limiting protein in corticosteroid production, was approximately 2-fold higher in adrenal glands of GX KO mice compared with WT mice, whereas StAR expression was suppressed in Y1 cells overexpressing GX sPLA(2). Results from StAR-promoter luciferase reporter gene assays indicated that GX sPLA(2) antagonizes StAR promoter activity and liver X receptor-mediated StAR promoter activation. In summary, GX sPLA(2) is expressed in mouse adrenal glands and functions to negatively regulate corticosteroid synthesis, most likely by negatively regulating StAR expression.

Pioglitazone suppresses the lipopolysaccharide-induced production of inflammatory factors in mouse macrophages by inactivating NF-kappaB.

TZDs (thiazolidinediones) are prescribed as anti-Type II diabetes drugs, but little is known regarding whether TZDs regulate the expression of sPLA2 (secretory phospholipase A2) in macrophages. We have investigated the effects of pioglitazone on LPS (lipopolysaccharide)-induced production of TNF-alpha (tumour necrosis factor alpha), sPLA2-V and -X (groups V and X sPLA2) in RAW 264.7 macrophages. TNF-alpha, sPLA2-V and -X mRNA and protein expression were determined by RT-PCR (reverse transcriptase-PCR) and Western blot analysis, respectively. The activity of NF-kappaB (nuclear factor kappaB) was determined by Western blot and confocal microscopy. LPS induced TNF-alpha, sPLA2-V and sPLA2-X mRNA and protein expression. Pretreatment with 10 mumol/l pioglitazone significantly suppressed LPS-induced TNF-alpha, sPLA2-V and sPLA2-X mRNA and protein expression. LPS induced NF-kappaB expression and translocation in the nucleus, but the inductive effects were inhibited by pioglitazone. Our findings indicate that pioglitazone inhibits production of inflammatory factors induced by LPS in murine macrophage cells by inactivating NF-kappaB. Pioglitazone appears to play an anti-inflammatory role in the atherosclerotic process.

Differential expression of secretory phospholipases A2 in normal and malignant prostate cell lines: regulation by cytokines, cell signaling pathways, and epigenetic mechanisms.

Upregulation of group IIA phospholipase A(2) (sPLA(2)-IIA) correlates with prostate tumor progression suggesting prooncogenic properties of this protein. Here, we report data on expression of three different sPLA(2) isozymes (groups IIA, V, and X) in normal (PrEC) and malignant (DU-145, PC-3, and LNCaP) human prostate cell lines. All studied cell lines constitutively expressed sPLA(2)-X, whereas sPLA(2)-V transcripts were identified only in malignant cells. In contrast, no expression of sPLA(2)-IIA was found in PrEC and DU-145 cells, but it was constitutively expressed by IFN-gamma in LNCaP and PC-3 cells. Expression of sPLA(2)-IIA is upregulated in PC-3 and in PrEC cell in a signal transducer and activator of transcription-1-dependent manner, but not in LNCaP cell. Additional signaling pathways regulating sPLA(2)-IIA expression include cAMP/protein kinase A, p38 mitogen-activated protein kinase, protein kinase C, Rho-kinase, and mitogen-activated/extracellular response protein kinase / extracellular signal-regulated kinase. No deletions were revealed in the sPLA(2)-IIA gene from DU-145 cells lacking the expression of sPLA(2)-IIA. Reexpression of sPLA(2)-IIA was induced by 5-aza-2'-deoxycytidine demonstrating that DNA methylation is implicated in the regulation of sPLA(2)-II. Together, these data suggest that sPLA(2)-IIA and sPLA(2)-V, but not sPLA(2)-X, are differentially expressed in normal and malignant prostate cells under the control of proinflammatory cytokines; epigenetic mechanisms appear involved in the regulation of sPLA(2)-IIA expression, at least in DU-145 cells.