Role of dihydroartemisinin in regulating prostaglandin E2 synthesis cascade and inflammation in endothelial cells.

Endothelial cells (ECs) are crucial in maintaining vascular homeostasis. Endothelial dysfunction was involved in many cardiovascular diseases (CVDs). Recently, antimalarial medicine artemisinin and its derivatives including dihydroartemisinin (DHA) were found to be beneficial in some diseases including CVDs. Prostaglandin (PG) E2 is a known inflammatory mediator and plays important roles in cardiovascular system. This study was to investigate the role of DHA in regulating cyclooxygenase (COX)/PGE synthase (PGES)/PGE2 cascade and inflammation in ECs. After DHA treatment, the mRNA and protein levels of COX-2 were strikingly upregulated in time- and dose-dependent manners. In contrast, COX-1 was significantly downregulated. As expected, inhibition of COX-1 or COX-2 further reduced PGE2 production after DHA treatment. Moreover, DHA enhanced microsomal PGE2 synthase (mPGES)-2 and moderately modulated cytosolic PGE2 synthase (cPGES) with no effect on mPGES-1 expression. Importantly, DHA significantly reduced PGE2 levels in line with the upregulation of 15-hydroxyprostaglandin dehydrogenase (15-PGDH, a key enzyme for prostaglandin degradation). Lastly, we observed that DHA not only reduced the PGE2 levels in tumor necrosis factor-α (TNF-α)-treated ECs but also blunted the upregulation of inflammatory cytokines of interleukin (IL)-6 and IL-1ß induced by TNF-α or PGE2. These findings demonstrated an important role of DHA in regulating PGE2 synthesis cascade and inflammation in ECs, suggesting a potential of DHA for the treatment of inflammatory vascular diseases.

Loss of luteotropic prostaglandin E plays an important role in the regulation of luteolysis in women.

STUDY QUESTION: Do intraluteal prostaglandins (PG) contribute to luteal regulation in women? SUMMARY ANSWER: Prostaglandin E (PGE), which is produced in human granulosa-lutein cells stimulated with luteotropic hCG, exerts similar luteotropic effects to hCG, and the expression of PG synthetic and metabolic enzymes in the human CL is driven toward less PGE but more prostaglandin F (PGF) during luteolysis. WHAT IS KNOWN ALREADY: Uterine PGF is a major luteolysin in many non-primate species but not in women. Increases in the PGF synthase, aldo-ketoreductase family one member C3 (AKR1C3), have been observed in the CL of marmoset monkeys during luteolysis. PGE prevents spontaneous or induced luteolysis in domestic animals. STUDY DESIGN, SIZE, DURATION: Human CL tissues staged as the early-luteal (n = 6), mid-luteal (n = 6), late-luteal (n = 5) and menstrual (n = 3) phases were obtained at the time of hysterectomy for benign gynecological conditions. Luteinized granulosa cells (LGCs) were purified from follicular fluids obtained from patients undergoing assisted conception. PARTICIPANTS/MATERIALS, SETTING, METHODS: Upon collection, one half of the CL was snap-frozen and the other was fixed with formalin and processed for immunohistochemical analysis of a PGE synthase (PTGES). Quantitative RT-PCR was employed to examine changes in the mRNA abundance of PG synthetic and metabolic enzymes, steroidogenic enzymes, and luteolytic molecules in the staged human CL and in human LGCs in vitro treated with hCG, PGE and PGF. A PGE withdrawal experiment was also conducted in order to reveal the effects of the loss of PGE in LGCs. Progesterone concentrations in the culture medium were measured. MAIN RESULTS AND THE ROLE OF CHANCE: The key enzyme for PGE synthesis, PTGES mRNA was abundant in the functional CL during the mid-luteal phase (P < 0.01), while mRNA abundance for genes involved in PGF synthesis (AKR1B1 and AKR1C1-3) increased in the CL during the late-luteal phase and menstruation (P < 0.05-0.001). PTGES mRNA expression positively correlated with that of 3ß-hydroxysteroid dehydrogenase (HSD3B1; r = 0.7836, P < 0.001), while AKR1C3 expression inversely correlated with that of HSD3B1 (r = -0.7514, P = 0.0012) and PTGES (r = -0.6923, P = 0.0042). PGE exerted similar effects to hCG-promoting genes, such as steroidogenic acute regulatory protein (STAR) and HSD3B1, to produce progesterone and luteotropic PGE, suppress PGF synthetic enzymes and down-regulate luteolytic molecules such as ßA- and ßB-inhibin subunits (INHBA and INHBB) and bone morphogenetic proteins (BMP2, BMP4 and BMP6). PGE withdrawal resulted in reductions in the enzymes that produce progesterone (STAR; P < 0.001) and PGE (PTGES; P < 0.001), and the capacity to produce PGE decreased, while the capacity to produce PGF increased during the culture. The addition of PGF did not recapitulate the luteolytic effects of PGE withdrawal. LARGE SCALE DATA: None. LIMITATIONS, REASONS FOR CAUTION: Changes in mRNA expression of PG synthetic and metabolic enzymes may not represent actual increases in PGF during luteolysis in the CL. The effects of PGF on luteal cells currently remain unclear and the mechanisms responsible for decreases in the synthesis of PGE in vitro and at luteolysis have not been elucidated in detail. WIDER IMPLICATIONS OF THE FINDINGS: The results obtained strongly support a luteotropic function of PGE in regulation of the human CL. They suggest that the main PG produced in human luteal tissue changes from PGE to PGF during the maturation and regression of the CL, and the loss of PGE is more important than the effects of PGF during luteolysis in women. This may be accompanied by reduced effects of LH/hCG in luteal cells, particularly decreased activation of cAMP/protein kinase A; however, the underlying mechanisms remain unknown. STUDY FUNDING AND COMPETING INTEREST(S): This study was supported by the Cunningham Trust to WCD, a Postdoctoral Fellowship for Research Abroad from the Japan Society for the Promotion of Science and the Suntory Foundation for Life Sciences to J.N.-K.; W.C.D. is supported by an MRC Centre Grant G1002033 and a Scottish Senior Clinical Fellowship. The authors have nothing to disclose.

Paederia foetida Linn. inhibits adjuvant induced arthritis by suppression of PGE(2) and COX-2 expression via nuclear factor-κB.

The current investigation was undertaken to determine the anti-inflammatory and antioxidant effects of Paederia foetida Linn. (PF) along with its mechanism of action when implemented in tissue protection. HPTLC was used in the identification of the compound quercetin, while in vitro analysis confirmed the significance of the antioxidant and anti-inflammatory action of PF. We initially demonstrated the in vivo anti-inflammatory effect of PF, evaluating it against a variety of phlogistic agents as well as turpentine oil, prostaglandin and arachidonic acid. Groups of rats, fasted overnight, were treated as follows: Group I: normal control (vehicle), Group II: PF (100 mg kg(-1)), Group III: arthritic control (CFA only, 0.05 ml), Group IV, V, VI: CFA (0.05 ml) + PF (25, 50 and 100 mg kg(-1)) and Group VII: CFA (0.05 ml) + indomethacin (10 mg per kg b.w.). PF significantly protected against paw edema, arthritic index and body weight alteration induced by Complete Fruend's Adjuvant (CFA). Other observations, like histological and macroscopic changes, were observed in CFA induced inflammation in knee joints. Subcutaneous administration of CFA was accompanied by proinflammatory cytokine status, as appraised by the amplification of interleukin-2 (IL-2), interleukin-1ß (IL-1ß) and tumor necrosis factor (TNF-α); oxidative stress status was estimated by the enhancement of the level of lipid peroxidation (LPO) and the depletion of superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione (GSH). Pre-treatment with PF significantly (P < 0.001) protected against CFA induced oxidative stress and proinflammatory cytokines. More prominently, CFA administration augmented tissue and plasma superoxide (O2) and hydrogen peroxide (H2O2) levels, while the PF pre-treatment significantly (P < 0.001) reversed all CFA induced intracellular interruption. Following CFA induced arthritis, PF was tested for its free radical scavenging activity against the DPPH and ABTS radicals and its inhibitory proficiency against COX-1 and COX-2 in vitro. Considering the above, the current research confirmed momentous protection against CFA induced arthritis, which could be attributed to its anti-inflammatory and pro-oxidant nature.

Estrogen receptor-related receptor α regulation by interleukin-1ß in prostaglandin E(2)- and cAMP-dependent pathways in osteoarthritic chondrocytes.

OBJECTIVE: We reported previously that the orphan nuclear receptor, estrogen receptor-related receptor α (ERRα), is expressed in articular chondrocytes and is dysregulated in a mouse model of inflammatory arthritis. The aim of this study, therefore, was to determine whether ERRα is also dysregulated in patients with osteoarthritis (OA). METHODS: ERRα messenger RNA (mRNA) and protein were quantified in normal and OA cartilage samples and in OA chondrocytes in vitro, with and without short-term treatment with a variety of OA-associated factors and signaling pathway agonists and inhibitors. RESULTS: ERRα expression was lower in OA than in normal articular cartilage. Interleukin-1ß (IL-1ß) markedly up-regulated ERRα expression in OA chondrocytes in vitro, and agonist or inhibitor treatment indicated that the up-regulation was dependent on cyclooxygenase 2 (COX-2; NS398), prostaglandin E(2), cAMP (8-bromo-cAMP), and protein kinase A (PKA; KT5720). Treatment with the ERRα inverse agonist XCT790 decreased the expression of SOX9 and the up-regulation of ERRα by IL-1ß, suggesting autoregulation of ERRα in the IL-1ß pathway. Matrix metalloproteinase 13 (MMP-13) expression was also decreased by treatment with XCT790 plus IL-1ß versus IL-1ß alone, and the down-regulation of MMP-13 mRNA and protein observed with XCT790 alone suggests that the up-regulation of MMP-13 by IL-1ß is ERRα-dependent. CONCLUSION: We report the first evidence that ERRα expression is regulated by IL-1ß in COX-2-, cAMP-, and PKA-dependent pathways in OA chondrocytes. We confirmed that SOX9 is an ERRα target gene in human, as in rodent, chondrocytes and identified MMP-13 as a potential new target gene, which suggests that ERRα may both respond to the healing signal and contribute to extracellular degradation in OA cartilage.

Regulation of prostaglandin synthesis in ovaries of sexually-mature zebrafish (Danio rerio).

This study investigates the regulation of prostaglandin (PG) synthesis in the ovaries of sexually-mature zebrafish (Danio rerio). We examined the ovarian expression of genes within the arachidonic acid (AA) pathway, and the ovarian levels of 17alpha,20beta-dihydroxy-4-pregnen-3-one (17alpha,20beta-P), 17beta-estradiol (E(2)), and PGF(2alpha) in spawning and nonspawning fish during the ovulatory cycle. Real-time RT-PCR analysis revealed that the expression levels of cytosolic phospholipase A(2) (cpla2) and cyclooxygenases (COX)-2 (ptgs2) in ovarian fragments and in isolated full-grown follicles of spawning fish were highest at 6:00 when ovulation was expected to occur. In nonspawning fish, cpla2 expression levels declined over time while ptgs2 expression displayed the same temporal pattern as in spawning fish. Elevated levels of 17alpha,20beta-P in the spawning fish occurred at 3:30, but there were no changes in the nonspawning fish. In other studies conducted to investigate the hormonal regulation of AA pathway genes, fish exposed via the water for 24 or 96 hr to 17alpha,20beta-P or E(2) exhibited reduced ovarian expression levels of COX-1 (ptgs1) and PG E synthase-2 (ptgsl), and E(2) reduced the expression of cpla2. Injection of human chorionic gonadotropin (hCG) (100 IU) led to increased expression levels of cpla2 and ptgs2 at 2 and 18 hr post-treatment, but consistently reduced ptgs1 and ptgsl expression. In these fish, ovarian levels of 17alpha,20beta-P were elevated at all time points and PGF(2alpha) levels in the hCG-treated group were significantly higher than the control fish at 18 hr. Collectively, these in vivo results suggest that gonadotropins and steroids are involved in the regulation of the AA pathway in ovarian follicles of zebrafish.

Mapping of the genes encoding mouse prostaglandin D, E, and F and prostacyclin receptors.

Prostaglandins and prostacyclin are metabolites of arachidonic acid and exert a variety of actions to maintain local homeostasis in the body. Their actions are mediated by cell surface receptors specific to the respective ligands. Using a panel of interspecific back-cross mice, we have mapped the prostaglandin D receptor gene (Ptgdr), prostaglandin E receptor subtype EP(1) gene (Ptgerepl), prostaglandin F receptor gene (Ptgfr), and prostacyclin receptor gene (Ptgir). Ptgdr mapped to proximal Chr 14, Ptgfr mapped to distal Chr 3, Ptgerepl mapped to middle Chr 8, and Ptgir mapped to proximal Chr 7.

Genetically determined alcohol preference and cyclic AMP binding proteins in mouse brain.

Free-choice consumption of alcohol by mice with differing phenotypic alcohol preferences caused uniformly large decreases in brain cyclic AMP-dependent protein kinase activity toward an exogenous substrate (histone 2b) but the effect of alcohol on brain cyclic AMP binding activity was strain-specific. Furthermore, particulate kinase phosphorylating activity toward an endogenous protein (kinase regulatory subunit, RII) was altered by alcohol consumption in a strain-specific manner. The changes in cyclic AMP binding and phosphorylating activity appeared to result from phenotypic differences in the brain's response to alcohol. Thus, low preference animals were sensitive to alcohol and showed a large decrease in cyclic AMP binding and an increase in phosphorylation of regulatory subunit in response to alcohol. In contrast, high preference strain had only a small decrease in cyclic AMP binding and a decrease in phosphorylation, even though these animals consumed a significantly larger dose of alcohol. These data suggest that changes in cyclic AMP binding and/or phosphorylation of kinase regulatory subunit may be phenotypic markers of alcohol preference in inbred mice.