Regulation Of Prostaglandin E2 Synthesis Research Articles

Epigenetic Regulation of Prostaglandin E2 Synthesis Alters Macrophage Function to Promote Inflammation and Impair Diabetic Wound Healing

Davis, Frank M. MD; Joshi, Amrita PhD; Kimball, Andrew S. MD; denDekker, Aaron PhD; Gallagher, Katherine MD Author Information

Abstract LB-250: Establishment of a positive feedback between PGE2 and COX2 diverts DC differentiation into MDSCs and sustains their immunosuppressive functions in ovarian cancer

Abstract Myeloid-derived suppressor cells (MDSCs) are essential players in cancer-associated immune dysfunction. We report that the differentiation and functional stability of human monocytic MDSCs is driven by a single cancer-associated inflammatory mediator, prostaglandin E2 (PGE2), and depends on the establishment of a positive feedback between PGE2 and the key regulator of PGE2 synthesis, COX-2. PGE2 abrogates the differentiation of CD1a+ DC from monocytes, redirecting it towards CD14+CD33+CD34+ cells expressing all MDSC-associated suppressive factors: IDO, arginase, NOS2 and IL-10, and inducing their CTL-suppressive functions. Moreover, PGE2 promotes the expression of endogenous COX2 and autocrine secretion of PGE2 in MDSCs, establishing a positive feedback loop that stabilizes the MDSC phenotype. Undisturbed COX2 activity and persistent production of PGE2 by MDSCs, isolated from cancer patients, was required for their continued expression of CXCR4 and responsiveness to CXCL12. Moreover, PGE2 also proved to be the key factor promoting the production of CXCL12 in cancer microenvironment, a chemokine exerting multiple tumor-promoting effects either directly by acting on cancer cells or indirectly by inducing angiogenesis and recruiting other immunosuppressive cells, i.e. Tregs and pDCs. In accordance with the physiological role of PGE2 in the development and functional stability of human cancer -associated MDSCs, we observed that the frequencies of CD11b+ CD33+ MDSCs in the ovarian cancer (OvCa) ascites closely correlate with the local production of PGE2 and expression of COX2, while the OvCa ascites environment can promote the induction of MDSCs from differentiating monocytes in a COX2-dependent manner. Importantly. disruption of COX2-PGE2 feedback in fully-developed MDSCs isolated from cancer patients, using COX2 inhibitors or inhibitors of EP2/EP4-dependent PGE2 signaling, eliminates the production of all other suppressive factors and terminates the CTL-suppressive function of MDSCs as well as eliminates both the expression of CXCR4 on MDSCs and the production of CXCL12, highlighting the key role of PGE2 in the tumor-accumulation of immunosuppressive cells. The currently demonstrated key role of the positive COX2-PGE2 feedback loop in the differentiation and functional stability of MDSCs facilitates the targeting of MDSCs in cancer immunotherapy and facilitates the development of additional MDSC-targeting therapies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-250. doi:10.1158/1538-7445.AM2011-LB-250

The effect of the pro-inflammatory cytokine tumor necrosis factor-alpha on human joint capsule myofibroblasts

IntroductionPrevious studies have shown that the number of myoblastically differentiated fibroblasts known as myofibroblasts (MFs) is significantly increased in stiff joint capsules, indicating their crucial role in the pathogenesis of post-traumatic joint stiffness. Although the mode of MFs' function has been well defined for different diseases associated with tissue fibrosis, the underlying mechanisms of their regulation in the pathogenesis of post-traumatic joint capsule contracture are largely unknown.MethodsIn this study, we examined the impact of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α) on cellular functions of human joint capsule MFs. MFs were challenged with different concentrations of TNF-α with or without both its specifically inactivating antibody infliximab (IFX) and cyclooxygenase-2 (COX2) inhibitor diclofenac. Cell proliferation, gene expression of both alpha-smooth muscle actin (α-SMA) and collagen type I, the synthesis of prostaglandin derivates E2, F1A, and F2A, as well as the ability to contract the extracellular matrix were assayed in monolayers and in a three-dimensional collagen gel contraction model. The α-SMA and COX2 protein expressions were evaluated by immunofluorescence staining and Western blot analysis.ResultsThe results indicate that TNF-α promotes cell viability and proliferation of MFs, but significantly inhibits the contraction of the extracellular matrix in a dose-dependent manner. This effect was associated with downregulation of α-SMA and collagen type I by TNF-α application. Furthermore, we found a significant time-dependent upregulation of prostaglandin E2 synthesis upon TNF-α treatment. The effect of TNF-α on COX2-positive MFs could be specifically prevented by IFX and partially reduced by the COX2 inhibitor diclofenac.ConclusionsOur results provide evidence that TNF-α specifically modulates the function of MFs through regulation of prostaglandin E2 synthesis and therefore may play a crucial role in the pathogenesis of joint capsule contractures.

Regulation of prostaglandin E2 synthesis in cells derived from chondrocytes of patients with osteoarthritis

BackgroundOsteoarthritis (OA) is a disorder that causes pain and degeneration of the joint over a chronic time course. Chondrocytes in OA play important roles in maintaining the homeostasis of the joint while they produce many cytokines and pathological mediators, including interleukin-1 P (IL-1β), cyclooxygenases (COX), and prostaglandin E2 (PGE2). To elucidate the mechanisms of pain due to OA, the pathway of PGE2 synthesis was analyzed using cells derived from chondrocytes obtained from patients with OA. MethodsChondrocytes were isolated from cartilage samples obtained at the time of joint replacement surgery from patients with OA. The chondrocytes at the second passage were cultured with or without IL-1β, dexamethasone (DEX), or COX inhibitors such as NS-398, meloxicam, and indomethacin. Reverse transcription-polymerase chain reaction and Western blotting analysis were performed to study the levels of mRNA and protein, respectively. An enzyme-linked immunosorbent assay was performed to investigate the translocation of nuclear factor-KB (NF-ΚB) to the nucleus, and Western blotting analysis was performed to study the phosphorylation of mitogen-activated protein kinases. ResultsIL-1β markedly enhanced the expression of COX-2 and microsomal prostaglandin E synthase-1 (mPGES-1) at both the mRNA and protein levels. The up-regulation was suppressed by DEX or COX inhibitors. IL-1β strongly increased the translocation of NF-ΚB to the nucleus and the phosphorylation of extracellular-signal-regulated kinase, p38, and c-Jun amino-terminal kinase; but the up-regulation was not inhibited by DEX or COX inhibitors. Interestingly, in a dose-dependent manner, PGE2 recovered mPGES-i expression from suppression by DEX, whereas it did not restore the expression of COX-2 in the presence of DEX and IL-1β. ConclusionsThese results suggested that in cells derived from OA chondrocytes different mechanisms of regulation exist between mPGES-1 and COX-2, and the expression of mPGES-1 was, at least partially, regulated through the autocrine positive feedback by PGE2.

Regulation of prostaglandin E2 synthesis after brain irradiation

A local tissue reaction, termed neuroinflammation, occurs after irradiation of brain tissue. Previous work suggested that cyclooxygenase (COX)-2 activity was important for changes in gene expression associated with neuroinflammation as well as increased prostaglandin E2 (PGE2) levels seen after radiation treatment. To begin to determine the contributions of other enzymes involved in PGE2 production, we examined protein levels of COX-1 and COX-2 as well as 2 PGE synthases (membrane and cytosolic PGES) 4 h after 35 Gy single dose irradiation to the brains of C3HeN mice. We also evaluated the effects of specific COX inhibitors on PGE2 production and PGES expression. As expected, COX-2 expression increased after radiation exposure. Brain irradiation also increased tissue protein levels for both PGES isoforms. Specific COX-2 inhibition with NS398 lowered brain PGE2 levels by about 60%. Surprisingly, COX-1 inhibition with SC560 completely prevented the elevation of PGE2 seen after irradiation. Interestingly, NS398 reduced the membrane-associated PGES isoform, whereas SC560 treatment lowered cytosolic isoform levels below those seen in unirradiated controls. Taken together, these data indicate that both cyclooxygenases contribute to PGE2 production in irradiated brain and reveal dependence of PGES isoforms expression on specific cyclooxygenase activities.

Opposite regulation of prostaglandin E2 synthesis by transforming growth factor-β1 and interleukin 10 in activated microglial cultures

We have recently shown that prostaglandin E2 (PGE2) synthesis in activated microglia is tightly regulated by several substances (NO, neurotransmitters, pro-inflammatory cytokines), that might originate from intrinsic brain cells or from hematogenous cells infiltrating the brain in the course of inflammatory diseases. In view of the important immunoregulatory and neuroprotective functions recently attributed to PGE2, in the present study we extended our analysis of factors regulating PGE2 synthesis in rat microglial cultures to two anti-inflammatory and immunosuppressive cytokines, transforming growth factor β1 (TGF-β1) and interleukin 10 (IL-10), which share with PGE2 the ability to strongly deactivate peripheral macrophages and microglial cells. Moreover, we looked at the effect of the two cytokines on nitric oxide (NO) synthesis, another important microglial effector, whose synthesis is linked to that of PGE2 by complex feed-back mechanisms. We found that while both cytokines inhibited LPS-induced NO release, they had distinct and opposite regulatory activities on PGE2 production. In fact, while TGF-β1 enhanced LPS-induced PGE2 synthesis, IL-10 showed an inhibitory effect. The two cytokines acted mainly by regulating the LPS-induced expression of the rate limiting enzymes of the two metabolic pathways, cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS). Moreover, TGF-β1 counteracted the effect of the pro-inflammatory cytokine interferon-γ, which in the same cultures has been shown to downregulate PGE2 and to upregulate NO synthesis. Although the present in vitro observations cannot be directly extrapolated to the in vivo situation, they may provide a novel clue for understanding the specific role of TGF-β1 and IL-10 in several neurological diseases such as multiple sclerosis, in which their cerebral level was found to be elevated.

Regulation of prostaglandin E2 synthesis by cyclooxygenase induction in osteoblast MC3T3-E1

Regulation of prostaglandin E2 synthesis by cyclooxygenase induction in osteoblast MC3T3-E1

Protein kinase C and chondrocyte activation.

We have begun to examine the role of protein kinase C (PKC) in chondrocyte activation by interleukin-1 (IL-1) in search of a possible signal transduction mechanism. Untreated chondrocytes synthesised little or no collagenase or gelatinase and only modest amounts of prostaglandin E2 (PGE2), while IL-1 induced considerable amounts of these. An activator of PKC, phorbol myristate acetate (PMA), alone did not influence the synthesis of the metalloproteinases to any great degree, but enhanced PGE2 production. Sphingosine and staurosporin, inhibitors of PKC, each eliminated the synergistic effect of PMA upon enzyme induction by high doses (10 U/ml) of IL-1, but failed to influence enzyme induction by this dose of IL-1 alone. However, a low dose (1 U/ml) of IL-1 in combination with these inhibitors was synergistic upon enzyme induction. Although these inhibitors reduced the synthesis of PGE2 in response to PMA, synthesis of PGE2 in response to both doses of IL-1 was greatly enhanced by the inhibitors. PMA, but not IL-1, enhanced the phosphorylation of an 80 K protein which is characteristic of PKC activity in certain types of cells. From these data, we conclude that PKC is unlikely to be involved in the induction of neutral metalloproteinases by IL-1, although once induction has occurred, PKC may modulate this effect. PKC may also act as regulator of PGE2 synthesis, although this requires further investigation.

Regulation of prostaglandin E2 synthesis by angiotensin II, potassium, osmolality, and dexamethasone

We investigated the regulation of cellular prostaglandin E2 (PGE2) biosynthesis in rabbit renomedullary interstitial cells in tissue culture. Arachidonic acid markedly stimulated PGE2 biosynthesis by these cells. Repeated exposure to arachidonic acid, resulted in progressively less stimulation of PGE2 biosynthesis. Potassium and dexamethasone diminished PGE2 biosynthesis by decreasing the rate of arachidonic acid release from the endogenous arachidonic acid storage pool. Hyperosmolality, like angiotensin II, bradykinin, and arginine vasopressin, stimulated PGE2 biosynthesis by increasing the rate of arachidonic acid release. Inhibitors of protein synthesis diminished angiotensin II-, bradykinin-, and arginine-vasopressin-stimulated PGE2 biosynthesis by decreasing hormone-stimulated arachidonic acid release. The effects of potassium, dexamethasone, arachidonic acid, and hyperosmolality on PGE2 biosynthesis were unaffected by protein synthesis inhibitors. Hormone-stimulated phospholipase activation is dependent on protein synthesis, whereas potassium, hyperosmolality, and dexamethasone alter the release of arachidonic acid from cellular lipids via a mechanism that is independent of protein synthesis.